用于治疗癌症的新型免疫细胞因子

技术领域

本发明涉及新型免疫细胞因子及其在治疗癌症中的用途。

背景技术

尽管针对各种类型的血液系统恶性肿瘤和一些实体瘤(例如转移性睾丸癌)的治疗已经取得成功，但是大多数播散性实体癌症仍然无法治愈。常规癌症治疗方法的治疗功效通常受到有机小分子无法在疾病部位累积足够的量的限制(例如参见van der Veldt AA等人，Clin Cancer Res.19：4163–4173,2013)。

现在开发了新的策略，其优先激活相关的免疫亚群，例如效应T细胞、单核细胞和NK细胞，同时限制调节性T细胞的激活。但是，显著的副作用和不利的药代动力学特性已经成为阻碍治疗相关剂量施用的主要缺点。尤其是，细胞因子免疫疗法通常会导致产生严重的剂量限制副作用(Pachella等人，Pract Oncol 6：212–221,2015)。大多数细胞因子共有的两个特性被认为在与治疗相关的不良反应的发生中起着至关重要的作用。首先，细胞因子具有多效性，意味着其可以影响多于一种的细胞类型。此外，细胞因子具有短的血清半衰期，因此需要以高剂量施用以达到其治疗效果。在有效提高治疗功效的同时，高剂量会加剧多效活性，在患者中显现为不良反应。

一种旨在提高功效的方法试图通过将细胞因子遗传融合到抗体或抗体成分(例如单链可变片段(scFv))上，以将细胞因子递送到肿瘤部位。这样的融合蛋白称为免疫细胞因子，结合了抗体的结合特异性与细胞因子(例如IL-2)的效力(Sondel与Gillies,Antibodies 1：149-171,2012；Skrombolas与Frelinger,Expert Rev Clin Immunol.10(2)：207–217,2014；Kiefer与Neri,Immunol Rev.270(1)：178–192,2016)。通过使用免疫细胞因子改善了细胞因子向肿瘤部位的递送，尤其是对于具有易于接近的肿瘤的癌症。在另一个例子中，免疫细胞因子包含细胞因子(IL-12)，其通过MMP9裂解位点与特异性抑制性抗IL-12scFv连接(Skrombolas等人,JInterferon Cytokine Res.39(4)：233-245,2019)。但是，播散性全身性疾病的治疗可能受益于针对肿瘤靶向和在肿瘤部位的激活而优化的免疫细胞因子(Sondel和Gillies,Antibodies 1：149-171,2012)。特别地，抗体在肿瘤之外的结合可能导致不需要的细胞因子活性和潜在的副作用。由于已经报道某些有效载荷完全消除了癌症小鼠模型中亲本抗体的肿瘤靶向潜力，因此这个问题变得更加关键(参见例如Hess,博士学位论文,ETH Zürich,2015)。

因此，仍然需要一种免疫细胞因子，其能够安全有效地将细胞因子递送至肿瘤部位并激活细胞因子。

附图说明

图1：用于产生免疫细胞因子(ICC)的融合位点。

图2：去糖基化RP-LC分离后获得的c9G4PVGLIG-IL15的去卷积MS质谱图。

图3：去糖基化、IdEs消化和RP-LC分离后获得的Fc/2cG4PVGLIG-IL15的去卷积MS质谱图。

图4：与mAb重链C-末端融合时，MMP-9/2连接子的可裂解性评估。报道为不能被MMP-9/2裂解的GIVGPL连接子被用作裂解特异性的阴性对照。

图5：与mAb重链N-末端融合时，MMP-9/2连接子的可裂解性评估。报道为不能被MMP-9/2裂解的GIVGPL连接子被用作裂解特异性的阴性对照。HC：重链，LC：轻链，Ck：细胞因子。

图6：通过人和鼠MMP-9和MMP-2(HC C-末端融合，PVGLIG连接子)评估基于c9G4的免疫细胞因子以及H16/L16-IL15和HHS76-IL15免疫细胞因子的可裂解性。(A)c9G4-Il15、H16/L16-IL15及HHS76-IL15；(B)c9G4-CCL4及c9G4-IFNa。HC：重链，LC：轻链，Ck：细胞因子。

图7：MMP-9/2连接子可裂解性评估的总结。

图8：PVGLIG和GIVGPL连接子在存在MMP-9活性时，在缓冲液(50mM Tris pH 7.5、150mM NaCl、20mM CaCl

图9：小鼠血清中ICC裂解的分析：免疫沉淀，还原和反相分离后获得的抗

图10：IL15诱导IL2Rβ和IL2Rγ受体亚基的二聚化。来自三个独立实验的代表性数据。

图11：血浆样品(RENCA移植小鼠)的蛋白质印迹分析。

图12：血浆样品蛋白质印迹的光密度分析。X表示该样品缺失。

图13：循环ICC(血浆样品)(RENCA移植小鼠)的统计分析。

图14：肿瘤样品(RENCA移植小鼠)的蛋白质印迹分析。

图15：肿瘤样品蛋白质印迹的光密度分析。X表示该样品缺失。

图16：寻址至(addressed to)RENCA肿瘤的ICC的统计分析。

图17：RENCA移植小鼠血浆与肿瘤中ICC-PVGLIG表现的统计分析。

图18：去糖基化RP-LC分离后获得的NHS67-PVGLIG-IL15的去卷积MS质谱图。

图19：去糖基化、IdEs消化和RP-LC分离后获得的Fc/2NHS67-PVGLIG-IL15的去卷积MS质谱图。

图20：去糖基化RP-LC分离后获得的H16L16-PVGLIG-IL15的去卷积MS质谱图。

图21：去糖基化、IdEs消化和RP-LC分离后获得的Fc/2H16L16-PVGLIG-IL15的去卷积MS质谱图。

图22：纯化的c9G4-PVGLIG-hIL15、NHS76-PVGLIG-hIL15和H16L16-PVGLIG-hIL15ICC在非还原/加热(NRH)和还原/加热(RH)条件下的SDS-PAGE分析。

图23：小鼠T细胞的ICC激活与对照相比。在裂解和未裂解的NHS76-PVGLIG-IL15或对照存在时，通过T细胞CD69(A)或CD25(B)表达情况，以及在裂解和未裂解的H16L16-PVGLIG-IL15或对照存在时，通过T细胞CD69(C)或CD25(D)表达情况，来测量激活情况。

图24：人T细胞的ICC激活与对照相比。在裂解和未裂解的NHS76-PVGLIG-IL15或对照存在时，通过T细胞CD69(A)或CD25(B)表达情况，以及在裂解和未裂解的H16L16-PVGLIG-IL15或对照存在时，通过T细胞CD69(C)或CD25(D)表达情况，来测量激活情况。

图25：人T细胞的ICC激活与对照相比。在裂解和未裂解的NHS76-PVGLIG-IL15或两个不同供体(供体1(A)和供体2(B))的对照存在时，通过T细胞的INFγ分泌，来测量激活情况。上图：在裂解和未裂解的H16L16-PVGLIG-IL15或两个不同供体(供体1(C)和供体2(D))的对照存在时，通过T细胞的INFγ的分泌，来测量激活情况。

图26：A431条件介质与不同样品孵育24小时后IL-8的产量水平分析。IL8相对含量使用DUOSET ELISA得以确定，并以450nm下的光学单位表示。

图27：hIFNα2a诱导ISRE依赖性的荧光素酶依赖性生产。在c9G4-PVGLIG-hIFNα2a(A)、NHS76-PVGLIG-hIFNα2a(B)和H16/L16-PVGLIG-hIFNα2a中对hIFNα2a活性进行了测定，将细胞同10μg/ml H6/L16抗体(C)或没有同10μg/ml H6/L16抗体(D)进行预孵育，通过监测GloResponse

图28：有/无尿激酶孵育6小时后IL15的活性。IL15相对含量使用IL15生物测定法得以确定，并以发光表示。

图29A：在uPA介导的H16/L16-SGRSA hIFNα2a(A)和H16/L16-PSSRRRVN hIFNα2a(B)裂解后的hIFN活性评估。将H16/L16-SGRSA hIFNα2a(A)和H16/L16-PSSRRRVN hIFNα2a(B)在有/无尿激酶的条件下孵育24h，然后在ISRE-luc2/HEK293中将IGF1R受体饱和后，测定hIFNα活性。相对hIFNα活性使用GloResponse ISRE-luc2P生物测定法得以确定，并以发光表示。

图30：c9G4-GSRS-CXCL10(A)、NHS76-GSRS-CXCL10(B)和H16/L16-SGRS-CXCL10(C)在uPA介导的裂解后的hCXCL10活性评估。相对hCXCL10活性使用PathHunter eXpressCXCR3 CHOK1β-抑制蛋白(arrestin)GPCR测定法得以确定，并以发光表示。

发明内容

令人惊讶地发现，抗体与细胞因子部分融合的特定组合，其可在裂解可裂解肽连接子时选择性释放，提供了新型的和治疗上有效的融合蛋白。

本发明涉及“免疫细胞因子”，即抗体或其片段或衍生物与细胞因子之间的融合体。本发明免疫细胞因子中的抗体部分靶向肿瘤，在所述肿瘤中细胞因子被释放以发挥其作用。其赋予了融合蛋白更大的特异性，即与仅依赖于局部蛋白水解将细胞因子活性靶向至肿瘤的现有技术的免疫细胞因子相比，其产生的副作用更少(Skrombolas等人，2019)。

尽管现有技术的其他免疫细胞因子在抗体和细胞因子之间不包含任何连接子或仅包含结构连接子(即没有任何特定生物学活性的连接子)，但本发明的融合蛋白包含肽连接子，所述肽连接子在两部分之间可被裂解，使得可以更好地控制分子的治疗活性。实际上，发明人已经发现融合蛋白在血液中令人惊讶地失活，但是在到达肿瘤部位时被激活。可裂解肽连接子优先在肿瘤微环境中裂解，从而释放细胞因子。细胞因子的靶向递送因此增强了其抗肿瘤活性，同时降低了细胞因子相关毒性的风险。

在第一个方面，本发明涉及一种融合蛋白，其包含抗体或其抗原结合蛋白、可裂解肽连接子、及细胞因子或其功能片段。

“融合蛋白”是指编码两个或更多个分开的蛋白序列的嵌合蛋白，所述蛋白序列重组表达为单个部分。该术语意在涵盖所有偶联物，其中所述抗体或其抗原结合蛋白通过某种方式与可裂解肽连接子及细胞因子或其功能片段结合，所述方式例如共价和/或非共价的，例如离子键。该术语涵盖所有结合的安排方式。优选的安排方式包括抗体-连接子-细胞因子，及细胞因子-连接子-抗体。

抗体

如本文所用，“抗体”是指免疫球蛋白(Ig)分子，其通过位于免疫球蛋白分子可变区的至少一个抗原识别位点，能够特异性结合靶标即“抗原”(例如碳水化合物、多核苷酸、脂质、多肽等)。本发明融合蛋白的抗体或其抗原结合蛋白介导免疫细胞因子向疾病环境和/或向特定细胞亚群的靶向递送。优选的靶抗原为在患病组织中过表达而在其他部位保持低水平的那些抗原。这样的抗原是技术人员众所周知的，因为其多年来已成为许多研究的主题。例如，本发明免疫细胞因子的抗体部分可以靶向在恶性细胞表面上过表达的抗原(例如上皮细胞粘附分子、EGFR、IGF-1R、GD2双唾液酸神经节苷脂、HER2/neu、CD20和CD30)，以及靶向在肿瘤和慢性炎症部位发现的新血管生成抗原(例如纤连蛋白、剪接变体EDA/EDB和腱生蛋白C的A1结构域)。

如本文所用，术语“抗体”不仅涵盖完整的多克隆或单克隆抗体，而且涵盖任何抗原结合片段(即antigen-binding fragment)或其单链，包含抗体的融合蛋白，以及任何其他免疫球蛋白分子的修饰的构型，其包含抗原识别位点(包括但不限于scFv、单结构域抗体(例如鲨鱼和骆驼抗体)、巨抗体(maxibody)、微抗体、内抗体、双抗体、三抗体、四抗体、v-NAR和bis-scFv)。如本文所用，术语“抗体”涵盖了全长抗体及其抗原结合片段，以及其任何衍生物。优选地，根据本发明的抗体或其衍生的化合物或抗原结合片段为单克隆抗体。

如本文所用，“单克隆抗体”是指源自几乎同质的抗体群体的抗体。更特别地，除了少数可按照最小比例发现的可能天然存在的突变以外，群体中的单个抗体为相同的。换句话说，单克隆抗体由源自单细胞克隆生长的同质抗体组成(例如杂交瘤、被编码同质抗体的DNA分子转染的真核宿主细胞、被编码同质抗体的DNA分子转染的原核宿主细胞等)，且通常有且仅有一种类型和亚类的重链和仅一种类型的轻链为特征。单克隆抗体具有高度特异性，并且针对单一抗原。另外，与多克隆抗体(通常包括针对各种决定簇或表位的各种抗体)的制备相反，每种单克隆抗体针对抗原的单个表位。由于这些抗体针对单个表位，其为高度特异性的。

“表位”为抗原上与抗体结合的位点。其可以由连续的残基、或由于抗原蛋白折叠而变得非常接近的非连续残基形成。由连续氨基酸形成的表位通常在暴露于变性溶剂时保留，而由非连续氨基酸形成的表位通常在所述暴露下丢失。

与特定抗原反应的抗体的产生可以通过本领域技术人员已知的任何方法来实现，例如骨髓瘤细胞与来自免疫小鼠或与选定的骨髓瘤细胞相容的其他物种的脾细胞融合(Kohler和Milstein,Nature,256：495-497,1975)。免疫动物可以包括具有人免疫球蛋白基因座的转基因小鼠，其随后直接产生人抗体。或者，可以通过重组方法来产生抗体，例如选择重组抗体在噬菌体或类似载体中的文库。通常，为了制备单克隆抗体或其功能片段，特别是鼠源的，可以参考特别地在手册“抗体”中描述的技术(Harlow和Lane，《抗体：实验室手册》，纽约州冷泉港，冷泉港实验室，726页，1988)，或Kohler和Milstein描述的由杂交瘤制备的技术(Nature,256：495-497,1975)。

抗体包括任何类别的抗体，例如IgG、IgA或IgM(或其亚类)，且抗体不必为任何特定类别。免疫球蛋白可根据抗体重链恒定区的氨基酸序列分为不同类别。

典型的IgG抗体由通过二硫键连接的两条相同的重链和两条相同的轻链组成。每条重链和轻链包含恒定区和可变区。每个可变区包含三个片段，称为“互补决定区”(“CDRs”)或“高变区”，其主要负责结合抗原的表位。其通常称为CDR1、CDR2和CDR3，从N-端开始顺序编号。可变区的更高度保守的部分称为“框架区”。

有三个重链CDRs和三个轻链CDRs。此处使用术语“CDR”或“CDRs”是为了根据情况表示这些区域中的一个或多个，或甚至整个区域，其包含负责通过抗体对抗原或其识别的表位的亲和力结合的大部分氨基酸残基。

如本文所用，“VH”或“V

抗体恒定结构域不直接参与抗体与抗原的结合，但具有多种效应子功能。对应于不同类别的免疫球蛋白的重链恒定区分别称为α、δ、ε、γ和μ。根据其重链恒定区的氨基酸序列，可以将抗体或免疫球蛋白分为不同类别，即IgA、IgD、IgE、IgG和IgM，其中一些可以进一步分为亚类(同种型)，例如IgG1、IgG2、IgG3和IgG4；IgA1和IgA2(参见W.E.Paul编辑，1993，《基础免疫学》，Raven出版社，纽约州纽约)。

木瓜蛋白酶消化抗体会产生两个相同的抗原结合片段，称为Fab片段，每个片段都具有一个抗原结合位点，以及一个残留的“Fc”片段。已经确定了人IgG Fc结构域的晶体结构(Deisenhofer,Biochemistry,20,2361-2370,1981)。如说明书和权利要求书中所用，“免疫球蛋白Fc结构域或Fc”是指免疫球蛋白重链恒定区的羧基末端部分。如本文所用，“天然序列Fc结构域”包含与天然发现的Fc结构域的氨基酸序列相同的氨基酸序列。天然序列人Fc结构域包括天然序列人IgG1 Fc结构域(非A和A同种异型)；天然序列人IgG2 Fc结构域；天然序列人IgG3 Fc结构域；及天然序列人IgG4 Fc结构域，及其天然存在的变体。

尽管免疫球蛋白重链Fc结构域的边界可能会变化，但通常将人IgG重链Fc结构域定义为从铰链区Cys226或Pro230位的氨基酸残基延伸到包含重链的CH2和CH3结构域的羧基末端。在整个本说明书和权利要求书中，免疫球蛋白重链中的残基编号为EU索引编号，如Kabat等人，Sequences of Proteins of Immunological Interest，第5版，美国国立卫生研究院公共卫生服务，马里兰州贝塞斯达(1991)。“如Kabat的EU索引编号”是指人IgG1 EU抗体的残基编号。

术语“铰链区”通常定义为从人IgG1的Glu216延伸到Pro230(Burton,MoIImmunol,22：161-206,1985)。通过将形成重链间S-S键的第一个和最后一个半胱氨酸残基置于相同位置，可以将其他IgG同种型的铰链区与IgG1序列对齐。人IgG Fc部分的“CH2结构域”(也称为“Cγ2”结构域)通常从约氨基酸231延伸至约氨基酸340。CH2结构域的独特之处在于其不与另一个结构域紧密配对。而是，两个N-连接的支链碳水化合物链插入完整的天然IgG分子的两个CH2结构域之间。据推测，碳水化合物可为结构域-结构域配对提供替代物，并有助于稳定CH2结构域(Burton,MoI Immunol,22：161-206,1985)。“CH3结构域”包括在Fc部分中C-末端残基延伸至CH2结构域(即从IgG的约氨基酸残基341至约氨基酸残基447)。

Fc结构域在确定免疫球蛋白的生物学功能中至关重要，所述生物学功能被称为“效应子功能”。这些Fc结构域介导的活性通过免疫效应细胞(如杀伤细胞、自然杀伤细胞和激活的巨噬细胞)或各种补体成分介导。通过抗体的Fc结构域与受体或补体成分的结合,这些效应子功能涉及激活所述效应细胞表面上的所述受体。抗体依赖性细胞毒性(antibody-dependent cellular cytotoxicity，ADCC)和补体依赖性细胞毒性(complement-dependent cytotoxicity，CDC)活性涉及Fc结构域与效应细胞的Fc受体(如FcγRI、FcγRII、FcγRIII)或补体成分(如C1q)的结合。在各种人免疫球蛋白类别中，人IgG11和IgG3比IgG2和IgG4更有效地介导ADCC。

本发明的抗体还包含嵌合或人源化抗体。

嵌合抗体是含有衍生自给定物种的抗体的天然可变区(轻链和重链)及与所述给定物种异源的物种的抗体的轻链和重链的恒定区结合的抗体。

抗体或其嵌合片段可以通过使用重组遗传学技术来制备。例如，嵌合抗体可以通过克隆重组DNA来生产，所述重组DNA包含启动子和编码本发明的非人单克隆抗体的可变区的序列，尤其是是鼠的序列，以及编码人抗体恒定区的序列。由这样的重组基因编码的根据本发明的嵌合抗体可以为例如小鼠-人嵌合体，该抗体的特异性由鼠DNA衍生的可变区决定，而其同种型由人DNA衍生的恒定区决定。应当理解，在这种情况下，嵌合抗体的Fc结构域是人源的。参阅Verhoeyn等人(BioEssays,8：74,1988)制备嵌合抗体的方法。

另外，本发明还涉及源自上述鼠抗体的人源化抗体。“人源化抗体”在本文中是指包含衍生自非人源抗体的CDR区的抗体，所述抗体分子的其他部分衍生自一种(或几种)人抗体。另外，一些骨架片段残基(称为FR)可以被修饰以保持结合亲和力(Jones等人,Nature,321：522-525,1986；Verhoeyen等人,Science,239：1534-1536,1988；Riechmann等人,Nature,332：323-327,1988)。如在嵌合抗体中一样，人源化抗体的Fc结构域将是人源的。

本发明的人源化抗体或其片段可以通过本领域技术人员已知的技术来制备(例如在文献Singer等人,J.Immun.,150：2844-2857,1992；Mountain等人,Biotechnol.Genet.Eng.Rev.,10：1-142,1992；及Bebbington等人,Bio/Technology,10：169-175,1992中描述的技术)。这种人源化抗体优选用于涉及体外诊断或体内预防和/或治疗性处理的方法。本领域技术人员还已知其他人源化技术，例如PDL在专利EP 0 451 261，EP 0 682 040、EP 0 939 127、EP 0 566 647或US 5,530,101、US 6,180,370、US 5,585,089和US 5,693,761中描述的“CDR移植”技术。也可以引用美国专利5,639,641或6,054,297、5,886,152和5,877,293。

尽管可以在本发明的免疫细胞因子中使用抗体片段，但是优选使用全长的二价抗体。单价抗体(例如Fab或scFv)仅具有一个抗原结合位点(不同于天然的“二价”抗体)，即由一个抗原结合臂组成。如技术人员所知，免疫球蛋白的化合价(抗原结合位点数)越大，其结合的抗原量就越大。单价和二价结合之间的亲和力变化显著，Kd值变化了1500倍。因此，可以以较低剂量使用二价抗体以获得与单价Fab或scFv片段相似的治疗效率，从而限制继发效应的风险。

优选地，可用于本文所述的免疫细胞因子的抗体为不与所述免疫细胞因子的细胞因子部分特异性结合的抗体。例如，如果细胞因子为IL-12，则根据该实施方案的抗体不是结合IL-12的抗体。

在另一个实施方案中，用于本发明免疫细胞因子的抗体为特异性结合肿瘤相关抗原(tumour-associated antigens，TAA)或肿瘤特异性抗原(tumour-specific antigens，TSA)的抗体。如本文所用，“肿瘤特异性抗原”为在癌细胞上发现的蛋白质或其他分子，而“肿瘤特异性抗原”为在癌细胞上而不在正常细胞上发现的蛋白质或其他分子。肿瘤特异性抗原是本领域已知的。

肿瘤抗原可以以多种方式分类。肿瘤抗原包括由经历了染色体改变的基因编码的抗原。这些抗原中有许多是在淋巴瘤和白血病中发现的。即使在这种分类下，也可以将抗原表征为涉及静止基因激活的抗原。其包括BCL-1和IgH(套细胞淋巴瘤)、BCL-2和IgH(滤泡性淋巴瘤)、BCL-6(弥漫性大B细胞淋巴瘤)、TAL-1和TCRδ或SIL(T细胞急性淋巴细胞白血病)、c-MYC和IgH或IgL(Burkitt淋巴瘤)、MUN/IRF4和IgH(骨髓瘤)、PAX-5(BSAP)(免疫细胞瘤)。

其他涉及染色体改变从而产生新的融合基因和/或蛋白质的肿瘤抗原包括RARoa、PML、PLZF、NPM或NuM4(急性早幼粒细胞白血病)、BCR和ABL(慢性髓样/急性淋巴细胞白血病)、MLL(HRX)(急性白血病)、E2A和PBX或HLF(B细胞急性淋巴细胞性白血病)、NPM、ALK(间变性大细胞白血病)和NPM、MLF-1(骨髓增生异常综合症/急性骨髓性白血病)。

其他肿瘤抗原对于组织或细胞系具有特异性。其包括细胞表面蛋白，例如CD20、CD22(非霍奇金淋巴瘤、B细胞淋巴瘤、慢性淋巴细胞性白血病(CLL))、CD52(B细胞CLL)、CD33(急性骨髓性白血病(AML))、CD10(gp100)(常见(B前)急性淋巴细胞白血病和恶性黑色素瘤)、CD3/T细胞受体(TCR)(T细胞淋巴瘤和白血病)、CD79/B细胞受体(BCR)(B细胞淋巴瘤和白血病)、CD26(上皮和淋巴样恶性肿瘤)、人白细胞抗原(HLA)-DR、HLA-DP和HLA-DQ(淋巴样恶性肿瘤)、RCAS1(妇科癌症、胆道腺癌和胰腺导管腺癌)和前列腺特异性膜抗原(前列腺癌)。

组织或谱系特异性肿瘤抗原还包括表皮生长因子受体(高表达)，例如EGFR(HER1或erbB1)和EGFRvIII(脑、肺、乳腺、前列腺和胃癌)、erbB2(HER2或HER2/neu)(乳腺癌和胃癌)、erbB3(HER3)(腺癌)和erbB4(HER4)(乳腺癌)。

组织或谱系特异性肿瘤抗原还包括细胞相关蛋白，例如酪氨酸酶、Melan-A/MART-1，酪氨酸酶相关蛋白(TRP)-1/gp75(恶性黑色素瘤)，多态性上皮粘蛋白(PEM)(乳腺癌)和人上皮粘蛋白(MUC1)(乳腺、卵巢、结肠和肺癌)。

组织或谱系特异性的肿瘤抗原还包括分泌的蛋白质，例如单克隆免疫球蛋白(多发性骨髓瘤和浆细胞瘤)、免疫球蛋白轻链(多发性骨髓瘤)、甲胎蛋白(肝癌)、激肽释放酶6和10(卵巢癌)、胃泌素释放肽/蛙皮素(肺癌)和前列腺特异性抗原(前列腺癌)。

还有其他肿瘤抗原是在某些正常组织(例如睾丸)和某些情况下在胎盘中表达的癌-睾丸(CT)抗原。其表达在不同谱系的肿瘤中很常见，并且抗原作为一个整体形成免疫疗法的靶标。CT抗原的肿瘤表达的示例包括MAGE-A1、-A3、-A6、-A12、BAGE、GAGE、HAGE、LAGE-1、NY-ESO-1、RAGE、SSX-1、-2、-3、-4、-5、-6、-7、-8、-9、HOM-TES-14/SCP-1、HOM-TES-85和PRAME。CT抗原及表达CT抗原的癌症的其他例子包括SSX-2和-4(神经母细胞瘤)，SSX-2(HOM-MEL-40)，MAGE、GAGE、BAGE和PRAME(恶性黑色素瘤)，HOM-TES-14/SCP-1(脑膜瘤)，SSX-4(少突神经胶质瘤)，HOM-TES-14/SCP-1、MAGE-3和SSX-4(星形细胞瘤)，SSX家族成员(头颈癌、卵巢癌、淋巴瘤、结直肠癌和乳腺癌)，RAGE-1、-2、-4、GAGE-1-2、-3、-4、-5、-6、-7和-8(头颈部鳞状细胞癌(HNSCC))，HOM-TES14/SCP-1、PRAME、SSX-1和CT-7(非霍奇金淋巴瘤)和PRAME(急性淋巴细胞白血病(ALL)、急性骨髓性白血病(AML)和慢性淋巴细胞性白血病(CLL))。

其他肿瘤抗原对于特定组织或细胞系不具有特异性。其包括癌胚抗原(CEA)家族成员：CD66a、CD66b、CD66c、CD66d及CD66e。这些抗原可以在许多不同的恶性肿瘤中表达，并可以通过免疫疗法靶向。

还有其他肿瘤抗原为病毒蛋白，其包括人乳头瘤病毒蛋白(宫颈癌)和EBV编码核抗原(EBNA)-1(颈部淋巴瘤和口腔癌)。

还有其他肿瘤抗原为突变的或异常表达的分子，例如但不限于CDK4和β-连环蛋白(黑色素瘤)。

在一些实施方案中，抗原为肿瘤抗原。肿瘤抗原可以选自MART-1/Melan-A、gp100、腺苷脱氨酶结合蛋白(ADAbp)、FAP、亲环蛋白b、结直肠相关抗原(CRC)-C017-1A/GA733、癌胚抗原(CEA)、CAP-1、CAP-2、etv6、AML1、前列腺特异性抗原(PSA)、PSA-1、PSA-2、PSA-3、前列腺特异性膜抗原(PSMA)、T细胞受体/CD3-ζ链和CD20。肿瘤抗原还可以选自MAGE-A1、MAGE-A2、MAGE-A3、MAGE-A4、MAGE-A5、MAGE-A6、MAGE-A7、MAGE-A8、MAGE-A9、MAGE-A10、MAGE-A11、MAGE-A12、MAGE-Xp2(MAGE-B2)、MAGE-Xp3(MAGE-B3)、MAGE-Xp4(MAGE-B4)、MAGE-C1、MAGE-C2、MAGE-C3、MAGE-C4、MAGE-C5。在另一个实施方案中，肿瘤抗原选自GAGE-1、GAGE-2、GAGE-3、GAGE-4、GAGE-5、GAGE-6、GAGE-7、GAGE-8、GAGE-9。在一个进一步的实施方案中，肿瘤抗原选自BAGE、RAGE、LAGE-1、NAG、GnT-V、MUM-1、CDK4、酪氨酸酶、p53、MUC家族、HER2/neu、p21 ras、RCAS 1、甲胎蛋白、E-钙黏素、α-连环素、β-连环素、γ-连环素、p120ctn、gp100 Pmel117、PRAME、NY-ESO-1、cdc27、腺瘤性息肉病蛋白(APC)、胞衬蛋白(fodrin)、连接蛋白37、Ig独特型、p15、gp75、GM2神经节苷脂、GD2神经节苷脂、人乳头瘤病毒蛋白、Smad家族的肿瘤抗原、Imp-1、P1A、EBV编码的核抗原(EBNA)-1、脑糖原磷酸化酶、SSX-1、SSX-2(HOM-MEL-40)、SSX-1、SSX-4、SSX-5、SCP-1和CT-7和c-erbB-2。

癌症或肿瘤抗原也可以根据它们所关联(即被其表达)的癌症或肿瘤来分类。与肿瘤抗原相关的癌症或肿瘤包括急性淋巴细胞白血病(etv6；am11；亲环蛋白b)、B细胞淋巴瘤(Ig独特型)；Burkitt(非霍奇金)淋巴瘤(CD20)；胶质瘤(E-钙黏素；α-连环素；β-连环素；γ-连环素；p120ctn)、膀胱癌(p21 ras)、胆道癌(p21ras)、乳腺癌(MUC家族；HER2/neu；c-erbB-2)、宫颈癌(p53；p21 ras)、结肠癌(p21 ras；HER2/neu；c-erbB-2；MUC家族)、结直肠癌(结直肠相关抗原(CRC)-0017-1A/GA733；APC)、绒毛膜癌(CEA)、上皮细胞癌(亲环蛋白b)、胃癌(HER2/neu；c-erbB-2；ga733糖蛋白)、肝细胞癌(甲胎蛋白)、霍奇金淋巴瘤(lmp-1；EBNA-1)、肺癌(CEA；MAGE-3；NY-ESO-1)、淋巴样细胞源性白血病(亲环蛋白b)、黑素瘤(p15蛋白、gp75、癌胚(oncofoetal)抗原、GM2和GD2神经节苷脂)、骨髓瘤(MUC家族；p21 ras)、非小细胞肺癌(HER2/neu；c-erbB-2)、鼻咽癌(lmp-1；EBNA-1)、卵巢癌(MUC家族；HER2/neu；c-erbB-2)，前列腺癌(前列腺特异性抗原(PSA)及其免疫原性表位PSA-1、PSA-2和PSA-3；PSMA；HER2/neu；c-erbB-2)、胰腺癌(p21 ras；MUC家族；HER2/neu；c-erbB-2；ga733糖蛋白)、肾癌(HER2/neu；c-erbB-2)、宫颈和食道鳞状细胞癌(病毒产物，例如人乳头瘤病毒蛋白和非感染性颗粒)、睾丸癌(NY-ESO-1)、T细胞白血病(HTLV-1表位)和黑素瘤(Melan-A/MART-1；cdc27；MAGE-3；p21 ras；gp100

可以用于本发明的抗体的示例包括：阿仑单抗(CD52)、阿利珠单抗(Alirocumab)(PCSK9)、阿西莫单抗(Arcitumomab)(人CEA(癌胚抗原))、阿特珠单抗(PD-L1)、阿维单抗(PD-L1)、巴利昔单抗(CD25(IL2受体的一条链))、贝利木单抗(Belimumab)(BLyS)、贝伐单抗(CD19)、贝伐单抗(VEGF)、布罗达单抗(Brodalumab)(IL-17RA)、卡罗单抗(肿瘤表面抗原PSMA)、卡妥索单抗(Catumaxomab)(EpCAM和CD3)、卡妥索单抗(EpCAM)、PEG化赛妥珠单抗(TNFa)、西妥昔单抗(EGFR)、达雷木单抗(CD38)、达妥昔单抗(GD2)、度匹鲁单抗(IL-4Rα)、德瓦鲁单抗(PD-L1)、依法利珠单抗(CD11a)、埃罗妥珠单抗(SLAMF7)、依洛尤单抗(LDL-C/PCSK9)、法索单抗(CD15)、戈利木单抗(Golimumab)(TNFa)、替伊莫单抗(Ibritumomabtiuxetan)(CD20)、英夫利昔单抗(TNFα)、伊匹单抗(CTLA-4)、耐昔妥珠单抗(EGFR)、耐昔妥珠单抗(CD25(IL2受体的一条链))、纳武单抗(PD-1)、诺非单抗(肿瘤相关抗原)、奥滨尤妥珠单抗(Obinutuzumab)(CD20)、奥瑞珠单抗(CD20)、奥法木单抗(CD20)、奥拉木单抗(PDGFR-α)、帕尼单抗(EGFR)、派姆单抗(PD-1)、帕妥珠单抗(HER2)、雷莫芦单抗(VEGF)、雷珠单抗(VEGF-A)、利妥昔单抗(CD20)、司妥昔单抗(cCLB8)、托珠单抗(IL-6受体)、曲妥珠单抗(HER-2)、维多珠单抗(整合素-α4β7)、伏妥莫单抗(细胞角蛋白肿瘤相关抗原)。

可裂解肽连接子

如本文所用，“可裂解肽连接子”是指多价连接子，其与抗体或其抗原结合片段共价结合，并与细胞因子或其片段共价结合，其可被酶裂解(例如在裂解位点)。根据本发明，在可裂解肽连接子水解(蛋白水解裂解)后，释放细胞因子部分(优选IL-15)，使其能够发挥其治疗活性。在优选的实施方案中，可裂解肽连接子被重组表达为免疫细胞因子的一部分。在其他实施方案中，可裂解肽连接子是通过使用例如偶联物化学方法，使连接于连接子的官能(反应性)基团与抗体或其抗原结合片段反应而形成的连接子。在其他实施方案中，可裂解肽连接子是通过使用例如偶联物化学方法，使连接于连接子的官能(反应性)基团与细胞因子或其片段反应而形成的连接子。在一个优选的实施方案中，所述可裂解肽连接子将细胞因子或其片段连接于抗体或其抗原结合片段的重链。在另一个实施方案中，可裂解肽连接子将细胞因子或其片段连接于抗体或其抗原结合片段的轻链。可裂解肽连接子可将细胞因子或其片段连接于抗体或其抗原结合片段的重链和轻链之一的N-末端。可裂解肽连接子也可能将细胞因子或其片段连接于抗体或其抗原结合片段的重链和轻链的C-末端。更优选地，可裂解肽连接子将细胞因子或其片段连接于抗体或其抗原结合片段的重链的N-末端或C-末端。

本文提供的可裂解肽连接子可包括蛋白酶裂解位点。

如本文所用，“裂解位点”是指用于裂解如本文所述的免疫细胞因子中存在的一部分连接子(例如如上所述的可裂解肽连接子)的可识别的位点。因此，可在本文所述的可裂解肽连接子的序列中发现裂解位点，包括其实施方案。在实施方案中，裂解位点是被裂解剂识别和裂解的氨基酸序列(例如肽基序列)。示例性裂解剂包括蛋白质、酶、DNA酶、RNA酶、金属、酸和碱。示例性裂解位点在本文中定义(参见图7)。

如本文所用，“蛋白酶裂解位点”是被蛋白酶识别并特异性裂解的裂解位点。根据一个优选的实施方案，蛋白酶裂解位点为肿瘤相关的蛋白酶裂解位点。如本文所用，“肿瘤相关的蛋白酶裂解位点”是指被蛋白酶识别的氨基酸序列，其表达对于肿瘤细胞或其肿瘤细胞环境是特异性的，或与健康组织相比主要在肿瘤细胞或肿瘤环境中表达，或仅/主要在肿瘤细胞或肿瘤环境中具有活性。在实施方案中，蛋白酶裂解位点为基质金属蛋白酶(MMP)裂解位点、前列腺特异性抗原(PSA)蛋白酶裂解位点、膜型丝氨酸蛋白酶1(MT-SP1)蛋白酶裂解位点、uPA尿激酶纤溶酶原激活物裂解位点、或豆荚蛋白(legumain)蛋白酶裂解位点。在一些实施方案中，基质金属蛋白酶(MMP)裂解位点为MMP 9裂解位点、MMP 13裂解位点或MMP 2裂解位点。蛋白酶裂解位点可以由特定的氨基酸序列指定，但是也可以涵盖该标准氨基酸序列的任何变体，所述变体仍被目标蛋白酶识别和裂解。

优选地，可裂解肽连接子为基质金属蛋白酶(MMP)裂解位点。更优选地，可裂解肽连接子包含MMP 9裂解位点或MMP 2裂解位点。MMP裂解位点的示例包括GPLGIAGQ、GPLGLWAQ、GPLGMLSQ、PLGLAG和PVGLIG。

在另一个优选的实施方案中，可裂解肽连接子为尿激酶纤溶酶原激活物(uPA)的裂解位点。uPA裂解位点的示例包括SGRS、SGRSA和PSSSRRRVN。

如本文所用，术语“MMP 2”或“MMP 2蛋白酶”是指基质金属蛋白酶2(MMP 2)。MMP 2是由NCBI序列参考号GI 189217853识别的蛋白质。如本文所用，术语“MMP 9”或“MMP 9蛋白酶”是指基质金属蛋白酶9(MMP 9)。MMP 9是由NCBI序列参考号GI 74272287识别的蛋白质。本文所用的术语“MMP 13”或“MMP 13蛋白酶”是指基质金属蛋白酶13(MMP 13)。MMP 13是由NCBI序列参考号GL 4505209识别的蛋白质。如本文所用，术语“PSA”或“PSA蛋白酶”是指前列腺特异性抗原(PSA)，也称为γ精原蛋白或激肽释放酶-3。PSA是由NCBI序列参考号GL71834853识别的蛋白质。如本文所用，术语“PSMA”或“PSMA蛋白酶”是指前列腺特异性膜抗原(PSMA)，也称为谷氨酸羧肽酶II(GCPII)、N-乙酰基-L-天冬氨酰-L-谷氨酸肽酶I(NAALADase I)或NAAG肽酶。PSMA是由NCBI序列参考号GL 62548858识别的蛋白质。如本文所用，术语“成纤维细胞相关蛋白”是指成纤维细胞相关蛋白。成纤维细胞相关蛋白是由NCBI序列参考号GL 1888316识别的蛋白质。如本文所用，术语“MT-SP1”或“MT-SP1蛋白酶”是指膜型丝氨酸蛋白酶1(MT-SP1)。MT-SPl是由NCBI序列参考号GI 11415040识别的蛋白质。如本文所用，术语“豆荚蛋白”(legumain)或“豆荚蛋白酶”(legumain protease)是指豆荚蛋白。豆荚蛋白是由NCBI序列参考号GL 2842759识别的蛋白质。本文所用的术语uPA是指通过NCBI序列参考基因ID 5328识别的尿激酶型纤溶酶原激活剂。

在一些实施方案中，可裂解肽连接子包含至少2个、至少3个、至少4个、至少5个、至少6个、至少7个、至少8个、至少9个、至少10个、至少11个、至少12个或至少13个氨基酸。在其他实施方案中，可裂解肽连接子为5聚体(即长度为5个氨基酸的肽)、6聚体(即长度为6个氨基酸的肽)、7聚体(即长度为7个氨基酸的肽)、8聚体(即长度为8个氨基酸的肽)、9聚体(即长度为9个氨基酸的肽)、10聚体(即长度为10个氨基酸的肽)、11聚体(即长度为11个氨基酸的肽)、12聚体(即长度为12个氨基酸的肽)或13聚体(即长度为13个氨基酸的肽)。

最优选地，所述可裂解肽连接子的所述序列选自：GPLGIAGQ、GPLGLWAQ、GPLGMLSQ、PLGLAG、PVGLIG、SGRS、SGRSA、及PSSSRRRVN。

细胞因子

如本文所用，术语“细胞因子”是指对免疫系统有影响的小分泌调节蛋白家族的成员。细胞因子参与细胞间的通讯并调节许多细胞功能，例如细胞存活和生长以及许多基因表达的诱导。因此，细胞因子的分泌能够以多方面和有效的方式快速传播免疫信号。细胞因子通过对淋巴细胞和/或其他细胞产生多种作用来调节免疫反应的性质、强度和持续时间。实际上，细胞因子通常分为促炎和抗炎细胞因子。一些细胞因子还能够动员免疫系统抵抗癌症(参见例如Floros和Tarhini,Semin Oncol.42(4)：539–548,2015)。细胞因子可由多种细胞类型产生，包括免疫细胞和非免疫细胞。细胞因子的示例尤其包括白介素、淋巴因子、单核因子、干扰素、集落刺激因子和趋化因子。如本文所用，“细胞因子”可以为以下任一项：IL-1、IL-2、IL-3、IL-4、IL-5、IL-6、IL-7、IL-8、IL-9、IL-10、IL-11、IL-12、IL-13、IL-15、IL-16、IL-17、IL-18、IL-19、IL-20、IL-21、IL-22、IL-23、、IL-24、IL-26、IL-28、IL-29、IL-33、IL-36、IL37、IL-38、IFN-α(包括IFN-α1/13、IFN-α2、IFN-α4、IFN-α5、IFN-α6、IFN-α7、IFN-α8、IFN-α10、IFN-α14、IFN-α16、IFN-α17、及IFN-α21)、IFN-β、IFN-γ、IFN-λ、TNF-α、TNF-β、TGF-β1、M-CSF、G-CSF、GM-CSF、及CXL10。根据本发明，细胞因子优选在与ICC连接时被失活或减弱，并且仅在被蛋白酶与ICC裂解后才具有活性。

关于所述细胞因子的术语“功能片段”应基本上类似于抗体的相同术语来解释(见下文)。细胞因子的功能片段和衍生物是与天然存在的细胞因子基本具有相同的生理功能/活性的那些。

在一个优选的实施方案中，细胞因子为IL-15。术语“IL-15”或“白介素-15”在本文中是指调节T细胞和自然杀伤细胞激活和增殖的细胞因子。白细胞介素-15(IL-15)是4α-螺旋束细胞因子家族的14至15kDa的成员，其由114个氨基酸组成，以登录号NP_000576.1可获得其序列。在人和猿IL-15之间有97％的序列同一性，在人和小鼠之间有73％的序列同一性。这似乎足以使huIL-15在猿和小鼠细胞上具有生物活性。

IL-15与白介素2(IL-2)具有高度的结构相似性。像IL-2一样，IL-15结合并通过由IL-2/IL-15受体β链(CD122)和共同的γ链(γ-C，CD132)组成的复合物发出信号。IL15被其受体(IL15RA)的α单元所识别，从而确保了信号传导的特异性，而IL-2则与IL2RA结合。IL-15刺激促炎性细胞因子(例如TNFα、IL-1、IFNγ)的产生、激活的B细胞的增殖和Ig合成、T

在另一个优选的实施方案中，细胞因子为CXCL10。“CXCL10”或“C-X-C基序趋化因子10”或“干扰素γ诱导的蛋白质10”或“IP10”，在本文中是指8.7kDa的CXC趋化因子，其作用为将激活的和记忆淋巴细胞募集到炎症部位。其分泌的生物活性形式(在信号肽裂解后)是具有77个残基的多肽(对应于NP_001556的22-98位)，在本文中称为“长CXCL10”，其结合CXCR3受体。CXCL10通过趋化因子受体CXCR3进行的信号传导，在淋巴细胞迁移和功能中具有重要作用。值得注意的是，CXCL10似乎增强了T细胞依赖性抗癌免疫力(Karin和Razon,Cytokine,109：24-28,2018)。

在另一个优选的实施方案中，细胞因子为IL-36。如本文所用，表述“IL-36”或“hIL-36”或“白介素-36”是指具有促炎性质的IL-1家族的亚组(参见例如Murrieta-Coxca,Int J Mol Sci.,20(7).pii：E1649,2019)。“IL-36”尤其是指IL-36α(IL-1F6)、IL-36β(IL-1F8)和IL-36γ(IL-1F9)。如本文所用，“IL-36α”是指158个氨基酸的蛋白质，其序列可以由登录号NP_055255获得，“IL-36β”是指具有2个同种型的157个氨基酸的蛋白质(登录号NP_055253和NP_775270)，而“IL-36γ”是一种具有2个同种型的169个氨基酸的蛋白质(登录号：NP_001265497和NP_062564)。IL-36α、IL-36β和IL-36γ是具有促炎活性的激动剂配体。其促进多种炎症介质的诱导，所述炎症介质包括细胞因子、趋化因子、生长因子和抗菌肽。其均使用相同的受体IL-36R，该受体与IL-1RAcP二聚化以激活细胞内信号级联。该途径以AP-1(激活蛋白1)和NF-kB转录因子驱动的炎性细胞因子的表达达到顶点。

在另一个优选的实施方案中，细胞因子为IFNα。如本文所用，表述“IFNα”或“IFN-α”或“干扰素α”是指人I型干扰素(IFN-I)的亚型，其为干扰素蛋白的一大亚类，其有助于调节免疫系统的活性。所有的IFN-I(包括IFN-α)都与特定的细胞表面受体复合物结合，该复合物称为IFN-α受体(IFNAR)，由IFNAR1和IFNAR2链组成(参见例如López de Padilla和Niewold,Gene,576(1Pt 1)：14-21,2016)。有12种功能性人IFN-α蛋白(IFN-α1/13、IFN-α2、IFN-α4、IFN-α5、IFN-α6、IFN-α7、IFN-α8、IFN-α10、IFN-α14、IFN-α16、IFN-α17和IFN-α21)，其一级、二级和三级结构都具有高度同源性。每种IFNA基因(IFNA1、IFNA2、IFNA3、IFN4、IFN5、IFN6、IFN7、IFN8、IFNA10、IFNA14、IFNA16、IFNA17和IFNA21)编码包含23个氨基酸的信号肽的前蛋白，该蛋白在分泌时会被裂解，产生成熟的166个残基的蛋白质(Uniprot登录号：Q6QNB6)，仅由165个氨基酸组成的IFN-2除外(Uniprot登录号：P01563)。实际上，在IFN-α的其他亚型中存在的于44位的天冬氨酸残基在IFN-α2中缺失。

根据该实施方案，本发明涉及一种融合蛋白，其包含：

(i)抗体或其抗原结合片段，其融合至下列

(ii)可裂解肽连接子，及

(iii)细胞因子，优选IL-15、CXCL10、IL-36或IFN-α，或其功能片段。

可裂解肽连接子和细胞因子的鉴定方法

发明人已经表明，可以鉴定适用于本发明融合蛋白的连接子。实验数据表明，根据其在体外是否被裂解，可以区分几种连接子。这些体外结果的相关性是这样一个事实所强调的，即这样鉴定出的所有连接子都在体内被裂解，从而在肿瘤部位释放出活性细胞因子。

因此，在另一方面，本文提供了选择可裂解肽连接子的方法。所述方法包括以下步骤：

(i)提供如本文所述的融合蛋白，所述融合蛋白包含待测试的可裂解肽连接子；

(ii)使所述融合蛋白与相关蛋白酶接触；及

(iii)检测所述融合蛋白的裂解情况。

在一个实施方案中，所述方法在体外进行。在另一个实施方案中，所述方法在体内进行。根据该具体实施方案，所述方法的步骤(ii)包括将步骤(i)的融合蛋白施用至哺乳动物，优选啮齿动物，最优选小鼠。

可以通过本领域技术人员可用的任何方式来检测连接子的裂解。其尤其可以使用特异性抗体来进行，特别是使用众所周知的技术，例如用特异性抗体进行免疫沉淀、蛋白质印迹、ELISA或ELISPOT、抗体微阵列、或与免疫组织化学偶联的组织微阵列。其他合适的技术包括FRET或BRET、使用单个或多个激发波长并应用任何适合的光学方法的单细胞显微镜或组织化学方法，例如电化学方法(伏安法和安培法)、原子力显微镜法和射频法，例如多极共振光谱法、共聚焦和非共聚焦、检测荧光、发光、化学发光、吸光度、反射率、透射率以及双折射或折射率(例如表面等离子体共振、椭圆偏振法、共振镜方法、光栅耦合波导法或干涉法)、细胞ELISA、流式细胞仪、放射性同位素、磁共振成像、聚丙烯酰胺凝胶电泳(SDS-PAGE)分析；HPLC-质谱法；液相色谱/质谱/质谱(LC-MS/MS)。

可选地，还可以通过功能测试来检测连接子的裂解。值得注意的是，连接子的水解释放了活性细胞因子部分，而所述细胞因子部分作为融合蛋白的一部分时被减弱。在该实施方案中，所述方法的步骤(iii)包括测量细胞因子部分的活性。优选地，步骤(iii)还包括将已经与步骤(ii)中的相关蛋白酶接触的所述融合蛋白的细胞因子部分的活性与尚未与蛋白酶接触的所述融合蛋白的细胞因子部分的活性进行比较。根据一个优选的实施方案，连接子在下述情况下为被裂解的：在步骤(ii)中已经与相关蛋白酶接触的所述融合蛋白的细胞因子部分的活性，相对于未经处理的融合蛋白的细胞因子部分的活性，增加了至少2倍，优选至少3倍，优选至少4倍，优选至少5倍，优选至少10倍，优选至少20倍，优选至少50倍，优选至少100倍。更优选地，如果细胞因子部分的活性增加至少10倍，则连接子被裂解。

另外，立即显而易见的是，可以容易地测试细胞因子及其变体在本发明免疫细胞因子中的适用性。如本文所用，“细胞因子变体”是指不同于主要物种细胞因子的细胞因子。例如，细胞因子变体可以具有不同于主要物种细胞因子的氨基酸序列。通常，变体与主要物种细胞因子具有至少约70％的同源性，优选其与主要物种细胞因子具有至少约80％，更优选至少约90％的同源性。细胞因子变体在主要物种细胞因子的氨基酸序列内或附近的某些位置具有取代、缺失和/或添加。可选地，细胞因子变体可以在至少一种翻译后修饰中与主要物种细胞因子不同。例如，细胞因子变体可以带有一个或多个附着在其上的碳水化合物部分，所述碳水化合物部分与附着在主要物种抗体上的一个或多个碳水化合物部分不同。

因此，另一方面，本文提供了选择细胞因子或其变体的方法。所述方法包括以下步骤：

(i)提供如本文所述的融合蛋白，所述融合蛋白包含待测试的细胞因子或其变体；

(ii)使所述融合蛋白与相关蛋白酶接触；及

(iii)检测所述细胞因子的活性。

在一个实施方案中，所述方法在体外进行。在另一个实施方案中，所述方法在体内进行。根据该具体实施方案，所述方法的步骤(ii)包括将步骤(i)的融合蛋白施用至哺乳动物，优选啮齿动物，最优选小鼠。

如果细胞因子或变体适合用于本文所述的免疫细胞因子，则连接子的水解将释放活性细胞因子部分，而当所述细胞因子部分为融合蛋白的一部分时被减弱。在该实施方案中，所述方法的步骤(iii)包括测量细胞因子部分的活性。优选地，步骤(iii)还包括将已经与步骤(ii)中的相关蛋白酶接触的所述融合蛋白的细胞因子部分的活性与尚未与蛋白酶接触的所述融合蛋白的细胞因子部分的活性进行比较。根据一个优选的实施方案，细胞因子或细胞因子变体在下述情况下为有活性的：在步骤(ii)中已经与相关蛋白酶接触的所述融合蛋白的细胞因子部分的活性，相对于未经处理的融合蛋白的细胞因子部分的活性，增加了至少2倍，优选至少3倍，优选至少4倍，优选至少5倍，优选至少10倍，优选至少20倍，优选至少50倍，优选至少100倍。更优选地，如果所述活性增加至少10倍，则细胞因子或细胞因子变体为有活性的。

技术人员将知道如何根据所述细胞因子的性质来测量细胞因子部分的活性。细胞因子活性可以通过多种方法确定，包括但不限于酶片段互补技术(Eglen,JBiomolScreen.2004Aug；9(5)：398-408)、邻近连接法(Andersen等人,Cytokine.2013Oct；64(1)：54-7)、NF-κB易位(Trask 2012，测定指导手册[互联网].马里兰州贝塞斯达：礼来公司和国家转化科学促进中心；2004-2012Oct 1.)、β-抑制蛋白募集(Wang测定指导手册[互联网].马里兰州贝塞斯达：礼来公司和国家转化科学促进中心；2004-2017Nov 20.)、生物发光共振能量转移(Compan Methods Mol Biol.2016；1417：89-95)。这些方法已用于多种细胞因子，并可根据需要轻松适应特定目标细胞因子的需要。

技术人员将特别参考本申请的实验部分，其中描述了这样的测定的示例。

编码免疫细胞因子的多核苷酸

本文还提供了包含编码如上所述的融合蛋白的核苷酸序列的多核苷酸。本文还提供了在高严格、中等或较低严格杂交条件下(例如，如上文所定义)与编码本文提供的融合蛋白或修饰的融合蛋白的多核苷酸杂交的多核苷酸。

在某些实施方案中，本文提供的核酸分子包含或由以下组成：编码V

抗体的重组表达

如本文所述，多种表达系统可用于表达本发明的免疫细胞因子。一方面，这样的表达系统表示可以产生目的编码序列并随后纯化的载体，也表示当用合适的核苷酸编码序列瞬时转染时可以原位表达本发明抗体的细胞。

本发明提供了包含本文所述的多核苷酸的载体。在一个实施方案中，载体包含编码本发明的免疫细胞因子的重链的多核苷酸，其中所述重链与可裂解肽连接子和细胞因子融合或不融合。在另一个实施方案中，所述多核苷酸编码本发明的免疫细胞因子的轻链，其中所述轻链与可裂解肽连接子和细胞因子融合或不融合。本发明还提供了包含编码融合蛋白、修饰的抗体、抗体片段及其探针的多核苷酸分子的载体。

为了表达本文公开的免疫细胞因子的重链和/或轻链，将编码所述重链和/或轻链的多核苷酸插入表达载体，使得基因可操作地连接至转录和翻译序列。

“可操作地连接”的序列包括与目的基因连续的表达控制序列，和以反式或远距离作用来控制目的基因的表达控制序列。如本文所用，术语“表达控制序列”是指实现与它们连接的编码序列的表达和加工所必需的多核苷酸序列。表达控制序列包括适当的转录起始、终止、启动子和增强子序列；有效的RNA处理信号，例如剪接和聚腺苷酸化信号；稳定细胞质mRNA的序列；增强翻译效率的序列(即Kozak共有序列)；增强蛋白质稳定性的序列；以及在需要时，增强蛋白质分泌的序列。这种控制序列的性质因宿主生物而不同；在原核生物中，这种控制序列通常包括启动子、核糖体结合位点和转录终止序列；在真核生物中，这种控制序列通常包括启动子和转录终止序列。术语“控制序列”旨在至少包括其存在对于表达和加工必不可少的所有组分，并且还可以包括其存在为有利的其他组分，例如前导序列和融合伴侣序列。

如本文所用，术语“载体”旨在指能够转运已与其连接的另一核酸的核酸分子。载体的一种类型为“质粒”，其是指环状双链DNA环，其中可以连接其他DNA区段。载体的另一种类型为病毒载体，其中可以将其他DNA区段连接到病毒基因组中。某些载体能够在引入它们的宿主细胞中自主复制(例如具有细菌复制起点的细菌载体和附加体型哺乳动物载体)。在导入宿主细胞后，其他载体(例如非附加体型的哺乳动物载体)可以整合到宿主细胞的基因组中，从而与宿主基因组一起复制。

某些载体能够指导与其可操作地连接的基因的表达。这样的载体在本文中称为“重组表达载体”(或简称为“表达载体”)。通常，在重组DNA技术中有用的表达载体为质粒形式。在本说明书中，“质粒”和“载体”可以互换使用，因为质粒是最常用的载体形式。但是，本发明旨在包括表达载体的形式，例如细菌质粒、YAC、粘粒、逆转录病毒、EBV衍生的附加体、以及本领域技术人员会知道可方便地确保本发明抗体的重链和/或轻链的表达的所有其他载体。技术人员将认识到，可以将编码重链和轻链的多核苷酸克隆到不同的载体或相同的载体中。在一个优选的实施方案中，将所述多核苷酸克隆到两个载体中。

本发明的多核苷酸和包含这些分子的载体可以用于转化合适的宿主细胞。如本文所用，术语“宿主细胞”旨在指已导入重组表达载体以表达本发明免疫细胞因子的细胞。应当理解的是，这些术语不仅旨在指特定的对象细胞，而且还指所述细胞的后代。因为由于突变或环境影响，某些修饰可能在后代中发生，所以这样的后代实际上可能与亲本细胞不同，但仍包括在本文所用的术语“宿主细胞”的范围内。

可以通过将多核苷酸引入细胞宿主的任何已知方法进行转化。这样的方法是本领域技术人员众所周知的，并且包括葡聚糖介导的转化、磷酸钙沉淀、聚乙烯介导的转染、原生质体融合、电穿孔、将多核苷酸包封到脂质体中、基因枪注射以及将DNA直接显微注射到核中。

宿主细胞可以与两个或更多个表达载体共转染，所述载体包括表达本发明蛋白的载体。特别地，其他表达载体可以编码参与翻译后修饰(例如糖基化)的酶。例如，可以用编码如上所述的免疫细胞因子的第一载体和编码糖基转移酶多肽的第二载体转染宿主细胞。可选地，可以用如上所述的编码免疫细胞因子的第一载体、编码糖基转移酶的第二载体和编码另一糖基转移酶的第三载体转化宿主细胞。哺乳动物细胞通常用于表达重组治疗性免疫球蛋白，特别是用于表达完整的重组抗体。例如，哺乳动物细胞(例如HEK293或CHO细胞)与含有表达信号的载体(例如携带来自人巨细胞病毒的主要中间早期基因启动子元件的表达信号)一起，是表达本发明免疫细胞因子的有效系统(Foecking等人,1986,Gene 45：101；Cockett等人,1990,Bio/Technology 8：2)。

还可能选择一种宿主细胞，其以所需的特定方式调节插入序列的表达，或修饰和加工基因产物。蛋白质产物的这种修饰(例如糖基化)和加工对于蛋白质的功能可能是重要的。不同的宿主细胞具有蛋白质和基因产物的翻译后加工和修饰的特征和特定机制。选择适当的细胞系或宿主系统以确保表达的目的抗体的正确修饰和加工。因此，可以使用真核宿主细胞，其具有用于适当加工初级转录物和糖基化基因产物的细胞机制。此类哺乳动物宿主细胞包括但不限于CHO、COS、HEK293、NS/0、BHK、Y2/0、3T3或骨髓瘤细胞(所有这些细胞系均可从公共保藏处获得，例如法国巴黎的国立微生物培养物保藏中心，或美国弗吉尼亚州马纳萨斯的美国典型培养物保藏中心)。

为了长期、高产量地生产重组蛋白，优选稳定表达。在本发明的一个实施方案中，可以工程化稳定表达免疫细胞因子的细胞系。在合适的表达调控元件(包括启动子、增强子、转录终止子、聚腺苷酸化位点和本领域技术人员已知的其他适当序列，以及筛选标记)的控制下用DNA转化宿主细胞，而不使用包含病毒复制起点的表达载体。引入外源DNA后，可以使工程细胞在富集介质中生长一至二天，然后移至选择性介质中。重组质粒上的筛选标记赋予对筛选的抗性，并使细胞将质粒稳定整合到染色体中并扩增成细胞系。用于构建稳定细胞系的其他方法是本领域已知的。特别地，已经开发了用于特定位点整合的方法。根据这些方法，在合适的表达调控元件(包括启动子、增强子、转录终止子、聚腺苷酸化位点和其他合适的序列)的控制下，将转化的DNA整合到宿主细胞基因组中先前已被裂解的特定靶位点上(Moele等人,Proc.Natl.Acad.Sci.U.S.A.,104(9)：3055-3060；US 5,792,632；US 5,830,729；US 6,238,924；WO 2009/054985；WO 03/025183；WO 2004/067753，其均通过引用并入本文)。

可以使用许多选择系统，包括但不限于单纯疱疹病毒胸苷激酶(Wigler等人,Cell11：223,1977)、次黄嘌呤-鸟嘌呤磷酸核糖基转移酶(Szybalska等人,Proc Natl Acad SciUSA 48：202,1992)、在甲硫氨酸亚磺酰亚胺存在下用谷氨酸合酶选择(Adv Drug Del Rev,58：671,2006，及Lonza集团有限公司的网站或文献)、及腺嘌呤磷酸核糖基转移酶(Lowy等人，Cell 22：817,1980)基因，其分别位于tk、hgprt或aprt细胞中。同样，抗代谢物抗性可以用作选择以下基因的基础：dhfr，其赋予对甲氨蝶呤的抗性(Wigler等人,Proc Natl AcadSci USA 77：357,1980)；gpt，其赋予对麦考酚酸的抗性(Mulligan等人,Proc Natl AcadSci USA 78：2072,1981)；neo，其赋予对氨基糖苷G-418的抗性(Wu等人,Biotherapy 3：87,1991)；及hygro，其赋予对潮霉素的抗性(Santerre等人，Gene 30：147,1984)。重组DNA技术领域中已知的方法可以常规地用于选择所需的重组克隆，这样的方法例如在Ausubel等人编辑的《Current Protocols in Molecular Biology》，John Wiley&Sons出版社(1993)中进行了描述。免疫细胞因子的表达水平可以通过载体扩增来增加。当表达抗体的载体系统中的标记是可扩增的时，培养物中存在的抑制剂水平的增加将增加标记基因的拷贝数。由于扩增的区域与编码目的免疫细胞因子的基因相关，因此所述免疫细胞因子的产生也将增加(Crouse等人,Mol Cell Biol 3：257,1983)。存在表达本发明基因的替代方法，并且是本领域技术人员已知的。例如，修饰的锌指蛋白可以经工程化改造，而使其能够结合本发明基因上游的表达调控元件；所述工程化的锌指蛋白(ZFN)在本发明宿主细胞中的表达导致蛋白质产量的增加(参见例如Reik等人,Biotechnol.Bioeng.,97(5),1180-1189,2006)。此外，ZFN可以刺激DNA整合到预定的基因组位置，导致高效的位点特异性的基因添加(Moehle等人,Proc Natl Acad Sci USA 104：3055,2007)。

本发明的免疫细胞因子可以通过在表达所需抗体所必需的培养条件下培养转化的宿主细胞的培养物来制备。然后可以从培养介质或细胞提取物中纯化得到的表达的免疫细胞因子。可以从培养物上清液中回收可溶性形式的免疫细胞因子。然后可以通过本领域已知的用于纯化免疫球蛋白分子的任何方法来纯化，例如通过色谱法(例如离子交换、亲和力、特别是通过蛋白A对Fc的亲和力等)、离心、差异溶解度或任何其他蛋白质纯化标准技术。合适的纯化方法对本领域普通技术人员将是显而易见的。

当递送至肿瘤部位时，本文提供的免疫细胞因子被裂解，从而释放具有抗肿瘤活性的细胞因子，例如IL-15。另外，一些抗体部分能够诱导ADCC和/或CDC应答和/或具有固有的抗肿瘤活性。因此，本领域技术人员将理解，本文提供的免疫细胞因子可用于治疗转移性肿瘤和疾病，例如癌症。

术语“治疗”或“处理”是指给予或施用如本文所述的组合物，其使用量、方式和/或模式可有效改善与病症有关的病症、症状、或与疾病或与预防疾病进展或恶化(包括由疾病引起的继发性损伤)相关的参数，所述改善达到统计学上显著的程度或本领域技术人员可检测的程度。

本发明的另一方面涉及本文所述的免疫细胞因子的药物组合物。

除了本发明的免疫细胞因子外，本发明的药物组合物还可包含各种稀释剂、填充剂、盐、缓冲剂、稳定剂、增溶剂和本领域公知的其他材料。

如本文所用，“药学上可接受的载体”包括生理相容的任何和所有溶剂、缓冲剂、盐溶液、分散介质、包衣、抗菌和抗真菌剂、等渗剂和吸收延迟剂等。可以基于预期的施用途径选择载体的类型。在各种实施方案中，载体适合于静脉内、腹腔内、皮下、肌内、局部、透皮或口服施用。药学上可接受的载体包括无菌水溶液或分散液以及用于临时制备无菌注射溶液或分散液的无菌粉末。用于药物活性物质的介质和试剂的使用是本领域众所周知的。如下所述，还可以将其他活性化合物掺入组合物中，例如抗癌剂和/或抗血管生成剂；特别地，所述其他活性化合物可以为抗血管生成剂、化学治疗剂或低分子量剂。用于静脉内输注的典型药物组合物可以制成包含250mL无菌林格氏液和100mg组合物。制备肠胃外施用的化合物的实际方法对于本领域技术人员来说是已知的或显而易见的，并且在例如《Remington'sPharmaceutical Science》第17版，宾夕法尼亚州伊斯顿Mack出版公司(1985)及其第18版和第19版中进行了详细描述，在此引用作为参考。

存在于组合物中的免疫细胞因子优选以有效量配制。“有效量”是指在所需剂量和时间段内有效达到所需结果的量，所述结果例如诱导肿瘤细胞凋亡。“治疗有效量”是指足以影响特定疾病状态的治疗过程的量。治疗有效量也是其中药物的治疗有益作用远大于任何毒性或有害作用的量。

对于治疗应用，将免疫细胞因子以药学上可接受的剂量形式(例如以上讨论的那些)施用至哺乳动物(优选人)，包括可以静脉推注(bolus)或在一段时间内连续输注、肌肉内、腹腔内、脊髓内、皮下、关节内、滑膜内、鞘内、口服、局部、或吸入途径施用至人的剂型。免疫细胞因子还可以通过瘤内、瘤周、病灶内或病灶周围途径适当地施用，以发挥局部及全身治疗作用。腹腔内途径有望在例如卵巢肿瘤的治疗中特别有用。可以调整剂量方案以提供最佳响应。例如，可以施用单次推注，可以随时间施用数个分开的剂量，或者可以成比例地减少或增加剂量。制备制剂领域的本领域技术人员可以容易地选择适当的形式和施用方式，取决于所选择的产品的特定特征、要治疗的疾病或病症、疾病或病症的阶段及其他相关情况。

可以将本发明的组合物施用受试者，以影响受试者的细胞生长活性。如本文所用，术语“受试者”旨在包括其中可诱导凋亡的活的生物体，并且具体包括哺乳动物，例如兔、狗、猫、小鼠、大鼠、猴的转基因物种，优选人。

免疫细胞因子预防或治疗癌症的有效性改善，可通过连续施用所述免疫细胞因子或与对那些目的有效的其他药物联合施用来进行，所述其他药物例如肿瘤坏死因子(TNF)、能够抑制或中和酸性或碱性成纤维细胞生长因子(FGF)的血管生成活性的拮抗剂、血小板源性生长因子(PDGF)、或肝细胞生长因子(HGF)、能够抑制或中和组织因子蛋白C或蛋白S的凝血活性的拮抗剂(参见WO91/01753)、拮抗剂例如能够与HER2受体结合的抗体(参见US 5,772,997)、或一种或多种常规治疗剂(例如烷化剂、叶酸拮抗剂、核酸代谢的抗代谢物、抗生素、嘧啶类似物、5-氟尿嘧啶、顺铂、嘌呤核苷、胺、氨基酸、三唑核苷或皮质类固醇)。

另外，本发明的药物组合物还可以包含其他试剂，其能够调节免疫细胞(尤其是T细胞或单核细胞)的激活和/或功能。特别地，本发明的药物组合物可以进一步包含治疗有效量的共抑制分子拮抗剂。在一些实施方案中，所述共抑制分子选自CD86、CD80、PDL-1、PDL-2、CTLA-4、PD1、LAG3、BTNL2、B7-H3、B7-H4、嗜乳脂蛋白、CD48、CD244、TIM-3、CD200R、CD200、CD160、BTLA、HVEM、LAIR1、TIM1、半乳糖凝集素9、TIM3、CD48、2B4、CD155、CD112、CD113和TIGIT。所述共抑制分子拮抗剂包括抗共抑制分子的抗体。已经认识到，其他共抑制分子拮抗剂是本领域众所周知的，例如在Mercier等人,Frontiers in Immunology,6：418(2015)，Kyi等人,FEBS Letters,588：368-376(2014)和Pardoll,Nature Reviews,12：252-264(2012)中所描述的那些。在一些其他实施方案中，本文所述的药物组合物还包含治疗有效量的共刺激分子激动剂。在一些实施方案中，所述共刺激分子选自CD154、TNFRSF25、GITR、4-1BB、OX40、CD27、TMIGD2、ICOS、CD28、CD40、TL1A、GITRL、41BBL、OX40L、CD70、HHLA2、ICOSL、细胞因子、LIGHT、HVEM、CD30、CD30L、B7-H2、CD80、CD86、CD40L、TIM4、TIM1、SLAM、CD48、CD58、CD155、CD112、DR3、GITR、CD2、及CD226。所述共刺激分子激动剂包括抗共刺激分子的激动抗体。已经认识到，共刺激分子激动剂是本领域众所周知的，例如在Mercier等人,Frontiers in Immunology,6：418(2015)，Kyi等人,FEBS Letters,588：368-376(2014)和Capece等人,J.Biomed.Biotechnol.2012：926321,17页(2012)中所描述的那些。

在本发明的另一方面，该施用与治疗有效量的化学治疗剂例如紫杉醇(taxol,paclitaxel)或泰索帝(taxotere)(多西紫杉醇)的施用进行联合。

化学治疗剂包括但不限于：抗微管剂，例如二萜和长春花生物碱；铂配合物；烷基化剂，例如氮芥、氧杂氮杂膦、烷基磺酸盐/酯、亚硝基脲和三氮烯；抗生素，例如蒽环素、放线菌素和博来霉素；拓扑异构酶II抑制剂，例如表鬼臼毒素；抗代谢物，例如嘌呤和嘧啶类似物以及抗叶酸化合物；拓扑异构酶I抑制剂，例如喜树碱；激素和激素类似物；信号转导途径抑制剂；非受体酪氨酸激酶血管生成抑制剂；免疫治疗剂；促凋亡剂；及细胞周期信号抑制剂。另外，本发明的方法可以与另一种抗癌治疗、抗血管生成剂或化学治疗剂或放射疗法组合。一个优选的例子为多西紫杉醇或泰索帝。其他实例包括：吉西他滨、顺铂二萜和长春花碱生物碱、紫杉醇、长春碱、长春新碱和长春瑞滨、卡铂、环磷酰胺、美法仑和苯丁酸氮芥、白消安、卡莫司汀、达卡巴嗪、环磷酰胺、美法仑、苯丁酸氮芥、白消安、卡莫司汀、达卡巴嗪、抗肿瘤药物，包括但不限于放线菌素(例如更生霉素)、蒽环霉素(例如柔红霉素和多柔比星)、博来霉素、表鬼臼毒素、依托泊苷和替尼泊苷；抗代谢肿瘤药物，5-氟尿嘧啶、甲氨蝶呤、阿糖胞苷、巯基嘌呤、硫代鸟嘌呤、喜树碱、伊立替康盐酸盐和托泊替康盐酸盐。

也可以选择多种不同的化学治疗剂或抗癌多肽。信息源例如www.clinicaltrials.gov、www.ncbi.nlm.nih和www.drugs.com包括可以选择的多肽和试剂的参考。

本发明的免疫细胞因子和药物组合物尤其可用于治疗或预防几种类型的癌症。

因此，本发明的另一方面涉及用于治疗癌症的本文所述的免疫细胞因子。

本发明还涉及用于治疗癌症的包含本文所述的免疫细胞因子的药物组合物。

可以用本发明的免疫细胞因子治疗的癌症为表达了被所述免疫细胞因子中抗体部分识别的抗原的癌症。这些癌症包括(但不限于)以下：癌和腺癌，包括膀胱癌、乳腺癌、结肠癌、头颈癌、前列腺癌、肾癌、肝癌、肺癌、卵巢癌、胰腺癌、胃癌、宫颈癌、甲状腺癌和皮肤癌，包括鳞状细胞癌；淋巴系的造血肿瘤，包括多发性骨髓瘤、白血病、急性和慢性淋巴细胞性(或淋巴样)白血病、急性和慢性淋巴细胞性白血病、B细胞淋巴瘤、T细胞淋巴瘤、非霍奇金淋巴瘤(例如Burkitt淋巴瘤)；骨髓系的造血肿瘤，包括急性和慢性骨髓性(髓样或髓细胞性)白血病和早幼粒细胞性白血病；间充质起源的肿瘤，包括纤维肉瘤、骨肉瘤和横纹肌肉瘤；中枢和周围神经系统的肿瘤，包括星形细胞瘤、神经母细胞瘤、神经胶质瘤和神经鞘瘤；和其他肿瘤，包括黑素瘤、畸胎癌、色素性干皮病、角棘皮瘤和精原细胞瘤、以及尚待确定表达所述抗原的其他癌症。表达所述免疫细胞因子的抗体部分识别的抗原的癌症，在本文中是指相对于正常成体细胞上所述抗原的表达水平而言，高表达所述抗原的癌症。

上述其他试剂(例如抗血管生成剂或化学治疗剂)可以存在于正在施用或可以单独施用的组合物中。在本发明的一个方面，施用是与另一种活性成分一起，同时进行、分别进行或随时间依次进行。当同时进行施用时，可以将两种活性成分组合在包含两种组合物的单一药物组合物中，例如片剂或凝胶胶囊。另一方面，两种活性成分无论是否同时施用，都可以存在于分开的药物组合物中。为此，组合物可以是试剂盒的形式，该试剂盒一方面包含本文所述的免疫细胞因子，另一方面包含第二活性成分，本文所述的免疫细胞因子和第二活性成分位于分开的隔室中，并且旨在同时、分别或随时间依次施用。

本发明的组合物可以特别地与化学疗法、蛋白质疗法(即使用诸如抗体或重组蛋白的治疗剂)、基因疗法、放射疗法、免疫疗法、外科手术或其的组合疗法联合用于治疗癌症。如上所述，在其他治疗策略的背景下，长期治疗与辅助治疗同样可能。

细胞因子(例如IL-15)能够刺激高度特异性的肿瘤特异性T细胞应答。例如，施用IL-15可诱导CD8 T细胞和NK细胞选择性激活和增殖，其为最适合介导抗肿瘤反应的特定细胞类型(Waldmann,J Investig Dermatol Symp Proc.16(1)：S28-30,2013)。

在一些实施方案中，免疫细胞因子可以用于调节免疫细胞功能的方法中，所述功能由所述免疫细胞因子的细胞因子部分(优选IL-15)的结合介导。优选地，所述免疫细胞为T细胞或单核细胞。这样的方法可以包括使免疫细胞(优选T细胞或单核细胞)与本文所述的免疫细胞因子接触。在一些实施方案中，用于调节免疫细胞(尤其是T细胞或单核细胞)功能的方法包括向受试者施用有效量的包含本文提供的免疫细胞因子的组合物。在一些方面，被调节的T细胞功能包括增加T细胞的激活。这种T细胞的激活可以进一步包括增加T细胞的增殖。在一些方面，被调节的单核细胞功能包括增加抗癌细胞因子的分泌。用于测定免疫应答调节的方法是本领域技术人员众所周知的，并且应理解的是，技术人员将能够容易地进行此类测定。

在一些实施方案中，免疫细胞因子或包含免疫细胞因子的组合物，包括本文所述的，可以单独使用或与另外的化合物或治疗组合使用。例如，在一些实施方案中，另外的化合物为共抑制分子拮抗剂或共刺激分子激动剂。在这样的实施方案中，联合疗法通过激活的T细胞导致免疫系统的重振或从头激活，所述效应大于单独施用一个化合物或治疗。免疫系统的这种激活将在癌症治疗中产生高度有益的生理反应。

在一些实施方案中，本文所述的方法可以包括施用治疗有效量的免疫细胞因子联合治疗有效量的共抑制分子拮抗剂。在一些实施方案中，所述共抑制分子选自CD86、CD80、PDL-1、PDL-2、CTLA-4、PD1、LAG3、BTNL2、B7-H3、B7-H4、嗜乳脂蛋白、CD48、CD244、TIM-3、CD200R、CD200、CD160、BTLA、HVEM、LAIR1、TIM1、半乳糖凝集素9、TIM3、CD48、2B4、CD155、CD112、CD113和TIGIT。所述共抑制分子的拮抗剂包括抗共抑制分子的抗体。已经认识到，其他共抑制分子拮抗剂是本领域众所周知的，例如在Mercier等人,Frontiers inImmunology,6：418(2015)，Kyi等人,FEBS Letters,588：368-376(2014)和Pardoll,NatureReviews,12：252-264(2012)中所描述的那些。根据该实施方案，本发明涉及一种用于治疗如上所述的癌症的免疫细胞因子，所述用途还包括施用共抑制分子拮抗剂，其中所述共抑制分子选自：CD86、CD80、PDL-1、PDL-2、CTLA-4、PD1、LAG3、BTNL2、B7-H3、B7-H4、嗜乳脂蛋白、CD48、CD244、TIM-3、CD200R、CD200、CD160、BTLA、HVEM、LAIR1、TIM1、半乳凝素9、TIM3、CD48、2B4、CD155、CD112、CD113和TIGIT。

在一些实施方案中，本文所述的方法可以包括施用治疗有效量的免疫细胞因子联合治疗有效量的共刺激分子激动剂。在一些实施方案中，所述共刺激分子选自CD154、TNFRSF25、GITR、4-1BB、OX40、CD27、TMIGD2、ICOS、CD28、CD40、TL1A、GITRL、41BBL、OX40L、CD70、HHLA2、ICOSL、细胞因子、LIGHT、HVEM、CD30、CD30L、B7-H2、CD80、CD86、CD40L、TIM4、TIM1、SLAM、CD48、CD58、CD155、CD112、DR3、GITR、CD2、及CD226。所述共刺激分子激动剂包括抗共刺激分子的激动抗体。已经认识到，共刺激分子激动剂是本领域众所周知的，例如在Mercier等人,Frontiers in Immunology,6：418(2015)，Kyi等人,FEBS Letters,588：368-376(2014)和Capece等人，J.Biomed.Biotechnol.2012：926321,17页(2012)中所描述的那些。根据该实施方案，本发明涉及一种用于治疗如上所述的癌症的免疫细胞因子，所述用途还包括施用共刺激分子激动剂，其中所述共刺激分子选自：CD154、TNFRSF25、GITR、4-1BB、OX40、CD27、TMIGD2、ICOS、CD28、CD40、TL1A、GITRL、41BBL、OX40L、CD70、HHLA2、ICOSL、细胞因子、LIGHT、HVEM、CD30、CD30L、B7-H2、CD80、CD86、CD40L、TIM4、TIM1、SLAM、CD48、CD58、CD155、CD112、DR3、GITR、CD2、及CD226。

以下实施例仅是本发明范围和本公开内容的示例。本领域技术人员可以在不脱离本发明范围的情况下设计和构造对以下所列实施例的多种修改。

具体实施方式

表1：抗体可变区

表2：恒定区

由人IgG1而非人IgG1ΔK获得相似结果。

表3：细胞因子

表4：连接子

表5：克隆位点(对于某些

表6：ICC

表7：ICC-完整构建体

将编码整个ICC的序列(参阅1.序列)通过使用HindIII/BamHI限制性位点，包括信号肽(Thermo Fisher Scientific)，克隆到pCDNA3.4载体中。

图2解释了连接子-细胞因子在Ab上的位置(即融合位点)。

所有纯化的ICC通过SDS-PAGE在非还原和加热条件下、及还原和加热条件下表征。c9G4-PVGLIG-hIL15、NHS76-PVGLIG-hIL15和H16/L16-PVGLIG-hIL15的SDS-PAGE迁移如图22所示。从SDS PAGE推导出的表观ICC分子量与理论计算所期望的一致。每个ICC至少有80％为完整的ICC(H2L2)。通过尺寸排阻色谱法确定ICC的单体含量。

ICC的完整性还通过糖基化ICC和IdeS消化后的去糖基化ICC的LC-MS进行了验证。反相分离在与Synapt G2si质谱仪连接的超高效液相色谱(UHPLC)系统(Acquity UPLC H-Class Bio系统，Waters)上进行，使用

对于c9G4PVGLIG-IL15、c9G4PVGLIG-hINFa、c9G4PVGLIG-hCCL4，Fc区的去糖基化是通过将ICC溶液与

Fc/2亚基片段是将ICC溶液与IdeS酶(

在MS色谱峰上，提取m/z质谱图，并在m/z质谱图去卷积后确定质量，并将计算出的去糖基化ICC和去糖基化fc/2的质量与实验质量进行比较。

去糖基化的c9G4-PVGLIG-hIL15、NHS76-PVGLIG-hIL15和H16/L16-PVGLIG-hIL15免疫细胞因子的LS-MS分析如图2和图3所示。其模式与预期完全形成的ICC应有的一致，如表8所示：

表8：C-末端HC ICC：ICC和Fc/2的实验质量与理论质量的比较

整体实验质量证实了ICC构建体(2细胞因子/抗体)，而Fc/2+细胞因子质量证实了细胞因子的位置。

对其他ICC进行了类似的实验，并获得了类似的结论。

来自文献的五个连接子被评估为MMP-9的底物时，其与PDL1 Ab序列上的四个不同位点融合：N-末端HC、N-末端LC、C-末端HC和C-末端LC。“GIVGPL”连接子被描述为不可裂解的(Chau等人,Bioconjug Chem(2004)15(4)：931-41)并用作阴性对照。各构建体和实验结果的总结示于图7。

将2μg纯化的ICC在40ng重组MMP-9或MMP-2(摩尔比25：1)存在下，于20μL总体积的测试缓冲液(含有20mM Tris pH 7.5、10mM CaCl

然后通过SDS-PAGE评估裂解效率。将20μL样品连同蛋白质标准品(PrecisionPlus Protein Standards，未染色，BioRad)一起，在Tris-甘氨酸-SDS缓冲液(BioRad)中的Criterion TGX 4-15％免染胶(BioRad)上以300V运行20分钟。然后使用ChemiDoc TouchImager(BioRad)显示蛋白条带。

3.2.1抗PDL1抗体与两种不同分子的HC或LC C-末端融合物中不同连接子的可裂解性：IL2和

结果如图4所示。

3.2.2抗PDL1抗体与两种不同分子的HC或LC N-末端融合物中不同连接子的可裂解性：IL2和

结果如图5所示。

3.2.3不同的基于c9G4的ICC和H16/L16-IL15和HHS76-IL15 ICC被人和鼠MMP-9/2(HC C-末端融合蛋白和PVGLIG连接子)的可裂解性。结果如图6所示。

注1：高水平的蛋白质糖基化削弱了IL15和IFNa裂解后的可视化。裂解前样品的去糖基化使得释放的细胞因子可视化，表明该蛋白未被MMP-9/2水解(数据未显示)。

图4、图5和图6的结果汇总在图7中。鉴定了两个融合位点，其允许被蛋白酶正确地裂解：Ab重链的N-末端和C-末端。

另一方面，当在Ab轻链的N-末端或C-末端融合时，MMP-9连接子仅被MMP-2或MMP-9弱裂解。

鼠和人MMP-9和MMP-2均裂解了所有5种c9G4融合蛋白(图6)。

将ICC以100μg/ml的浓度掺入缓冲液(50mM Tris pH 7.5、150mM NaCl、20mMCaCl

通过使用包被4μg CaptureSelect

反相分离在与Synapt G2si质谱仪连接的超高效液相色谱(UHPLC)系统(AcquityUPLC H-Class Bio系统，Waters)上进行，使用

将还原后的样品用洗脱液A稀释至体积比(volume to volume)，然后注入加热到80℃的PLRP-S色谱柱中，流速为0.5mL/min。用水作为洗脱液A、乙腈作为洗脱液B(均含有0.05％TFA)进行洗脱。梯度条件在5％B下保持5分钟，在45分钟内升至70％，在2分钟内升至95％。

对于MS色谱图的每个峰，提取m/z谱图并在m/z谱图去卷积后确定质量。

4.2结果

在没有MMP-9的情况下，抗

如实施例2所述，通过LC/MS检查了在MMP-9存在下的裂解情况。结果示于图8A和图8B。

在MMP-9存在下，在37℃下24小时后，100％的抗体抗

结果如图9所示。有或没有MMP-9掺入时，在小鼠肝素血浆、小鼠、食蟹猴、人血清和小鼠肝素全血中的T0和T24处均获得相似的图谱。MMP-9在所有测试的生物液体中均被抑制，LC-MS谱图示例如图9所示。

总之，在肝素血浆、血清和肝素全血中无法监测体外抗体抗

为了评估和比较hIL15的生物学活性，将其连接至ICC或被MMP9裂解后，进行了IL-15受体二聚化测试。活性形式的IL15通过其两个受体亚基IL2Rβ/IL2Rγ的二聚化发出信号。

5.1材料与方法

重组的人重组前-MMP-9购自R&D Systems，并在缓冲液(含50mM Tris、150mMNaCl、10mM CaCl

评估了三种基于IL15的免疫细胞因子的活性：NHS76-PVGLIG-hIL15、H16/L16-PVGLIG-hIL15和c9G4-PVGLIG-hIL15。重组hIL15和hMMP-9的作用也用相同的方法评估。将所有样品在重组hMMP-9存在(+MMP-9)或不存在(-MMP-9)的情况下，在测定缓冲液(含有50mM Tris、150mM NaCl、10mM CaCl

5.2结果

三种被测ICC的hIL15诱导的受体二聚化结果随hIL15浓度的变化示于图10。

每种化合物或ICC的EC

如图10和表9所示，hIL15和hIL15+MMP-9高度诱导受体二聚化，因此验证了本实验。相反，如预期的那样，单独的MMP-9对IL2Rβ/IL2Rγ二聚化没有影响。

在没有MMP-9的情况下，三个评估的ICC对受体二聚化没有显著的活性。另一方面，用MMP-9预处理相同分子可通过IL15诱导的受体二聚化以剂量依赖性方式恢复活性。

因此，从这些实验清楚地表明，当与所述三种抗体连接时，IL-15表现出高度减弱的活性。通过MMP-9裂解ICC后，IL15以其活性形式被释放，并再次能够履行其生物学活性。

表9：Ab结合的hIL15与游离的hIL15的EC

6.1材料与方法

六周大的具有免疫能力的BALB/c小鼠用于所有体内评估。其安置在无菌的顶部为过滤器的笼子里，保持无菌状态，并根据法国和欧洲的指导方针进行操作。

选择表达PDL1的鼠肾癌细胞系RENCA(ATCC：CRL-2947)进行体内评估。在D0将0.5×10

施用后0、3、6、24和48小时处死动物，并通过心脏穿刺将血液样品收集在肝素化钠管中。在1500g、4℃下将血液样品离心15分钟，然后在96孔U型底的板中收集血浆。分析前将血浆样品冷冻(-80℃)。

对肿瘤取样并在液氮中速冻，并储存在-80℃。

在评估之前，将肿瘤“重悬”在50mM Tris-HCl缓冲液中，其含有150mM NaCl、0.5％DOC、1％Igepal、1％Triton X100、抑制剂混合物(裂解缓冲液)，每毫升缓冲液200mg肿瘤。

肿瘤通过Minilys破坏，在钢球存在下5000rpm搅拌15秒、3个周期。在2个周期之间，将管在冰上保持2分钟。将溶液置于2mL锥形Eppendorf管中，并在4℃以11500g离心10分钟。回收上清液，即细胞裂解物。根据制造商的说明，通过二辛可宁酸(bicinchoninicacid)测定法测量其蛋白质含量。

通过蛋白质印迹分析血清样品和肿瘤样品。对于每个体内实验，一式三份进行蛋白质印迹。

简而言之，将待分析的样品加载到预制的4-15％聚丙烯酰胺凝胶上，血浆样品以0.1μL的速度加载，肿瘤裂解物加载2μL。样品在加热、非还原的条件下迁移。然后将蛋白质转移到硝酸纤维素膜上，使用

用Image Lab(Bio-Rad)进行蛋白质印迹的光密度分析。

使用GraphPad Prism6进行统计分析。

6.2结果

评估了ICC在小鼠血清中的稳定性随时间的变化。ICC注射后3h、6h和24h通过蛋白质印迹获得的结果示于图11。对于每种测试的ICC构建体，循环ICC的数量均随时间显著减少。不受理论的束缚，最容易通过在肿瘤中ICC的隔离(sequestration)和/或天然的体内Ab降解来解释该结果。

对蛋白质印迹进行光密度分析。图形表示(图12)显示了未裂解的ICC的百分比，计算方法如下：[未裂解ICC/(裂解+未裂解)ICC的强度]×100。两种ICC模型在血液循环中的表现相似，即在逐渐失去其

总体而言，循环c9G4 ICC和PDL1 ICC之间没有显著差异。

评估了ICC在小鼠肿瘤样品中的稳定性随时间的变化。ICC注射后3h、6h和24h通过蛋白质印迹获得的结果示于图14。

对蛋白质印记进行了光密度分析，图形表示(图15)显示了如前所述计算的未裂解ICC的百分比。

两种模型ICC的表现相似，即

c9G4 ICC的裂解较慢，注射后6小时，约20％的ICC在肿瘤中裂解。然而，在注射后24小时，存在于肿瘤中的总ICC的80％以上被裂解。对肿瘤隔离的ICC进行统计分析。通过在每个时间点获得的未裂解ICC和裂解的ICC的信号相加来确定总的肿瘤隔离的ICC的量。

肿瘤中存在的总ICC(裂解和未裂解的)随时间变化的定量分析示于图16。

总体而言，与对照c9G4 ICC相比，PDL1-ICC在RENCA肿瘤中的隔离度更高。

这些结果表明，肿瘤特异性ICC能够在肿瘤环境中快速积累。积累后，ICC被肿瘤特异性蛋白酶裂解，在肿瘤环境中特异性释放细胞因子。

将两个ICC在RENCA移植小鼠血浆和肿瘤样品中的表现随时间的变化进行比较。

图17表明，无论抗体部分的性质如何，RENCA肿瘤中的ICC均比小鼠血浆中的ICC裂解程度显著更高。

7.1材料与方法

重组人IL15购自Peprotech。

测试了下列ICC构建体：

·NHS76-PVGLIG-IL15；

·H16/L16-PVGLIG-IL15；及

·c9G4-PVGLIG-IL15

加入了下列对照：hIL15、MMP9、hIL15+MMP9及单个抗体。

评估了三种基于IL15的免疫细胞因子(即NHS76-IL15、H16/L16-IL15和c9G4-IL15)的活性，评估其激活鼠和人CD3

根据制造商的说明，通过负选择从balb/c小鼠(Charles River)的脾脏中使用鼠泛T细胞分离试剂盒II(Miltenyi，130-095-130)分离鼠CD3

从健康供体的细胞分离(cytapheresis)环的新鲜血液中通过密度梯度离心分离人外周血单个核细胞(PBMC)。然后，使用人泛T细胞分离试剂盒II(Miltenyi，130-096-535)通过从PBMC中进行负选择来纯化CD3 T细胞。

将鼠和人CD3

然后将其在存在或不存在MMP9的条件下，以100和200ng/ml的人IL-15作为阳性对照进行处理，或以6μg/ml的不同ICC构建体(等效于200ng/ml的IL-15)处理。将细胞在37℃、5％CO

处理6天后，通过CD25和CD69表面标志物的表达监测CD3

对于每个鼠和人T细胞激活实验，一式三份进行分析；测试了三名健康人供体。

使用Prism 7.01软件(GraphPad)进行数据分析。

7.2结果

将ICC激活鼠T细胞的能力与对照进行比较。通过CD69或CD25的T细胞表达来测量T细胞激活。图23A和图23B显示NHS76-PVGLIG-IL15和c9G4-PVLGIG-IL15在存在MMP-9、而非不存在MMP-9的情况下诱导CD69和CD25的表达。因此，当ICC未裂解时，未观察到鼠T细胞的激活。还测试了阴性对照(单独的抗体、单独的MMP-9、单独的缓冲液)或阳性对照(单独的IL15和IL15+MMP-9)，以确认实验的有效性。

如图23B和图23C所示，裂解的H16/L16-PVGLIG-IL15(但不是ICC的未裂解形式)也能够以类似方式激活鼠T细胞的激活。

用来自健康供体的人T细胞进行了相同的实验。通过CD69或CD25的T细胞表达来测量T细胞激活。NHS76-PVGLIG-IL15、c9G4-PVLGIG-IL15和H16L16-PVGLIG-IL15在存在MMP-9、而非不存在MMP-9的情况下诱导CD69和CD25的表达(图24A、图24B、图24C和图24D)。因此，当ICC未裂解时，未观察到人T细胞的激活。还测试了阴性对照(单独的抗体、单独的MMP-9、单独的缓冲液)或阳性对照(单独的IL15和IL15+MMP-9)，以确认实验的有效性。

在裂解(但不是未裂解)ICC存在下的这种T细胞激活，也对来自两个不同供体的人T细胞IFN-γ的分泌进行了测量。

为了评估和比较作为ICC的一部分或MMP9裂解后的hIL36γ的生物学活性，通过测量人表皮癌A431细胞培养物中CXCL8的产生进行了IL36γ活性测定。所述测定是基于一项发现：IL36γ信号通路可以引起CXCL8分泌。

8.1材料与方法

重组的人重组前-MMP-9购自R&D Systems，并在缓冲液(含50mM Tris、150mMNaCl、10mM CaCl

为了评估基于IL36γ的免疫细胞因子(H16/L16-PVGLIG-IL-36)的活性，将鳞状癌A431细胞与ICC或不同的对照分子孵育24小时。在重组hMMP-9存在(+MMP-9)或不存在(-MMP-9)的情况下，在测定缓冲液(含有50mM Tris、150mM NaCl、10mM CaCl

8.2结果

ICC和对照在A431培养物中的hIL8产生的结果如图26所示。数据以对应于hIL8表达水平的O.D(450nm)随hIL36γ浓度的变化表示。

如图26所示，rhIL36γ和rhIL36γ+MMP-9高度诱导A431细胞产生hIL8，从而验证了本实验程式。另一方面，如预期的，单独的MMP-9或抗体H16/L16(+/-MMP9)都不会对hIL8的产生有任何影响。

在没有MMP-9的情况下，H16/L16-PVGLIG-hIL36γICC对hIL8的产生没有显著的活性。另一方面，如通过剂量依赖式的IL-8产生观察到的，用MMP-9预处理相同分子可导致活性恢复。

总之，使用PVGLIG将hIL36γ的C-末端连接至H16/L16抗体完全减弱了其细胞因子活性。PVGLIG肽内的MMP9裂解可将其恢复到与等效剂量游离hIL36γ细胞因子相当的水平。

为了评估和比较hIFNα2a与ICC连接时或被MMP9裂解后的生物学活性，使用基于细胞的荧光素酶报告基因生物测定法测定了IFNα2a的活性。

9.1材料与方法

重组的人重组前-MMP-9购自R&D Systems，并在缓冲液(含50mM Tris、150mMNaCl、10mM CaCl

IFNα2a活性使用GloResponse

评估了三种基于hIFNα2a的免疫细胞因子的活性：NHS76-PVGLIG-h IFNα2a、H16/L16-PVGLIG-hIFNα2a和c9G4-PVGLIG-hIFNα2a。重组hIFNα2a的作用也用相同的方法评估。将所有样品在重组hMMP-9存在(+MMP-9)或不存在(-MMP-9)的情况下，在测定缓冲液(含有50mM Tris、150mM NaCl、10mM CaCl

通过SDS-PAGE分析控制裂解效率，并立即将样品储存在-20℃直至处理。用裂解的或未裂解的ICC或对照过夜处理ISRE-luc2P/HEK293细胞。然后加入检测剂(Bio-Glo

对于H16/L16-PVGLIG-hIFNα2a，使用或不使用10μg缺乏细胞因子融合物的H16/L16抗体预处理细胞，目的是使ISRE-luc2P/HEK293细胞表面的IGF1-R结合位点饱和。

9.2结果

三种被测ICC的hIFNα2a活性测定结果随hIFNα2a浓度的变化示于图27。每种化合物或ICC的EC

如荧光素酶表达所示，hIFNα2a和hIFNα2a+MMP-9均强烈诱导信号传导反应，从而验证了本实验(图27和表10)。

在没有MMP-9的情况下，三个评估ICC中的每个：c9G4-PVGLIG-hIFNα2a(图27A)、NHS76-PVGLIG-hIFNα2a(图27B)和H16/L16-PVGLIG-hIFNα2a，用(图27C)或未用(图27D)10μg/ml H16/L16抗体预孵育细胞，与等摩尔浓度的hIFNα2a活性相比，显示出高度减弱的活性。另一方面，用MMP-9预处理相同分子可导致剂量依赖式的活性恢复。

因此，从这些实验清楚地表明，当与所述三种抗体的任一个连接时，hIFNα2a表现出减弱的活性。通过MMP-9裂解ICC后，hIFNα2a以其活性形式被释放，并再次能够履行其生物学活性。

表10：Ab结合的hIFNα2a与游离hIFNα的EC

为了评估和比较hIL15的生物学活性，将其与ICC连接或被uPA裂解后，进行了IL15活性的生物学测试(Promega IL15生物测定法)。

10.1材料与方法

重组尿激酶(＝uPA)购自Abcam：ab92767(人)。重组人IL15购自Peprotech(200-15)。

使用Promega的IL-15生物测定法(Cat.#JA2015)监测IL15的活性。其为一种生物发光的基于细胞的测定法，旨在使用STAT-5响应元件作为读数来测量IL-15的刺激或抑制作用。当IL-15与其受体结合时，受体介导的信号通路会诱导发光，所述发光可以在添加底物后检测并用发光计进行定量。

待评估的构建体在PBS中用uPA 37℃孵育24小时。

通过SDS-PAGE分析控制裂解效率，并立即将样品储存在-20℃直至处理。

用裂解的或未裂解的ICC或对照处理IL15细胞6h。然后加入检测剂(Bio-Glo

10.2结果

ICC和对照的hIL15活性的结果如图28所示。数据以对应于hIL15活性水平的相对发光(RLU)随STAT-5激活的变化表示。

rhIL15和rhIL15+uPA均高度激活发光，从而验证了实验程式。在没有uPA的情况下，H16L16-SGRSA-hIL15 ICC没有显著的IL15活性。如通过STAT-5激活所观察到的，用uPA预处理同一分子可显著恢复活性(图28)。

总之，使用SGRSA将hIL15与H16/L16抗体在C-末端进行连接，完全减弱了其活性。SGRSA肽中的uPA裂解显著恢复了IL15活性。

为了评估和比较hIFNα的生物学活性，将其与ICC连接或被uPA裂解后，进行了IFNα活性的生物学测试(GloResponse ISRE-luc2P)。

11.1材料与方法

重组人尿激酶(＝uPA)购自Abcam(ab92767)。重组人IFNα购自PBL(11101-02)。使用GloResponse ISRE-luc2P(Promega CS190701)监测IFNα活性。其为一种生物发光的基于HEK293细胞的测定法，旨在使用干扰素刺激响应元件(ISRE)来测量IFNα的刺激或抑制作用，ISRE驱动荧光素酶报道基因luc2P(萤火虫Photinus pyralis)的转录。当IFNα与其受体结合时，受体介导的信号通路会诱导发光，所述发光可以在添加底物后检测并用发光计进行定量。

待评估的构建体在PBS中用uPA 37℃孵育24小时。通过SDS-PAGE分析控制裂解效率，并立即将样品储存在-20℃直至处理。

将ISRE-luc2P/HEK293细胞用10μg抗IGF1R抗体预处理1小时，以使细胞表面的IGF1R结合位点饱和。

用裂解的或未裂解的ICC和对照过夜处理ISRE-luc2P/HEK293细胞。然后加入检测剂(Bio-Glo

11.2结果

ICC和对照的hIFNα活性的结果如图29A和图29B所示。数据以对应于hIFNα活性水平的相对发光(RLU)随ISRE激活的变化表示。

如图29A所示，rhIFNα和rhIFNα+uPA高度激活发光，因此验证了实验程式。

在没有uPA的情况下，H16/L16-SGRSA-hIFNαICC的IFNαEC

总之，使用SGRSA将hIFNα与H16/L16抗体在C-末端进行连接减弱了其活性。在SGRSA肽中的uPA裂解使其恢复到与游离hIFNα细胞因子相似的水平。

如图29B所示，rhIFNα和rhIFNα+uPA高度激活发光，因此验证了实验程式。

在没有uPA的情况下，H16/L16-PSSRRRVN-hIFNαICC的IFNαEC

总之，使用PSSRRRVN将hIFNα与H16/L16抗体在C-末端进行连接减弱了其活性。在PSSRRRVN肽中的uPA裂解使其恢复到与游离hIFNα细胞因子相似的水平。

为了评估和比较hCXCL10与ICC连接或被uPA裂解后的生物学活性，进行了由CXCL10与CXCR3受体结合诱导的β-抑制蛋白募集的生物学测试(PathHunter eXpressCXCR3 CHOK1β-抑制蛋白GPCR测定)。

12.1材料与方法

重组人尿激酶(＝uPA)购自Abcam(ab92767)。重组人CXCL10购自Peprotech(300-12)。CXCL10活性使用PathHunter eXpress CXCR3 CHOK1β-抑制蛋白GPCR分析(DiscoverX93-0271E2)监测。其为一种生物发光的基于CHO细胞的测定法，旨在测量CXCL10与CXCR3受体结合诱导的β-抑制蛋白募集。

GPCR-PK的激活诱导β-抑制蛋白-EA募集，迫使两个β-半乳糖苷酶片段(EA和PK)互补。所得的功能性酶水解底物以产生化学发光信号，该发光信号可在加入底物时检测并用发光计定量。

将待评估的构建体在PBS中用uPA 37℃孵育1小时。通过SDS-PAGE分析控制裂解效率，样品立即储存在-20℃直至处理。

用裂解的或未裂解的ICC或对照处理CHO-K1 CXCR3细胞1小时30分钟。然后加入检测剂，1小时后用酶标仪(Mithras,Berthold)记录化学发光强度。使用Prism 7.01软件(GraphPad)进行数据分析。

12.2结果

ICC和对照的hCXCL10活性的结果如图30A、图30B和图30C所示。数据以对应于hCXCL10活性水平的相对发光(RLU)随CXCR3β-抑制蛋白募集的变化表示。

重组hCXCL10和重组hCXCL10+uPA均强烈激活发光信号，从而验证了实验程式。在没有uPA的情况下，9G4-SGRS-CXCL10 ICC没有显著的活性。如通过CXCR3β-抑制蛋白募集所观察到的，用uPA预处理相同分子可导致活性恢复(图30A)。

总之，使用SGRS连接子将hCXCL10与9G4抗体在C-末端进行连接减弱了其活性。在SGRS肽中的uPA裂解使CXCL10活性几乎恢复到与游离hCXCL10细胞因子相似的水平。

重组hCXCL10和重组hCXCL10+uPA均强烈激活发光信号，从而验证了实验程式。在没有uPA的情况下，NHS76-SGRS-CXCL10 ICC没有显著的活性。如通过CXCR3β-抑制蛋白募集所观察到的，用uPA预处理相同分子可导致活性部分恢复(图30B)。未观察到全部活性，可能是由于亲本NHS76-SGRS-CXCL10的部分蛋白水解所致。

总之，使用SGRS将hCXCL10与NHS76抗体在C-末端进行连接减弱了其活性。SGRS肽内uPA的裂解可恢复CXCL10活性。

重组hCXCL10和重组hCXCL10+uPA均强烈激活发光信号，从而验证了实验程式。在没有uPA的情况下，H16/L16-SGRS-CXCL10 ICC没有显著的活性。如通过CXCR3β-抑制蛋白募集所观察到的，用uPA预处理相同分子可导致活性部分恢复(图30C)。未观察到全部活性，可能是由于亲本H16/L16-SGRS-CXCL10的部分蛋白水解所致。

总之，使用SGRS将hCXCL10与H16/L16抗体在C-末端进行连接减弱了其活性。SGRS肽内uPA的裂解可恢复CXCL10活性。

序列表

<110> 皮埃尔法布雷医药公司

<120> 用于治疗癌症的新型免疫细胞因子

<130> B376706PCTD38826

<150> EP 19305336.0

<151> 2019-03-19

<150> US 62/738391

<151> 2018-09-28

<160> 186

<170> PatentIn version 3.5

<210> 1

<211> 354

<212> DNA

<213> 人工序列

<220>

<223> 阿特珠单抗重链可变区

<400> 1

gaggtccagc ttgtcgagtc aggtggaggt cttgtacagc caggaggaag tttgcgtttg 60

agctgcgctg cttctgggtt cacttttagc gactcctgga tccactgggt cagacaggca 120

cctggaaagg ggctggaatg ggttgcatgg atttccccct acggcggctc tacttattac 180

gctgacagtg ttaaaggcag gtttaccata agcgccgaca cctccaaaaa caccgcctac 240

ctgcagatga atagtctgcg agccgaagat actgccgtgt actattgcgc ccggcgccat 300

tggcccggcg gctttgatta ttgggggcaa gggacactgg tgacagtgtc tgct 354

<210> 2

<211> 118

<212> PRT

<213> 人工序列

<220>

<223> 阿特珠单抗重链可变区

<400> 2

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala

115

<210> 3

<211> 320

<212> DNA

<213> 人工序列

<220>

<223> 阿特珠单抗轻链可变区

<400> 3

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa 320

<210> 4

<211> 107

<212> PRT

<213> 人工序列

<220>

<223> 阿特珠单抗轻链可变区

<400> 4

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys

100 105

<210> 5

<211> 357

<212> DNA

<213> 人工序列

<220>

<223> 盖尼塔单抗重链可变区

<400> 5

caggtgcagc tgcaggagtc gggcccagga ctggtgaagc cttcggggac cctgtccctc 60

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 120

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 300

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagc 357

<210> 6

<211> 119

<212> PRT

<213> 人工序列

<220>

<223> 盖尼塔单抗重链可变区

<400> 6

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Gly

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser Ile Ser Ser Ser

20 25 30

Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile Trp Gly Gln Gly

100 105 110

Thr Met Val Thr Val Ser Ser

115

<210> 7

<211> 336

<212> DNA

<213> 人工序列

<220>

<223> 盖尼塔单抗轻链可变区

<400> 7

gatgttgtga tgactcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60

atctcctgca ggtctagtca gagcctcctg catagtaatg gatacaacta tttggattgg 120

tacctgcaga agccagggca gtctccacag ctcctgatct atttgggttc taatcgggcc 180

tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240

agcagggtgg aggctgagga tgttggggtt tattactgca tgcaaggtac acactggcct 300

ctgacgttcg gccaagggac caaggtggag atcaaa 336

<210> 8

<211> 112

<212> PRT

<213> 人工序列

<220>

<223> 盖尼塔单抗轻链可变区

<400> 8

Asp Val Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly

1 5 10 15

Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu His Ser

20 25 30

Asn Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Pro Gly Gln Ser

35 40 45

Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro

50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile

65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gln Gly

85 90 95

Thr His Trp Pro Leu Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys

100 105 110

<210> 9

<211> 351

<212> DNA

<213> 人工序列

<220>

<223> NHS76重链可变区

<400> 9

caggtgcagc tgcaggagtc cggcccagga ctggtgaagc cttcggagac cctgtccctc 60

acctgcgctg tctctggtta ctccatcagc agtggttact actggggctg gattcggcag 120

cccccaggga aggggctgga gtggattggg agtatctatc atagtgggag cacctactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca cgtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcagac acggccgtgt attactgtgc aagagggaag 300

tggtcgaagt ttgactattg gggccaaggc accctggtca ccgtctcttc a 351

<210> 10

<211> 117

<212> PRT

<213> 人工序列

<220>

<223> NHS76重链可变区

<400> 10

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly

20 25 30

Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu

100 105 110

Val Thr Val Ser Ser

115

<210> 11

<211> 324

<212> DNA

<213> 人工序列

<220>

<223> NHS76轻链可变区

<400> 11

tcctctgagc tgactcagga ccctgctgtg tctgtggcct tgggacagac agtcaggatc 60

acatgccaag gagacagcct cagaagctat tatgcaagct ggtaccagca gaagccagga 120

caggcccctg tacttgtcat ctatggtaaa aacaaccggc cctcagggat tccagaccga 180

ttctctggct ccagctcagg aaacacagct tccttgacca tcactggggc tcaggcggaa 240

gatgaggctg actattactg taactcccgg gacagcagtg gtaaccatgt ggtattcggc 300

ggagggacca agctgaccgt ccta 324

<210> 12

<211> 108

<212> PRT

<213> 人工序列

<220>

<223> NHS76轻链可变区

<400> 12

Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln

1 5 10 15

Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala

20 25 30

Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr

35 40 45

Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser

50 55 60

Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu

65 70 75 80

Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His

85 90 95

Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu

100 105

<210> 13

<211> 339

<212> DNA

<213> 人工序列

<220>

<223> 9G4重链可变区

<400> 13

gaagtgatgt tggtggagtc tgggggaggc ttagtgaagc ctggagggtc cctgaaactc 60

tcctgtacag cctctggatt cactttcagt acctatgaca tgtcttgggt tcgccagact 120

ccggagaaga ggctggagtg ggtctcaacc attagtagta gtcgtactta cacctactat 180

tcagccagtg tgaaggggcg attcaccatc tccagagaca atgccaggaa catcctgtac 240

ctgcaaatga gcagtctgag gtctgaggac acggccatgt attactgtgc aagacaattt 300

tctcactggg gccaagggac tctggtcact gtctctgca 339

<210> 14

<211> 113

<212> PRT

<213> 人工序列

<220>

<223> 9G4重链可变区

<400> 14

Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly

1 5 10 15

Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Thr Tyr

20 25 30

Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val

35 40 45

Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser Ala Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr

65 70 75 80

Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val Thr Val Ser

100 105 110

Ala

<210> 15

<211> 336

<212> DNA

<213> 人工序列

<220>

<223> 9G4轻链可变区

<400> 15

gatgttgtga tgacccagac tccactcact ttgtcggtta ccattggaca accagcctcc 60

atctcttgca agtcaagtca gagcctctta gatagtgatg gaaagacata tttgaattgg 120

ttgttacaga ggccaggcca gtctccaacg cgcctaatct atctggtgtc taaactggac 180

tctggagtcc ctgacaggtt cactggcagt ggatcaggga cagatttcac actgaagatc 240

agcagagtgg aggctgagga tttgggagtt tattattgct ggcaaggtac acattttcct 300

cacacgttcg gaggggggac caagctggaa ataaaa 336

<210> 16

<211> 112

<212> PRT

<213> 人工序列

<220>

<223> 9G4轻链可变区

<400> 16

Asp Val Val Met Thr Gln Thr Pro Leu Thr Leu Ser Val Thr Ile Gly

1 5 10 15

Gln Pro Ala Ser Ile Ser Cys Lys Ser Ser Gln Ser Leu Leu Asp Ser

20 25 30

Asp Gly Lys Thr Tyr Leu Asn Trp Leu Leu Gln Arg Pro Gly Gln Ser

35 40 45

Pro Thr Arg Leu Ile Tyr Leu Val Ser Lys Leu Asp Ser Gly Val Pro

50 55 60

Asp Arg Phe Thr Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile

65 70 75 80

Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Trp Gln Gly

85 90 95

Thr His Phe Pro His Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys

100 105 110

<210> 17

<211> 987

<212> DNA

<213> 人

<400> 17

gcgtccacaa agggcccaag cgtgttcccg ctagccccca gcagcaagag caccagcggc 60

ggcacagccg ccctgggctg cctggtgaag gactacttcc ccgagcccgt gaccgtgtcc 120

tggaacagcg gagccctgac ctccggcgtg cacaccttcc ccgccgtgct gcagagcagc 180

ggcctgtaca gcctgagcag cgtggtgacc gtgcccagca gcagcctggg cacccagacc 240

tacatctgta acgtgaacca caagcccagc aacaccaagg tggacaagag agtggagccc 300

aagagctgtg acaagaccca cacctgtccc ccctgcccag cccccgagct gctgggcgga 360

cccagcgtgt tcctgttccc ccccaagccc aaggacaccc tgatgatcag cagaaccccc 420

gaggtgacct gtgtggtggt ggacgtgtcc cacgaggacc cagaggtgaa gttcaactgg 480

tacgtggacg gcgtggaggt gcacaacgcc aagaccaagc ccagagagga gcagtacaac 540

agcacctaca gggtggtgtc cgtgctgacc gtgctgcacc aggactggct gaacggcaag 600

gagtacaagt gtaaggtgtc caacaaggcc ctgccagccc caatcgaaaa gaccatcagc 660

aaggccaagg gccagccaag agagccccag gtgtacaccc tgccacccag cagggaggag 720

atgaccaaga accaggtgtc cctgacctgt ctggtgaagg gcttctaccc aagcgacatc 780

gccgtggagt gggagagcaa cggccagccc gagaacaact acaagaccac ccccccagtg 840

ctggacagcg acggcagctt cttcctgtac agcaagctga ccgtggacaa gagcagatgg 900

cagcagggca acgtgttcag ctgctccgtg atgcacgagg ccctgcacaa ccactacacc 960

cagaagagcc tgagcctgtc cccaggc 987

<210> 18

<211> 329

<212> PRT

<213> 人

<400> 18

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys

1 5 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr

20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser

35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser

50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr

65 70 75 80

Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys

85 90 95

Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys

100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

115 120 125

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp

145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

165 170 175

Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

180 185 190

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

195 200 205

Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

210 215 220

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu

225 230 235 240

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

245 250 255

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn

290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

305 310 315 320

Gln Lys Ser Leu Ser Leu Ser Pro Gly

325

<210> 19

<211> 978

<212> DNA

<213> 人

<400> 19

gcgtccacaa agggccccag cgtgttccct ctggccccct gcagcagaag caccagcgag 60

tctacagccg ccctgggatg cctggtcaag gactacttcc ccgagcccgt gaccgtgtcc 120

tggaactctg gcgccctgac cagcggcgtg cacacctttc cagccgtgct gcagagcagc 180

ggcctgtaca gcctgagcag cgtcgtgacc gtgcctagca gcagcctggg caccaagacc 240

tacacctgta acgtggacca caagcccagc aacaccaagg tggacaagcg ggtggaatct 300

aagtacggcc ctccctgccc cccctgccca gcccctgaat ttctgggcgg accctccgtg 360

ttcctgttcc ccccaaagcc caaggacacc ctgatgatca gccggacccc cgaagtgacc 420

tgcgtggtgg tggacgtgtc ccaggaagat cccgaggtcc agttcaactg gtacgtggac 480

ggcgtggaag tgcacaacgc caagaccaag cccagagagg aacagttcaa cagcacctac 540

cgggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa agagtacaag 600

tgcaaggtct ccaacaaggg cctgcccagc tccatcgaga aaaccatcag caaggccaag 660

ggccagcccc gcgagcccca ggtgtacaca ctgcccccca gccaggaaga gatgaccaag 720

aaccaggtgt ccctgacctg cctcgtgaag ggcttctacc ccagcgatat cgccgtggaa 780

tgggagagca acggccagcc cgagaacaac tacaagacca ccccccctgt gctggacagc 840

gacggcagct tcttcctgta ctcccggctg accgtcgaca agtctagatg gcaggaaggc 900

aacgtcttca gctgcagcgt gatgcacgag gccctgcaca accactacac ccagaagtcc 960

ctgagcctga gcctggga 978

<210> 20

<211> 326

<212> PRT

<213> 人

<400> 20

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser Arg

1 5 10 15

Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr

20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser

35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser

50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys Thr

65 70 75 80

Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys

85 90 95

Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro

100 105 110

Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys

115 120 125

Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val

130 135 140

Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp

145 150 155 160

Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe

165 170 175

Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp

180 185 190

Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu

195 200 205

Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg

210 215 220

Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr Lys

225 230 235 240

Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp

245 250 255

Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys

260 265 270

Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser

275 280 285

Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser

290 295 300

Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser

305 310 315 320

Leu Ser Leu Ser Leu Gly

325

<210> 21

<211> 321

<212> DNA

<213> 人

<400> 21

cgtacggtgg ccgctcccag cgtgttcatc ttcccaccca gcgacgagca gctgaagtct 60

ggcaccgcca gcgtcgtgtg cctgctgaac aacttctacc cccgcgaggc caaggtgcag 120

tggaaggtgg acaacgccct gcagtccggc aacagccagg aaagcgtcac cgagcaggac 180

agcaaggact ccacctacag cctgagcagc accctgaccc tgagcaaggc cgactacgag 240

aagcacaagg tgtacgcctg cgaagtgacc caccagggcc tgtccagccc cgtgaccaag 300

agcttcaacc ggggcgagtg c 321

<210> 22

<211> 107

<212> PRT

<213> 人

<400> 22

Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu

1 5 10 15

Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe

20 25 30

Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln

35 40 45

Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser

50 55 60

Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu

65 70 75 80

Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser

85 90 95

Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys

100 105

<210> 23

<211> 318

<212> DNA

<213> 人

<400> 23

ggtcaaccca aggccgctcc cagcgtgacc ctgttccccc cctcgagcga ggagctgcag 60

gccaacaagg ccaccctggt gtgtctgatc agcgacttct acccaggcgc cgtgaccgtg 120

gcctggaagg ccgacagcag ccccgtgaag gccggcgtgg agaccaccac ccccagcaag 180

cagagcaaca acaagtacgc cgccagcagc tacctgagcc tgaccccaga gcagtggaag 240

agccacagga gctacagctg ccaggtcacc cacgagggca gcaccgtgga aaagaccgtg 300

gccccaaccg agtgctcc 318

<210> 24

<211> 106

<212> PRT

<213> 人

<400> 24

Gly Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser

1 5 10 15

Glu Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp

20 25 30

Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro

35 40 45

Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn

50 55 60

Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys

65 70 75 80

Ser His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val

85 90 95

Glu Lys Thr Val Ala Pro Thr Glu Cys Ser

100 105

<210> 25

<211> 342

<212> DNA

<213> 人

<400> 25

aactgggtga atgtaataag tgatttgaaa aaaattgaag atcttattca atctatgcat 60

attgatgcta ctttatatac ggaaagtgat gttcacccca gttgcaaagt aacagcaatg 120

aagtgctttc tcttggagtt acaagttatt tcacttgagt ccggagatgc aagtattcat 180

gatacagtag aaaatctgat catcctagca aacaacagtt tgtcttctaa tgggaatgta 240

acagaatctg gatgcaaaga atgtgaggaa ctggaggaaa aaaatattaa agaatttttg 300

cagagttttg tacatattgt ccaaatgttc atcaacactt ct 342

<210> 26

<211> 114

<212> PRT

<213> 人

<400> 26

Asn Trp Val Asn Val Ile Ser Asp Leu Lys Lys Ile Glu Asp Leu Ile

1 5 10 15

Gln Ser Met His Ile Asp Ala Thr Leu Tyr Thr Glu Ser Asp Val His

20 25 30

Pro Ser Cys Lys Val Thr Ala Met Lys Cys Phe Leu Leu Glu Leu Gln

35 40 45

Val Ile Ser Leu Glu Ser Gly Asp Ala Ser Ile His Asp Thr Val Glu

50 55 60

Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu Ser Ser Asn Gly Asn Val

65 70 75 80

Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu Leu Glu Glu Lys Asn Ile

85 90 95

Lys Glu Phe Leu Gln Ser Phe Val His Ile Val Gln Met Phe Ile Asn

100 105 110

Thr Ser

<210> 27

<211> 399

<212> DNA

<213> 人

<400> 27

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgact 399

<210> 28

<211> 133

<212> PRT

<213> 人

<400> 28

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr

130

<210> 29

<211> 207

<212> DNA

<213> 人

<400> 29

gcaccaatgg gctcagaccc tcccaccgcc tgctgctttt cttacaccgc gaggaaactt 60

cctcgcaact ttgtggtaga ttactatgag accagcagcc tctgctccca gccagctgtg 120

gtattccaaa ccaaaagaag caagcaagtc tgtgctgatc ccagtgaatc ctgggtccag 180

gagtacgtgt atgacctgga actgaac 207

<210> 30

<211> 69

<212> PRT

<213> 人

<400> 30

Ala Pro Met Gly Ser Asp Pro Pro Thr Ala Cys Cys Phe Ser Tyr Thr

1 5 10 15

Ala Arg Lys Leu Pro Arg Asn Phe Val Val Asp Tyr Tyr Glu Thr Ser

20 25 30

Ser Leu Cys Ser Gln Pro Ala Val Val Phe Gln Thr Lys Arg Ser Lys

35 40 45

Gln Val Cys Ala Asp Pro Ser Glu Ser Trp Val Gln Glu Tyr Val Tyr

50 55 60

Asp Leu Glu Leu Asn

65

<210> 31

<211> 495

<212> DNA

<213> 人

<400> 31

tgtgatctgc ctcaaaccca cagcctgggt agcaggagga ccttgatgct cctggcacag 60

atgaggaaaa tctctctttt ctcctgcttg aaggacagac atgactttgg atttccccag 120

gaggagtttg gcaaccagtt ccaaaaggct gaaaccatcc ctgtcctcca tgagatgatc 180

cagcagatct tcaatctctt cagcacaaag gactcatctg ctgcttggga tgagaccctc 240

ctagacaaat tctacactga actctaccag cagctgaatg acctggaagc ctgtgtgata 300

cagggggtgg gggtgacaga gactcccctg atgaaggagg actccattct ggctgtgagg 360

aaatacttcc aaagaatcac tctctatctg aaagagaaga aatacagccc ttgtgcctgg 420

gaggttgtca gagcagaaat catgagatct ttttctttgt caacaaactt gcaagaaagt 480

ttaagaagta aggaa 495

<210> 32

<211> 165

<212> PRT

<213> 人

<400> 32

Cys Asp Leu Pro Gln Thr His Ser Leu Gly Ser Arg Arg Thr Leu Met

1 5 10 15

Leu Leu Ala Gln Met Arg Lys Ile Ser Leu Phe Ser Cys Leu Lys Asp

20 25 30

Arg His Asp Phe Gly Phe Pro Gln Glu Glu Phe Gly Asn Gln Phe Gln

35 40 45

Lys Ala Glu Thr Ile Pro Val Leu His Glu Met Ile Gln Gln Ile Phe

50 55 60

Asn Leu Phe Ser Thr Lys Asp Ser Ser Ala Ala Trp Asp Glu Thr Leu

65 70 75 80

Leu Asp Lys Phe Tyr Thr Glu Leu Tyr Gln Gln Leu Asn Asp Leu Glu

85 90 95

Ala Cys Val Ile Gln Gly Val Gly Val Thr Glu Thr Pro Leu Met Lys

100 105 110

Glu Asp Ser Ile Leu Ala Val Arg Lys Tyr Phe Gln Arg Ile Thr Leu

115 120 125

Tyr Leu Lys Glu Lys Lys Tyr Ser Pro Cys Ala Trp Glu Val Val Arg

130 135 140

Ala Glu Ile Met Arg Ser Phe Ser Leu Ser Thr Asn Leu Gln Glu Ser

145 150 155 160

Leu Arg Ser Lys Glu

165

<210> 33

<211> 513

<212> DNA

<213> 深海虾（Oplophorus gracilirostris）

<400> 33

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcg 513

<210> 34

<211> 173

<212> PRT

<213> 深海虾（Oplophorus gracilirostris）

<400> 34

Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln

1 5 10 15

Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser

20 25 30

Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile

35 40 45

Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile

50 55 60

Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile

65 70 75 80

Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu

85 90 95

His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp

100 105 110

Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys

115 120 125

Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu

130 135 140

Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn

145 150 155 160

Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

165 170

<210> 35

<211> 18

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 35

ccgctgggcc tggcgggc 18

<210> 36

<211> 6

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 36

Pro Leu Gly Leu Ala Gly

1 5

<210> 37

<211> 24

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 37

ggcccgctgg gcattgcggg ccag 24

<210> 38

<211> 8

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 38

Gly Pro Leu Gly Ile Ala Gly Gln

1 5

<210> 39

<211> 24

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 39

ggcccgctgg gcctgtgggc gcag 24

<210> 40

<211> 8

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 40

Gly Pro Leu Gly Leu Trp Ala Gln

1 5

<210> 41

<211> 24

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 41

ggcccgctgg gcatgctgag ccag 24

<210> 42

<211> 8

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 42

Gly Pro Leu Gly Met Leu Ser Gln

1 5

<210> 43

<211> 18

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 43

ccggtgggcc tgattggc 18

<210> 44

<211> 6

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 44

Pro Val Gly Leu Ile Gly

1 5

<210> 45

<211> 18

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 45

ggcattgtgg gaccgctg 18

<210> 46

<211> 6

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 46

Gly Ile Val Gly Pro Leu

1 5

<210> 47

<211> 6

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 47

ctcgag 6

<210> 48

<211> 2

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 48

Leu Glu

1

<210> 49

<211> 1704

<212> DNA

<213> 人工序列

<220>

<223> 盖尼塔单抗-PVGLIG-IL15重链

<400> 49

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 60

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 120

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 300

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 360

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 420

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 480

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 540

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 600

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 660

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 720

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 780

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 840

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 900

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 960

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1020

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1080

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1140

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1200

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1260

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1320

aagagcctga gcctgtcccc aggcccggtg ggcctgattg gcaactgggt gaatgtaata 1380

agtgatttga aaaaaattga agatcttatt caatctatgc atattgatgc tactttatat 1440

acggaaagtg atgttcaccc cagttgcaaa gtaacagcaa tgaagtgctt tctcttggag 1500

ttacaagtta tttcacttga gtccggagat gcaagtattc atgatacagt agaaaatctg 1560

atcatcctag caaacaacag tttgtcttct aatgggaatg taacagaatc tggatgcaaa 1620

gaatgtgagg aactggagga aaaaaatatt aaagaatttt tgcagagttt tgtacatatt 1680

gtccaaatgt tcatcaacac ttct 1704

<210> 50

<211> 568

<212> PRT

<213> 人工序列

<220>

<223> 盖尼塔单抗-PVGLIG-IL15重链

<400> 50

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Gly

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser Ile Ser Ser Ser

20 25 30

Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile Trp Gly Gln Gly

100 105 110

Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe

115 120 125

Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu

130 135 140

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp

145 150 155 160

Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu

165 170 175

Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser

180 185 190

Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro

195 200 205

Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys

210 215 220

Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro

225 230 235 240

Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser

245 250 255

Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp

260 265 270

Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn

275 280 285

Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val

290 295 300

Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu

305 310 315 320

Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys

325 330 335

Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr

340 345 350

Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr

355 360 365

Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu

370 375 380

Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu

385 390 395 400

Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys

405 410 415

Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu

420 425 430

Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

435 440 445

Pro Val Gly Leu Ile Gly Asn Trp Val Asn Val Ile Ser Asp Leu Lys

450 455 460

Lys Ile Glu Asp Leu Ile Gln Ser Met His Ile Asp Ala Thr Leu Tyr

465 470 475 480

Thr Glu Ser Asp Val His Pro Ser Cys Lys Val Thr Ala Met Lys Cys

485 490 495

Phe Leu Leu Glu Leu Gln Val Ile Ser Leu Glu Ser Gly Asp Ala Ser

500 505 510

Ile His Asp Thr Val Glu Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu

515 520 525

Ser Ser Asn Gly Asn Val Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu

530 535 540

Leu Glu Glu Lys Asn Ile Lys Glu Phe Leu Gln Ser Phe Val His Ile

545 550 555 560

Val Gln Met Phe Ile Asn Thr Ser

565

<210> 51

<211> 1698

<212> DNA

<213> 人工序列

<220>

<223> NHS76-PVGLIG-IL15重链

<400> 51

caggtgcagc tgcaggagtc cggcccagga ctggtgaagc cttcggagac cctgtccctc 60

acctgcgctg tctctggtta ctccatcagc agtggttact actggggctg gattcggcag 120

cccccaggga aggggctgga gtggattggg agtatctatc atagtgggag cacctactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca cgtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcagac acggccgtgt attactgtgc aagagggaag 300

tggtcgaagt ttgactattg gggccaaggc accctggtca ccgtctcttc agcgtccaca 360

aagggcccaa gcgtgttccc gctagccccc agcagcaaga gcaccagcgg cggcacagcc 420

gccctgggct gcctggtgaa ggactacttc cccgagcccg tgaccgtgtc ctggaacagc 480

ggagccctga cctccggcgt gcacaccttc cccgccgtgc tgcagagcag cggcctgtac 540

agcctgagca gcgtggtgac cgtgcccagc agcagcctgg gcacccagac ctacatctgt 600

aacgtgaacc acaagcccag caacaccaag gtggacaaga gagtggagcc caagagctgt 660

gacaagaccc acacctgtcc cccctgccca gcccccgagc tgctgggcgg acccagcgtg 720

ttcctgttcc cccccaagcc caaggacacc ctgatgatca gcagaacccc cgaggtgacc 780

tgtgtggtgg tggacgtgtc ccacgaggac ccagaggtga agttcaactg gtacgtggac 840

ggcgtggagg tgcacaacgc caagaccaag cccagagagg agcagtacaa cagcacctac 900

agggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa ggagtacaag 960

tgtaaggtgt ccaacaaggc cctgccagcc ccaatcgaaa agaccatcag caaggccaag 1020

ggccagccaa gagagcccca ggtgtacacc ctgccaccca gcagggagga gatgaccaag 1080

aaccaggtgt ccctgacctg tctggtgaag ggcttctacc caagcgacat cgccgtggag 1140

tgggagagca acggccagcc cgagaacaac tacaagacca cccccccagt gctggacagc 1200

gacggcagct tcttcctgta cagcaagctg accgtggaca agagcagatg gcagcagggc 1260

aacgtgttca gctgctccgt gatgcacgag gccctgcaca accactacac ccagaagagc 1320

ctgagcctgt ccccaggccc ggtgggcctg attggcaact gggtgaatgt aataagtgat 1380

ttgaaaaaaa ttgaagatct tattcaatct atgcatattg atgctacttt atatacggaa 1440

agtgatgttc accccagttg caaagtaaca gcaatgaagt gctttctctt ggagttacaa 1500

gttatttcac ttgagtccgg agatgcaagt attcatgata cagtagaaaa tctgatcatc 1560

ctagcaaaca acagtttgtc ttctaatggg aatgtaacag aatctggatg caaagaatgt 1620

gaggaactgg aggaaaaaaa tattaaagaa tttttgcaga gttttgtaca tattgtccaa 1680

atgttcatca acacttct 1698

<210> 52

<211> 566

<212> PRT

<213> 人工序列

<220>

<223> NHS76-PVGLIG-IL15重链

<400> 52

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly

20 25 30

Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu

100 105 110

Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu

115 120 125

Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys

130 135 140

Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser

145 150 155 160

Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser

165 170 175

Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser

180 185 190

Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn

195 200 205

Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His

210 215 220

Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val

225 230 235 240

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr

245 250 255

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu

260 265 270

Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys

275 280 285

Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser

290 295 300

Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys

305 310 315 320

Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile

325 330 335

Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro

340 345 350

Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu

355 360 365

Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn

370 375 380

Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser

385 390 395 400

Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg

405 410 415

Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu

420 425 430

His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val

435 440 445

Gly Leu Ile Gly Asn Trp Val Asn Val Ile Ser Asp Leu Lys Lys Ile

450 455 460

Glu Asp Leu Ile Gln Ser Met His Ile Asp Ala Thr Leu Tyr Thr Glu

465 470 475 480

Ser Asp Val His Pro Ser Cys Lys Val Thr Ala Met Lys Cys Phe Leu

485 490 495

Leu Glu Leu Gln Val Ile Ser Leu Glu Ser Gly Asp Ala Ser Ile His

500 505 510

Asp Thr Val Glu Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu Ser Ser

515 520 525

Asn Gly Asn Val Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu Leu Glu

530 535 540

Glu Lys Asn Ile Lys Glu Phe Leu Gln Ser Phe Val His Ile Val Gln

545 550 555 560

Met Phe Ile Asn Thr Ser

565

<210> 53

<211> 657

<212> DNA

<213> 人工序列

<220>

<223> 盖尼塔单抗轻链

<400> 53

gatgttgtga tgactcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60

atctcctgca ggtctagtca gagcctcctg catagtaatg gatacaacta tttggattgg 120

tacctgcaga agccagggca gtctccacag ctcctgatct atttgggttc taatcgggcc 180

tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240

agcagggtgg aggctgagga tgttggggtt tattactgca tgcaaggtac acactggcct 300

ctgacgttcg gccaagggac caaggtggag atcaaacgta cggtggccgc tcccagcgtg 360

ttcatcttcc cacccagcga cgagcagctg aagtctggca ccgccagcgt cgtgtgcctg 420

ctgaacaact tctacccccg cgaggccaag gtgcagtgga aggtggacaa cgccctgcag 480

tccggcaaca gccaggaaag cgtcaccgag caggacagca aggactccac ctacagcctg 540

agcagcaccc tgaccctgag caaggccgac tacgagaagc acaaggtgta cgcctgcgaa 600

gtgacccacc agggcctgtc cagccccgtg accaagagct tcaaccgggg cgagtgc 657

<210> 54

<211> 219

<212> PRT

<213> 人工序列

<220>

<223> 盖尼塔单抗轻链

<400> 54

Asp Val Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly

1 5 10 15

Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu His Ser

20 25 30

Asn Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Pro Gly Gln Ser

35 40 45

Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro

50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile

65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gln Gly

85 90 95

Thr His Trp Pro Leu Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys

100 105 110

Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu

115 120 125

Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe

130 135 140

Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln

145 150 155 160

Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser

165 170 175

Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu

180 185 190

Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser

195 200 205

Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys

210 215

<210> 55

<211> 645

<212> DNA

<213> 人工序列

<220>

<223> NHS76轻链

<400> 55

tcctctgagc tgactcagga ccctgctgtg tctgtggcct tgggacagac agtcaggatc 60

acatgccaag gagacagcct cagaagctat tatgcaagct ggtaccagca gaagccagga 120

caggcccctg tacttgtcat ctatggtaaa aacaaccggc cctcagggat tccagaccga 180

ttctctggct ccagctcagg aaacacagct tccttgacca tcactggggc tcaggcggaa 240

gatgaggctg actattactg taactcccgg gacagcagtg gtaaccatgt ggtattcggc 300

ggagggacca agctgaccgt cctaggtcaa cccaaggccg ctcccagcgt gaccctgttc 360

cccccctcga gcgaggagct gcaggccaac aaggccaccc tggtgtgtct gatcagcgac 420

ttctacccag gcgccgtgac cgtggcctgg aaggccgaca gcagccccgt gaaggccggc 480

gtggagacca ccacccccag caagcagagc aacaacaagt acgccgccag cagctacctg 540

agcctgaccc cagagcagtg gaagagccac aggagctaca gctgccaggt cacccacgag 600

ggcagcaccg tggaaaagac cgtggcccca accgagtgct cctga 645

<210> 56

<211> 214

<212> PRT

<213> 人工序列

<220>

<223> NHS76轻链

<400> 56

Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln

1 5 10 15

Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala

20 25 30

Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr

35 40 45

Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser

50 55 60

Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu

65 70 75 80

Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His

85 90 95

Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln Pro Lys

100 105 110

Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gln

115 120 125

Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly

130 135 140

Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly

145 150 155 160

Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr Ala Ala

165 170 175

Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His Arg Ser

180 185 190

Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val

195 200 205

Ala Pro Thr Glu Cys Ser

210

<210> 57

<211> 1875

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc重链（C-末端融合）

<400> 57

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcatgctg agccagctcg agatggtctt cacactcgaa 1380

gatttcgttg gggactggcg acagacagcc ggctacaacc tggaccaagt ccttgaacag 1440

ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg taactccgat ccaaaggatt 1500

gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg tcatcatccc gtatgaaggt 1560

ctgagcggcg accaaatggg ccagatcgaa aaaattttta aggtggtgta ccctgtggat 1620

gatcatcact ttaaggtgat cctgcactat ggcacactgg taatcgacgg ggttacgccg 1680

aacatgatcg actatttcgg acggccgtat gaaggcatcg ccgtgttcga cggcaaaaag 1740

atcactgtaa cagggaccct gtggaacggc aacaaaatta tcgacgagcg cctgatcaac 1800

cccgacggct ccctgctgtt ccgagtaacc atcaacggag tgaccggctg gcggctgtgc 1860

gaacgcattc tggcg 1875

<210> 58

<211> 625

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc重链（C-末端融合）

<400> 58

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Met Leu Ser Gln Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly

450 455 460

Asp Trp Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln

465 470 475 480

Gly Gly Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro

485 490 495

Ile Gln Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile

500 505 510

His Val Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln

515 520 525

Ile Glu Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe

530 535 540

Lys Val Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro

545 550 555 560

Asn Met Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe

565 570 575

Asp Gly Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys

580 585 590

Ile Ile Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg

595 600 605

Val Thr Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu

610 615 620

Ala

625

<210> 59

<211> 1185

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc轻链（C-末端融合）

<400> 59

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcatgctg 660

agccagctcg agatggtctt cacactcgaa gatttcgttg gggactggcg acagacagcc 720

ggctacaacc tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc 780

ggggtgtccg taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc 840

gacatccatg tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa 900

aaaattttta aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat 960

ggcacactgg taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat 1020

gaaggcatcg ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc 1080

aacaaaatta tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc 1140

atcaacggag tgaccggctg gcggctgtgc gaacgcattc tggcg 1185

<210> 60

<211> 395

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc轻链（C-末端融合）

<400> 60

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Met Leu Ser Gln Leu Glu

210 215 220

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

225 230 235 240

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

245 250 255

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

260 265 270

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

275 280 285

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

290 295 300

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

305 310 315 320

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

325 330 335

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

340 345 350

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

355 360 365

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

370 375 380

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390 395

<210> 61

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc重链（N-末端融合）

<400> 61

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcatgct gagccaggaa 540

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 600

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 660

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 720

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 780

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 840

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 900

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 960

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 1020

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 1080

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1140

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1200

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1260

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1320

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1380

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1440

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1500

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1560

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1620

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1680

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1740

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1800

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1860

agcctggga 1869

<210> 62

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc重链（N-末端融合）

<400> 62

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Met

165 170 175

Leu Ser Gln Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln

180 185 190

Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe

195 200 205

Ser Asp Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu

210 215 220

Glu Trp Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala

225 230 235 240

Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn

245 250 255

Thr Ala Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val

260 265 270

Tyr Tyr Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly

275 280 285

Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser

290 295 300

Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala

305 310 315 320

Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val

325 330 335

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala

340 345 350

Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val

355 360 365

Pro Ser Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His

370 375 380

Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly

385 390 395 400

Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser

405 410 415

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

420 425 430

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro

435 440 445

Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

450 455 460

Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val

465 470 475 480

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

485 490 495

Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr

500 505 510

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

515 520 525

Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys

530 535 540

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

545 550 555 560

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

565 570 575

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser

580 585 590

Arg Trp Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

595 600 605

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 63

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc轻链（N-末端融合）

<400> 63

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcatgct gagccaggac 540

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 600

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 660

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 720

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 780

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 840

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 900

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 960

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 1020

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 1080

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1140

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1179

<210> 64

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-NanoLuc轻链（N-末端融合）

<400> 64

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Met

165 170 175

Leu Ser Gln Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala

180 185 190

Ser Val Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val

195 200 205

Ser Thr Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys

210 215 220

Leu Leu Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg

225 230 235 240

Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser

245 250 255

Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr

260 265 270

His Pro Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr

275 280 285

Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu

290 295 300

Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro

305 310 315 320

Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly

325 330 335

Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr

340 345 350

Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His

355 360 365

Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val

370 375 380

Thr Lys Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 65

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2重链（C-末端融合）

<400> 65

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcatgctg agccaggcac ctacctcgag ttctacaaag 1380

aaaacacagc tacaactgga gcatttactg ctggatttac agatgatttt gaatggaatt 1440

aataattaca agaatcccaa actcaccagg atgctcacat ttaagtttta catgcccaag 1500

aaggccacag aactgaaaca tcttcagtgt ctagaagaag aactcaaacc tctggaggaa 1560

gtgctaaatt tagctcaaag caaaaacttt cacttaagac ccagggactt aatcagcaat 1620

atcaacgtaa tagttctgga actaaaggga tctgaaacaa cattcatgtg tgaatatgct 1680

gatgagacag caaccattgt agaatttctg aacagatgga ttaccttttg tcaaagcatc 1740

atctcaacac tgact 1755

<210> 66

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2重链（C-末端融合）

<400> 66

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Met Leu Ser Gln Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu

450 455 460

Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile

465 470 475 480

Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe

485 490 495

Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu

500 505 510

Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys

515 520 525

Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile

530 535 540

Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala

545 550 555 560

Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe

565 570 575

Cys Gln Ser Ile Ile Ser Thr Leu Thr

580 585

<210> 67

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2轻链（C-末端融合）

<400> 67

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcatgctg 660

agccaggcac ctacctcgag ttctacaaag aaaacacagc tacaactgga gcatttactg 720

ctggatttac agatgatttt gaatggaatt aataattaca agaatcccaa actcaccagg 780

atgctcacat ttaagtttta catgcccaag aaggccacag aactgaaaca tcttcagtgt 840

ctagaagaag aactcaaacc tctggaggaa gtgctaaatt tagctcaaag caaaaacttt 900

cacttaagac ccagggactt aatcagcaat atcaacgtaa tagttctgga actaaaggga 960

tctgaaacaa cattcatgtg tgaatatgct gatgagacag caaccattgt agaatttctg 1020

aacagatgga ttaccttttg tcaaagcatc atctcaacac tgact 1065

<210> 68

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2轻链（C-末端融合）

<400> 68

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Met Leu Ser Gln Ala Pro

210 215 220

Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu

225 230 235 240

Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro

245 250 255

Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala

260 265 270

Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu

275 280 285

Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro

290 295 300

Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly

305 310 315 320

Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile

325 330 335

Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser

340 345 350

Thr Leu Thr

355

<210> 69

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2重链（N-末端融合）

<400> 69

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg catgctgagc 420

caggaagtcc aactggtcga aagcggtgga ggtctggtac agccaggagg aagtttgaga 480

ctgtcatgcg ctgcttccgg gtttactttc agcgactcct ggattcactg ggttcgtcag 540

gcacctggaa aggggcttga gtgggttgca tggatttccc catacggcgg ctctacatac 600

tacgctgaca gcgtcaaggg caggtttaca ataagcgccg atacctctaa aaacaccgcc 660

tacctgcaga tgaatagttt gcgagccgag gataccgccg tgtattattg tgcccggcgc 720

cattggcccg gcggcttcga ttattggggg caggggacac ttgtgacagt gtctgcagcc 780

tctaccaagg gccccagcgt gttccctctg gccccctgca gcagaagcac cagcgagtct 840

acagccgccc tgggatgcct ggtcaaggac tacttccccg agcccgtgac cgtgtcctgg 900

aactctggcg ccctgaccag cggcgtgcac acctttccag ccgtgctgca gagcagcggc 960

ctgtacagcc tgagcagcgt cgtgaccgtg cctagcagca gcctgggcac caagacctac 1020

acctgtaacg tggaccacaa gcccagcaac accaaggtgg acaagcgggt ggaatctaag 1080

tacggccctc cctgcccccc ctgcccagcc cctgaatttc tgggcggacc ctccgtgttc 1140

ctgttccccc caaagcccaa ggacaccctg atgatcagcc ggacccccga agtgacctgc 1200

gtggtggtgg acgtgtccca ggaagatccc gaggtccagt tcaactggta cgtggacggc 1260

gtggaagtgc acaacgccaa gaccaagccc agagaggaac agttcaacag cacctaccgg 1320

gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga gtacaagtgc 1380

aaggtctcca acaagggcct gcccagctcc atcgagaaaa ccatcagcaa ggccaagggc 1440

cagccccgcg agccccaggt gtacacactg ccccccagcc aggaagagat gaccaagaac 1500

caggtgtccc tgacctgcct cgtgaagggc ttctacccca gcgatatcgc cgtggaatgg 1560

gagagcaacg gccagcccga gaacaactac aagaccaccc cccctgtgct ggacagcgac 1620

ggcagcttct tcctgtactc ccggctgacc gtcgacaaga gccggtggca ggaaggcaac 1680

gtcttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1740

agcctgagcc tggga 1755

<210> 70

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2重链（N-末端融合）

<400> 70

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Met Leu Ser Gln Glu Val Gln

130 135 140

Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg

145 150 155 160

Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His

165 170 175

Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile

180 185 190

Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg

195 200 205

Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met

210 215 220

Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg

225 230 235 240

His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr

245 250 255

Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro

260 265 270

Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val

275 280 285

Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala

290 295 300

Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly

305 310 315 320

Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly

325 330 335

Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys

340 345 350

Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

355 360 365

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

370 375 380

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

385 390 395 400

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

405 410 415

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

420 425 430

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

435 440 445

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

450 455 460

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

465 470 475 480

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu

485 490 495

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

500 505 510

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

515 520 525

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

530 535 540

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

545 550 555 560

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

565 570 575

Gln Lys Ser Leu Ser Leu Ser Leu Gly

580 585

<210> 71

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2轻链（N-末端融合）

<400> 71

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg catgctgagc 420

caggacatcc agatgaccca atccccatcc tccttgagcg catctgtagg agacagagtc 480

accatcacct gcagggcttc acaggatgtt agcacagctg tggcttggta ccagcagaaa 540

cctggaaagg ccccaaaact gctgatatac agcgcctcat tcctgtacag cggagtgccc 600

tctcgcttta gtggctctgg gagtggcact gatttcacac tgactatttc tagtctgcag 660

cccgaagatt tcgccactta ctattgccag cagtatctct atcaccccgc cacctttggg 720

cagggcacaa aggtcgagat taagcgtacg gtggccgctc ccagcgtgtt catcttccca 780

cccagcgacg agcagctgaa gtctggcacc gccagcgtcg tgtgcctgct gaacaacttc 840

tacccccgcg aggccaaggt gcagtggaag gtggacaacg ccctgcagtc cggcaacagc 900

caggaaagcg tcaccgagca ggacagcaag gactccacct acagcctgag cagcaccctg 960

accctgagca aggccgacta cgagaagcac aaggtgtacg cctgcgaagt gacccaccag 1020

ggcctgtcca gccccgtgac caagagcttc aaccggggcg agtgc 1065

<210> 72

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGMLSQ-IL2轻链（N-末端融合）

<400> 72

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Met Leu Ser Gln Asp Ile Gln

130 135 140

Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val

145 150 155 160

Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp

165 170 175

Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala

180 185 190

Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser

195 200 205

Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe

210 215 220

Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly

225 230 235 240

Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val

245 250 255

Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser

260 265 270

Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln

275 280 285

Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val

290 295 300

Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu

305 310 315 320

Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu

325 330 335

Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg

340 345 350

Gly Glu Cys

355

<210> 73

<211> 1875

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc重链（C-末端融合）

<400> 73

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcattgcg ggccagctcg agatggtctt cacactcgaa 1380

gatttcgttg gggactggcg acagacagcc ggctacaacc tggaccaagt ccttgaacag 1440

ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg taactccgat ccaaaggatt 1500

gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg tcatcatccc gtatgaaggt 1560

ctgagcggcg accaaatggg ccagatcgaa aaaattttta aggtggtgta ccctgtggat 1620

gatcatcact ttaaggtgat cctgcactat ggcacactgg taatcgacgg ggttacgccg 1680

aacatgatcg actatttcgg acggccgtat gaaggcatcg ccgtgttcga cggcaaaaag 1740

atcactgtaa cagggaccct gtggaacggc aacaaaatta tcgacgagcg cctgatcaac 1800

cccgacggct ccctgctgtt ccgagtaacc atcaacggag tgaccggctg gcggctgtgc 1860

gaacgcattc tggcg 1875

<210> 74

<211> 625

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc重链（C-末端融合）

<400> 74

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Ile Ala Gly Gln Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly

450 455 460

Asp Trp Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln

465 470 475 480

Gly Gly Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro

485 490 495

Ile Gln Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile

500 505 510

His Val Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln

515 520 525

Ile Glu Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe

530 535 540

Lys Val Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro

545 550 555 560

Asn Met Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe

565 570 575

Asp Gly Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys

580 585 590

Ile Ile Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg

595 600 605

Val Thr Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu

610 615 620

Ala

625

<210> 75

<211> 1185

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc轻链（C-末端融合）

<400> 75

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcattgcg 660

ggccagctcg agatggtctt cacactcgaa gatttcgttg gggactggcg acagacagcc 720

ggctacaacc tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc 780

ggggtgtccg taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc 840

gacatccatg tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa 900

aaaattttta aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat 960

ggcacactgg taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat 1020

gaaggcatcg ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc 1080

aacaaaatta tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc 1140

atcaacggag tgaccggctg gcggctgtgc gaacgcattc tggcg 1185

<210> 76

<211> 395

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc轻链（C-末端融合）

<400> 76

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Ile Ala Gly Gln Leu Glu

210 215 220

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

225 230 235 240

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

245 250 255

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

260 265 270

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

275 280 285

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

290 295 300

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

305 310 315 320

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

325 330 335

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

340 345 350

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

355 360 365

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

370 375 380

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390 395

<210> 77

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc重链（N-末端融合）

<400> 77

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcattgc gggccaggaa 540

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 600

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 660

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 720

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 780

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 840

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 900

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 960

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 1020

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 1080

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1140

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1200

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1260

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1320

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1380

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1440

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1500

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1560

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1620

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1680

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1740

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1800

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1860

agcctggga 1869

<210> 78

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc重链（N-末端融合）

<400> 78

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Ile

165 170 175

Ala Gly Gln Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln

180 185 190

Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe

195 200 205

Ser Asp Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu

210 215 220

Glu Trp Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala

225 230 235 240

Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn

245 250 255

Thr Ala Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val

260 265 270

Tyr Tyr Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly

275 280 285

Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser

290 295 300

Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala

305 310 315 320

Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val

325 330 335

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala

340 345 350

Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val

355 360 365

Pro Ser Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His

370 375 380

Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly

385 390 395 400

Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser

405 410 415

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

420 425 430

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro

435 440 445

Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

450 455 460

Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val

465 470 475 480

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

485 490 495

Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr

500 505 510

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

515 520 525

Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys

530 535 540

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

545 550 555 560

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

565 570 575

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser

580 585 590

Arg Trp Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

595 600 605

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 79

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc轻链（N-末端融合）

<400> 79

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcattgc gggccaggac 540

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 600

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 660

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 720

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 780

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 840

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 900

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 960

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 1020

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 1080

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1140

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1179

<210> 80

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -Nanoluc轻链（N-末端融合）

<400> 80

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Ile

165 170 175

Ala Gly Gln Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala

180 185 190

Ser Val Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val

195 200 205

Ser Thr Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys

210 215 220

Leu Leu Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg

225 230 235 240

Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser

245 250 255

Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr

260 265 270

His Pro Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr

275 280 285

Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu

290 295 300

Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro

305 310 315 320

Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly

325 330 335

Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr

340 345 350

Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His

355 360 365

Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val

370 375 380

Thr Lys Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 81

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2重链（C-末端融合）

<400> 81

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcattgcg ggccaggcac ctacctcgag ttctacaaag 1380

aaaacacagc tacaactgga gcatttactg ctggatttac agatgatttt gaatggaatt 1440

aataattaca agaatcccaa actcaccagg atgctcacat ttaagtttta catgcccaag 1500

aaggccacag aactgaaaca tcttcagtgt ctagaagaag aactcaaacc tctggaggaa 1560

gtgctaaatt tagctcaaag caaaaacttt cacttaagac ccagggactt aatcagcaat 1620

atcaacgtaa tagttctgga actaaaggga tctgaaacaa cattcatgtg tgaatatgct 1680

gatgagacag caaccattgt agaatttctg aacagatgga ttaccttttg tcaaagcatc 1740

atctcaacac tgact 1755

<210> 82

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2重链（C-末端融合）

<400> 82

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Ile Ala Gly Gln Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu

450 455 460

Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile

465 470 475 480

Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe

485 490 495

Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu

500 505 510

Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys

515 520 525

Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile

530 535 540

Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala

545 550 555 560

Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe

565 570 575

Cys Gln Ser Ile Ile Ser Thr Leu Thr

580 585

<210> 83

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2轻链（C-末端融合）

<400> 83

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcattgcg 660

ggccaggcac ctacctcgag ttctacaaag aaaacacagc tacaactgga gcatttactg 720

ctggatttac agatgatttt gaatggaatt aataattaca agaatcccaa actcaccagg 780

atgctcacat ttaagtttta catgcccaag aaggccacag aactgaaaca tcttcagtgt 840

ctagaagaag aactcaaacc tctggaggaa gtgctaaatt tagctcaaag caaaaacttt 900

cacttaagac ccagggactt aatcagcaat atcaacgtaa tagttctgga actaaaggga 960

tctgaaacaa cattcatgtg tgaatatgct gatgagacag caaccattgt agaatttctg 1020

aacagatgga ttaccttttg tcaaagcatc atctcaacac tgact 1065

<210> 84

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2轻链（C-末端融合）

<400> 84

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Ile Ala Gly Gln Ala Pro

210 215 220

Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu

225 230 235 240

Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro

245 250 255

Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala

260 265 270

Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu

275 280 285

Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro

290 295 300

Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly

305 310 315 320

Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile

325 330 335

Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser

340 345 350

Thr Leu Thr

355

<210> 85

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2重链（N-末端融合）

<400> 85

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg cattgcgggc 420

caggaagtcc aactggtcga aagcggtgga ggtctggtac agccaggagg aagtttgaga 480

ctgtcatgcg ctgcttccgg gtttactttc agcgactcct ggattcactg ggttcgtcag 540

gcacctggaa aggggcttga gtgggttgca tggatttccc catacggcgg ctctacatac 600

tacgctgaca gcgtcaaggg caggtttaca ataagcgccg atacctctaa aaacaccgcc 660

tacctgcaga tgaatagttt gcgagccgag gataccgccg tgtattattg tgcccggcgc 720

cattggcccg gcggcttcga ttattggggg caggggacac ttgtgacagt gtctgcagcc 780

tctaccaagg gccccagcgt gttccctctg gccccctgca gcagaagcac cagcgagtct 840

acagccgccc tgggatgcct ggtcaaggac tacttccccg agcccgtgac cgtgtcctgg 900

aactctggcg ccctgaccag cggcgtgcac acctttccag ccgtgctgca gagcagcggc 960

ctgtacagcc tgagcagcgt cgtgaccgtg cctagcagca gcctgggcac caagacctac 1020

acctgtaacg tggaccacaa gcccagcaac accaaggtgg acaagcgggt ggaatctaag 1080

tacggccctc cctgcccccc ctgcccagcc cctgaatttc tgggcggacc ctccgtgttc 1140

ctgttccccc caaagcccaa ggacaccctg atgatcagcc ggacccccga agtgacctgc 1200

gtggtggtgg acgtgtccca ggaagatccc gaggtccagt tcaactggta cgtggacggc 1260

gtggaagtgc acaacgccaa gaccaagccc agagaggaac agttcaacag cacctaccgg 1320

gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga gtacaagtgc 1380

aaggtctcca acaagggcct gcccagctcc atcgagaaaa ccatcagcaa ggccaagggc 1440

cagccccgcg agccccaggt gtacacactg ccccccagcc aggaagagat gaccaagaac 1500

caggtgtccc tgacctgcct cgtgaagggc ttctacccca gcgatatcgc cgtggaatgg 1560

gagagcaacg gccagcccga gaacaactac aagaccaccc cccctgtgct ggacagcgac 1620

ggcagcttct tcctgtactc ccggctgacc gtcgacaaga gccggtggca ggaaggcaac 1680

gtcttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1740

agcctgagcc tggga 1755

<210> 86

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2重链（N-末端融合）

<400> 86

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Ile Ala Gly Gln Glu Val Gln

130 135 140

Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg

145 150 155 160

Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His

165 170 175

Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile

180 185 190

Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg

195 200 205

Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met

210 215 220

Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg

225 230 235 240

His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr

245 250 255

Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro

260 265 270

Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val

275 280 285

Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala

290 295 300

Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly

305 310 315 320

Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly

325 330 335

Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys

340 345 350

Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

355 360 365

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

370 375 380

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

385 390 395 400

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

405 410 415

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

420 425 430

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

435 440 445

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

450 455 460

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

465 470 475 480

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu

485 490 495

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

500 505 510

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

515 520 525

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

530 535 540

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

545 550 555 560

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

565 570 575

Gln Lys Ser Leu Ser Leu Ser Leu Gly

580 585

<210> 87

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2轻链（N-末端融合）

<400> 87

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg cattgcgggc 420

caggacatcc agatgaccca atccccatcc tccttgagcg catctgtagg agacagagtc 480

accatcacct gcagggcttc acaggatgtt agcacagctg tggcttggta ccagcagaaa 540

cctggaaagg ccccaaaact gctgatatac agcgcctcat tcctgtacag cggagtgccc 600

tctcgcttta gtggctctgg gagtggcact gatttcacac tgactatttc tagtctgcag 660

cccgaagatt tcgccactta ctattgccag cagtatctct atcaccccgc cacctttggg 720

cagggcacaa aggtcgagat taagcgtacg gtggccgctc ccagcgtgtt catcttccca 780

cccagcgacg agcagctgaa gtctggcacc gccagcgtcg tgtgcctgct gaacaacttc 840

tacccccgcg aggccaaggt gcagtggaag gtggacaacg ccctgcagtc cggcaacagc 900

caggaaagcg tcaccgagca ggacagcaag gactccacct acagcctgag cagcaccctg 960

accctgagca aggccgacta cgagaagcac aaggtgtacg cctgcgaagt gacccaccag 1020

ggcctgtcca gccccgtgac caagagcttc aaccggggcg agtgc 1065

<210> 88

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGIAGQ -IL2轻链（N-末端融合）

<400> 88

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Ile Ala Gly Gln Asp Ile Gln

130 135 140

Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val

145 150 155 160

Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp

165 170 175

Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala

180 185 190

Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser

195 200 205

Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe

210 215 220

Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly

225 230 235 240

Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val

245 250 255

Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser

260 265 270

Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln

275 280 285

Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val

290 295 300

Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu

305 310 315 320

Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu

325 330 335

Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg

340 345 350

Gly Glu Cys

355

<210> 89

<211> 1875

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc重链（C-末端融合）

<400> 89

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcctgtgg gcgcagctcg agatggtctt cacactcgaa 1380

gatttcgttg gggactggcg acagacagcc ggctacaacc tggaccaagt ccttgaacag 1440

ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg taactccgat ccaaaggatt 1500

gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg tcatcatccc gtatgaaggt 1560

ctgagcggcg accaaatggg ccagatcgaa aaaattttta aggtggtgta ccctgtggat 1620

gatcatcact ttaaggtgat cctgcactat ggcacactgg taatcgacgg ggttacgccg 1680

aacatgatcg actatttcgg acggccgtat gaaggcatcg ccgtgttcga cggcaaaaag 1740

atcactgtaa cagggaccct gtggaacggc aacaaaatta tcgacgagcg cctgatcaac 1800

cccgacggct ccctgctgtt ccgagtaacc atcaacggag tgaccggctg gcggctgtgc 1860

gaacgcattc tggcg 1875

<210> 90

<211> 625

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc重链（C-末端融合）

<400> 90

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Leu Trp Ala Gln Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly

450 455 460

Asp Trp Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln

465 470 475 480

Gly Gly Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro

485 490 495

Ile Gln Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile

500 505 510

His Val Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln

515 520 525

Ile Glu Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe

530 535 540

Lys Val Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro

545 550 555 560

Asn Met Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe

565 570 575

Asp Gly Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys

580 585 590

Ile Ile Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg

595 600 605

Val Thr Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu

610 615 620

Ala

625

<210> 91

<211> 1185

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc轻链（C-末端融合）

<400> 91

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcctgtgg 660

gcgcagctcg agatggtctt cacactcgaa gatttcgttg gggactggcg acagacagcc 720

ggctacaacc tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc 780

ggggtgtccg taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc 840

gacatccatg tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa 900

aaaattttta aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat 960

ggcacactgg taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat 1020

gaaggcatcg ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc 1080

aacaaaatta tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc 1140

atcaacggag tgaccggctg gcggctgtgc gaacgcattc tggcg 1185

<210> 92

<211> 395

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc轻链（C-末端融合）

<400> 92

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Leu Trp Ala Gln Leu Glu

210 215 220

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

225 230 235 240

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

245 250 255

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

260 265 270

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

275 280 285

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

290 295 300

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

305 310 315 320

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

325 330 335

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

340 345 350

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

355 360 365

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

370 375 380

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390 395

<210> 93

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc重链（N-末端融合）

<400> 93

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcctgtg ggcgcaggaa 540

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 600

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 660

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 720

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 780

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 840

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 900

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 960

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 1020

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 1080

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1140

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1200

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1260

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1320

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1380

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1440

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1500

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1560

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1620

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1680

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1740

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1800

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1860

agcctggga 1869

<210> 94

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc重链（N-末端融合）

<400> 94

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Leu

165 170 175

Trp Ala Gln Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln

180 185 190

Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe

195 200 205

Ser Asp Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu

210 215 220

Glu Trp Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala

225 230 235 240

Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn

245 250 255

Thr Ala Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val

260 265 270

Tyr Tyr Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly

275 280 285

Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser

290 295 300

Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala

305 310 315 320

Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val

325 330 335

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala

340 345 350

Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val

355 360 365

Pro Ser Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His

370 375 380

Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly

385 390 395 400

Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser

405 410 415

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

420 425 430

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro

435 440 445

Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

450 455 460

Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val

465 470 475 480

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

485 490 495

Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr

500 505 510

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

515 520 525

Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys

530 535 540

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

545 550 555 560

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

565 570 575

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser

580 585 590

Arg Trp Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

595 600 605

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 95

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc轻链（N-末端融合）

<400> 95

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcccgc tgggcctgtg ggcgcaggac 540

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 600

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 660

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 720

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 780

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 840

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 900

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 960

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 1020

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 1080

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1140

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1179

<210> 96

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -NanoLuc轻链（N-末端融合）

<400> 96

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Pro Leu Gly Leu

165 170 175

Trp Ala Gln Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala

180 185 190

Ser Val Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val

195 200 205

Ser Thr Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys

210 215 220

Leu Leu Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg

225 230 235 240

Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser

245 250 255

Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr

260 265 270

His Pro Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr

275 280 285

Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu

290 295 300

Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro

305 310 315 320

Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly

325 330 335

Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr

340 345 350

Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His

355 360 365

Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val

370 375 380

Thr Lys Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 97

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2重链（C-末端融合）

<400> 97

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcccgct gggcctgtgg gcgcaggcac ctacctcgag ttctacaaag 1380

aaaacacagc tacaactgga gcatttactg ctggatttac agatgatttt gaatggaatt 1440

aataattaca agaatcccaa actcaccagg atgctcacat ttaagtttta catgcccaag 1500

aaggccacag aactgaaaca tcttcagtgt ctagaagaag aactcaaacc tctggaggaa 1560

gtgctaaatt tagctcaaag caaaaacttt cacttaagac ccagggactt aatcagcaat 1620

atcaacgtaa tagttctgga actaaaggga tctgaaacaa cattcatgtg tgaatatgct 1680

gatgagacag caaccattgt agaatttctg aacagatgga ttaccttttg tcaaagcatc 1740

atctcaacac tgact 1755

<210> 98

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2重链（C-末端融合）

<400> 98

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Pro Leu Gly

435 440 445

Leu Trp Ala Gln Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu

450 455 460

Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile

465 470 475 480

Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe

485 490 495

Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu

500 505 510

Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys

515 520 525

Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile

530 535 540

Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala

545 550 555 560

Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe

565 570 575

Cys Gln Ser Ile Ile Ser Thr Leu Thr

580 585

<210> 99

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2轻链（C-末端融合）

<400> 99

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcccgct gggcctgtgg 660

gcgcaggcac ctacctcgag ttctacaaag aaaacacagc tacaactgga gcatttactg 720

ctggatttac agatgatttt gaatggaatt aataattaca agaatcccaa actcaccagg 780

atgctcacat ttaagtttta catgcccaag aaggccacag aactgaaaca tcttcagtgt 840

ctagaagaag aactcaaacc tctggaggaa gtgctaaatt tagctcaaag caaaaacttt 900

cacttaagac ccagggactt aatcagcaat atcaacgtaa tagttctgga actaaaggga 960

tctgaaacaa cattcatgtg tgaatatgct gatgagacag caaccattgt agaatttctg 1020

aacagatgga ttaccttttg tcaaagcatc atctcaacac tgact 1065

<210> 100

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2轻链（C-末端融合）

<400> 100

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Pro Leu Gly Leu Trp Ala Gln Ala Pro

210 215 220

Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu

225 230 235 240

Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro

245 250 255

Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala

260 265 270

Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu

275 280 285

Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro

290 295 300

Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly

305 310 315 320

Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile

325 330 335

Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser

340 345 350

Thr Leu Thr

355

<210> 101

<211> 1755

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2重链（N-末端融合）

<400> 101

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg cctgtgggcg 420

caggaagtcc aactggtcga aagcggtgga ggtctggtac agccaggagg aagtttgaga 480

ctgtcatgcg ctgcttccgg gtttactttc agcgactcct ggattcactg ggttcgtcag 540

gcacctggaa aggggcttga gtgggttgca tggatttccc catacggcgg ctctacatac 600

tacgctgaca gcgtcaaggg caggtttaca ataagcgccg atacctctaa aaacaccgcc 660

tacctgcaga tgaatagttt gcgagccgag gataccgccg tgtattattg tgcccggcgc 720

cattggcccg gcggcttcga ttattggggg caggggacac ttgtgacagt gtctgcagcc 780

tctaccaagg gccccagcgt gttccctctg gccccctgca gcagaagcac cagcgagtct 840

acagccgccc tgggatgcct ggtcaaggac tacttccccg agcccgtgac cgtgtcctgg 900

aactctggcg ccctgaccag cggcgtgcac acctttccag ccgtgctgca gagcagcggc 960

ctgtacagcc tgagcagcgt cgtgaccgtg cctagcagca gcctgggcac caagacctac 1020

acctgtaacg tggaccacaa gcccagcaac accaaggtgg acaagcgggt ggaatctaag 1080

tacggccctc cctgcccccc ctgcccagcc cctgaatttc tgggcggacc ctccgtgttc 1140

ctgttccccc caaagcccaa ggacaccctg atgatcagcc ggacccccga agtgacctgc 1200

gtggtggtgg acgtgtccca ggaagatccc gaggtccagt tcaactggta cgtggacggc 1260

gtggaagtgc acaacgccaa gaccaagccc agagaggaac agttcaacag cacctaccgg 1320

gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga gtacaagtgc 1380

aaggtctcca acaagggcct gcccagctcc atcgagaaaa ccatcagcaa ggccaagggc 1440

cagccccgcg agccccaggt gtacacactg ccccccagcc aggaagagat gaccaagaac 1500

caggtgtccc tgacctgcct cgtgaagggc ttctacccca gcgatatcgc cgtggaatgg 1560

gagagcaacg gccagcccga gaacaactac aagaccaccc cccctgtgct ggacagcgac 1620

ggcagcttct tcctgtactc ccggctgacc gtcgacaaga gccggtggca ggaaggcaac 1680

gtcttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1740

agcctgagcc tggga 1755

<210> 102

<211> 585

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2重链（N-末端融合）

<400> 102

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Leu Trp Ala Gln Glu Val Gln

130 135 140

Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg

145 150 155 160

Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His

165 170 175

Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile

180 185 190

Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg

195 200 205

Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met

210 215 220

Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg

225 230 235 240

His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr

245 250 255

Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro

260 265 270

Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val

275 280 285

Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala

290 295 300

Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly

305 310 315 320

Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly

325 330 335

Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys

340 345 350

Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

355 360 365

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

370 375 380

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

385 390 395 400

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

405 410 415

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

420 425 430

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

435 440 445

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

450 455 460

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

465 470 475 480

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu

485 490 495

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

500 505 510

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

515 520 525

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

530 535 540

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

545 550 555 560

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

565 570 575

Gln Lys Ser Leu Ser Leu Ser Leu Gly

580 585

<210> 103

<211> 1065

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2轻链（N-末端融合）

<400> 103

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcccgctggg cctgtgggcg 420

caggacatcc agatgaccca atccccatcc tccttgagcg catctgtagg agacagagtc 480

accatcacct gcagggcttc acaggatgtt agcacagctg tggcttggta ccagcagaaa 540

cctggaaagg ccccaaaact gctgatatac agcgcctcat tcctgtacag cggagtgccc 600

tctcgcttta gtggctctgg gagtggcact gatttcacac tgactatttc tagtctgcag 660

cccgaagatt tcgccactta ctattgccag cagtatctct atcaccccgc cacctttggg 720

cagggcacaa aggtcgagat taagcgtacg gtggccgctc ccagcgtgtt catcttccca 780

cccagcgacg agcagctgaa gtctggcacc gccagcgtcg tgtgcctgct gaacaacttc 840

tacccccgcg aggccaaggt gcagtggaag gtggacaacg ccctgcagtc cggcaacagc 900

caggaaagcg tcaccgagca ggacagcaag gactccacct acagcctgag cagcaccctg 960

accctgagca aggccgacta cgagaagcac aaggtgtacg cctgcgaagt gacccaccag 1020

ggcctgtcca gccccgtgac caagagcttc aaccggggcg agtgc 1065

<210> 104

<211> 355

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GPLGLWAQ -IL2轻链（N-末端融合）

<400> 104

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Pro Leu Gly Leu Trp Ala Gln Asp Ile Gln

130 135 140

Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val

145 150 155 160

Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp

165 170 175

Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala

180 185 190

Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser

195 200 205

Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe

210 215 220

Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly

225 230 235 240

Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val

245 250 255

Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser

260 265 270

Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln

275 280 285

Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val

290 295 300

Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu

305 310 315 320

Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu

325 330 335

Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg

340 345 350

Gly Glu Cys

355

<210> 105

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc重链（C-末端融合）

<400> 105

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaccggtggg cctgattggc ctcgagatgg tcttcacact cgaagatttc 1380

gttggggact ggcgacagac agccggctac aacctggacc aagtccttga acagggaggt 1440

gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc cgatccaaag gattgtcctg 1500

agcggtgaaa atgggctgaa gatcgacatc catgtcatca tcccgtatga aggtctgagc 1560

ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg tgtaccctgt ggatgatcat 1620

cactttaagg tgatcctgca ctatggcaca ctggtaatcg acggggttac gccgaacatg 1680

atcgactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact 1740

gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat caaccccgac 1800

ggctccctgc tgttccgagt aaccatcaac ggagtgaccg gctggcggct gtgcgaacgc 1860

attctggcg 1869

<210> 106

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc重链（C-末端融合）

<400> 106

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Pro Val Gly Leu

435 440 445

Ile Gly Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp

450 455 460

Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly

465 470 475 480

Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln

485 490 495

Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val

500 505 510

Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu

515 520 525

Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe Lys Val

530 535 540

Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met

545 550 555 560

Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly

565 570 575

Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile

580 585 590

Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr

595 600 605

Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

610 615 620

<210> 107

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc轻链（C-末端融合）

<400> 107

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcccggtggg cctgattggc 660

ctcgagatgg tcttcacact cgaagatttc gttggggact ggcgacagac agccggctac 720

aacctggacc aagtccttga acagggaggt gtgtccagtt tgtttcagaa tctcggggtg 780

tccgtaactc cgatccaaag gattgtcctg agcggtgaaa atgggctgaa gatcgacatc 840

catgtcatca tcccgtatga aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt 900

tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgca ctatggcaca 960

ctggtaatcg acggggttac gccgaacatg atcgactatt tcggacggcc gtatgaaggc 1020

atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa 1080

attatcgacg agcgcctgat caaccccgac ggctccctgc tgttccgagt aaccatcaac 1140

ggagtgaccg gctggcggct gtgcgaacgc attctggcg 1179

<210> 108

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc轻链（C-末端融合）

<400> 108

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Pro Val Gly Leu Ile Gly Leu Glu Met Val

210 215 220

Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala Gly Tyr

225 230 235 240

Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu Phe Gln

245 250 255

Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu Ser Gly

260 265 270

Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr Glu Gly

275 280 285

Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys Val Val

290 295 300

Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr Gly Thr

305 310 315 320

Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe Gly Arg

325 330 335

Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr Val Thr

340 345 350

Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu Ile Asn

355 360 365

Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val Thr Gly

370 375 380

Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390

<210> 109

<211> 1863

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc重链（N-末端融合）

<400> 109

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgccggtgg gcctgattgg cgaagtccaa 540

ctggtcgaaa gcggtggagg tctggtacag ccaggaggaa gtttgagact gtcatgcgct 600

gcttccgggt ttactttcag cgactcctgg attcactggg ttcgtcaggc acctggaaag 660

gggcttgagt gggttgcatg gatttcccca tacggcggct ctacatacta cgctgacagc 720

gtcaagggca ggtttacaat aagcgccgat acctctaaaa acaccgccta cctgcagatg 780

aatagtttgc gagccgagga taccgccgtg tattattgtg cccggcgcca ttggcccggc 840

ggcttcgatt attgggggca ggggacactt gtgacagtgt ctgcagcctc taccaagggc 900

cccagcgtgt tccctctggc cccctgcagc agaagcacca gcgagtctac agccgccctg 960

ggatgcctgg tcaaggacta cttccccgag cccgtgaccg tgtcctggaa ctctggcgcc 1020

ctgaccagcg gcgtgcacac ctttccagcc gtgctgcaga gcagcggcct gtacagcctg 1080

agcagcgtcg tgaccgtgcc tagcagcagc ctgggcacca agacctacac ctgtaacgtg 1140

gaccacaagc ccagcaacac caaggtggac aagcgggtgg aatctaagta cggccctccc 1200

tgccccccct gcccagcccc tgaatttctg ggcggaccct ccgtgttcct gttcccccca 1260

aagcccaagg acaccctgat gatcagccgg acccccgaag tgacctgcgt ggtggtggac 1320

gtgtcccagg aagatcccga ggtccagttc aactggtacg tggacggcgt ggaagtgcac 1380

aacgccaaga ccaagcccag agaggaacag ttcaacagca cctaccgggt ggtgtccgtg 1440

ctgaccgtgc tgcaccagga ctggctgaac ggcaaagagt acaagtgcaa ggtctccaac 1500

aagggcctgc ccagctccat cgagaaaacc atcagcaagg ccaagggcca gccccgcgag 1560

ccccaggtgt acacactgcc ccccagccag gaagagatga ccaagaacca ggtgtccctg 1620

acctgcctcg tgaagggctt ctaccccagc gatatcgccg tggaatggga gagcaacggc 1680

cagcccgaga acaactacaa gaccaccccc cctgtgctgg acagcgacgg cagcttcttc 1740

ctgtactccc ggctgaccgt cgacaagagc cggtggcagg aaggcaacgt cttcagctgc 1800

agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag cctgagcctg 1860

gga 1863

<210> 110

<211> 621

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc重链（N-末端融合）

<400> 110

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Pro Val Gly Leu Ile

165 170 175

Gly Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly

180 185 190

Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp

195 200 205

Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp

210 215 220

Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser

225 230 235 240

Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala

245 250 255

Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr

260 265 270

Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly

275 280 285

Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe

290 295 300

Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu

305 310 315 320

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp

325 330 335

Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu

340 345 350

Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser

355 360 365

Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro

370 375 380

Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro

385 390 395 400

Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe

405 410 415

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

420 425 430

Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val

435 440 445

Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

450 455 460

Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val

465 470 475 480

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

485 490 495

Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser

500 505 510

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

515 520 525

Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

530 535 540

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

545 550 555 560

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

565 570 575

Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp

580 585 590

Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

595 600 605

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 111

<211> 1173

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc轻链（N-末端融合）

<400> 111

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgccggtgg gcctgattgg cgacatccag 540

atgacccaat ccccatcctc cttgagcgca tctgtaggag acagagtcac catcacctgc 600

agggcttcac aggatgttag cacagctgtg gcttggtacc agcagaaacc tggaaaggcc 660

ccaaaactgc tgatatacag cgcctcattc ctgtacagcg gagtgccctc tcgctttagt 720

ggctctggga gtggcactga tttcacactg actatttcta gtctgcagcc cgaagatttc 780

gccacttact attgccagca gtatctctat caccccgcca cctttgggca gggcacaaag 840

gtcgagatta agcgtacggt ggccgctccc agcgtgttca tcttcccacc cagcgacgag 900

cagctgaagt ctggcaccgc cagcgtcgtg tgcctgctga acaacttcta cccccgcgag 960

gccaaggtgc agtggaaggt ggacaacgcc ctgcagtccg gcaacagcca ggaaagcgtc 1020

accgagcagg acagcaagga ctccacctac agcctgagca gcaccctgac cctgagcaag 1080

gccgactacg agaagcacaa ggtgtacgcc tgcgaagtga cccaccaggg cctgtccagc 1140

cccgtgacca agagcttcaa ccggggcgag tgc 1173

<210> 112

<211> 391

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG -NanoLuc轻链（N-末端融合）

<400> 112

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Pro Val Gly Leu Ile

165 170 175

Gly Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val

180 185 190

Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr

195 200 205

Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu

210 215 220

Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser

225 230 235 240

Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln

245 250 255

Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro

260 265 270

Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala

275 280 285

Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser

290 295 300

Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu

305 310 315 320

Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser

325 330 335

Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu

340 345 350

Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val

355 360 365

Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys

370 375 380

Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 113

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2重链（C-末端融合）

<400> 113

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaccggtggg cctgattggc gcacctacct cgagttctac aaagaaaaca 1380

cagctacaac tggagcattt actgctggat ttacagatga ttttgaatgg aattaataat 1440

tacaagaatc ccaaactcac caggatgctc acatttaagt tttacatgcc caagaaggcc 1500

acagaactga aacatcttca gtgtctagaa gaagaactca aacctctgga ggaagtgcta 1560

aatttagctc aaagcaaaaa ctttcactta agacccaggg acttaatcag caatatcaac 1620

gtaatagttc tggaactaaa gggatctgaa acaacattca tgtgtgaata tgctgatgag 1680

acagcaacca ttgtagaatt tctgaacaga tggattacct tttgtcaaag catcatctca 1740

acactgact 1749

<210> 114

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2重链（C-末端融合）

<400> 114

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Pro Val Gly Leu

435 440 445

Ile Gly Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu

450 455 460

Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn

465 470 475 480

Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met

485 490 495

Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu

500 505 510

Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe

515 520 525

His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu

530 535 540

Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu

545 550 555 560

Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln

565 570 575

Ser Ile Ile Ser Thr Leu Thr

580

<210> 115

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2轻链（C-末端融合）

<400> 115

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcccggtggg cctgattggc 660

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 720

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 780

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 840

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 900

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 960

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 1020

tggattacct tttgtcaaag catcatctca acactgact 1059

<210> 116

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2轻链（C-末端融合）

<400> 116

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Pro Val Gly Leu Ile Gly Ala Pro Thr Ser

210 215 220

Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp

225 230 235 240

Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu

245 250 255

Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr Glu

260 265 270

Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu Glu Glu

275 280 285

Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro Arg Asp

290 295 300

Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly Ser Glu

305 310 315 320

Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu

325 330 335

Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu

340 345 350

Thr

<210> 117

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2重链（N-末端融合）

<400> 117

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactc cggtgggcct gattggcgaa 420

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 480

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 540

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 600

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 660

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 720

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 780

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 840

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 900

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 960

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1020

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1080

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1140

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1200

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1260

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1320

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1380

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1440

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1500

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1560

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1620

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1680

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1740

agcctggga 1749

<210> 118

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2重链（N-末端融合）

<400> 118

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Pro Val Gly Leu Ile Gly Glu Val Gln Leu Val

130 135 140

Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser

145 150 155 160

Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His Trp Val

165 170 175

Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile Ser Pro

180 185 190

Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr

195 200 205

Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met Asn Ser

210 215 220

Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg His Trp

225 230 235 240

Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser

245 250 255

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser

260 265 270

Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

275 280 285

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

290 295 300

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

305 310 315 320

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys

325 330 335

Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp

340 345 350

Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala

355 360 365

Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro

370 375 380

Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val

385 390 395 400

Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val

405 410 415

Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln

420 425 430

Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln

435 440 445

Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly

450 455 460

Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro

465 470 475 480

Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr

485 490 495

Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser

500 505 510

Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr

515 520 525

Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr

530 535 540

Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe

545 550 555 560

Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys

565 570 575

Ser Leu Ser Leu Ser Leu Gly

580

<210> 119

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2轻链（N-末端融合）

<400> 119

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactc cggtgggcct gattggcgac 420

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 480

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 540

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 600

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 660

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 720

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 780

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 840

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 900

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 960

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1020

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1059

<210> 120

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PVGLIG-IL2轻链（N-末端融合）

<400> 120

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Pro Val Gly Leu Ile Gly Asp Ile Gln Met Thr

130 135 140

Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile

145 150 155 160

Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp Tyr Gln

165 170 175

Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala Ser Phe

180 185 190

Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr

195 200 205

Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr

210 215 220

Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly Gln Gly

225 230 235 240

Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile

245 250 255

Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser Val Val

260 265 270

Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys

275 280 285

Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val Thr Glu

290 295 300

Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu

305 310 315 320

Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr

325 330 335

His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu

340 345 350

Cys

<210> 121

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc重链（C-末端融合）

<400> 121

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaccgctggg cctggcgggc ctcgagatgg tcttcacact cgaagatttc 1380

gttggggact ggcgacagac agccggctac aacctggacc aagtccttga acagggaggt 1440

gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc cgatccaaag gattgtcctg 1500

agcggtgaaa atgggctgaa gatcgacatc catgtcatca tcccgtatga aggtctgagc 1560

ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg tgtaccctgt ggatgatcat 1620

cactttaagg tgatcctgca ctatggcaca ctggtaatcg acggggttac gccgaacatg 1680

atcgactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact 1740

gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat caaccccgac 1800

ggctccctgc tgttccgagt aaccatcaac ggagtgaccg gctggcggct gtgcgaacgc 1860

attctggcg 1869

<210> 122

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc重链（C-末端融合）

<400> 122

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Pro Leu Gly Leu

435 440 445

Ala Gly Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp

450 455 460

Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly

465 470 475 480

Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln

485 490 495

Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val

500 505 510

Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu

515 520 525

Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe Lys Val

530 535 540

Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met

545 550 555 560

Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly

565 570 575

Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile

580 585 590

Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr

595 600 605

Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

610 615 620

<210> 123

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc轻链（C-末端融合）

<400> 123

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcccgctggg cctggcgggc 660

ctcgagatgg tcttcacact cgaagatttc gttggggact ggcgacagac agccggctac 720

aacctggacc aagtccttga acagggaggt gtgtccagtt tgtttcagaa tctcggggtg 780

tccgtaactc cgatccaaag gattgtcctg agcggtgaaa atgggctgaa gatcgacatc 840

catgtcatca tcccgtatga aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt 900

tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgca ctatggcaca 960

ctggtaatcg acggggttac gccgaacatg atcgactatt tcggacggcc gtatgaaggc 1020

atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa 1080

attatcgacg agcgcctgat caaccccgac ggctccctgc tgttccgagt aaccatcaac 1140

ggagtgaccg gctggcggct gtgcgaacgc attctggcg 1179

<210> 124

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc轻链（C-末端融合）

<400> 124

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Pro Leu Gly Leu Ala Gly Leu Glu Met Val

210 215 220

Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala Gly Tyr

225 230 235 240

Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu Phe Gln

245 250 255

Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu Ser Gly

260 265 270

Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr Glu Gly

275 280 285

Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys Val Val

290 295 300

Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr Gly Thr

305 310 315 320

Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe Gly Arg

325 330 335

Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr Val Thr

340 345 350

Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu Ile Asn

355 360 365

Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val Thr Gly

370 375 380

Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390

<210> 125

<211> 1863

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc重链（N-末端融合）

<400> 125

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgccgctgg gcctggcggg cgaagtccaa 540

ctggtcgaaa gcggtggagg tctggtacag ccaggaggaa gtttgagact gtcatgcgct 600

gcttccgggt ttactttcag cgactcctgg attcactggg ttcgtcaggc acctggaaag 660

gggcttgagt gggttgcatg gatttcccca tacggcggct ctacatacta cgctgacagc 720

gtcaagggca ggtttacaat aagcgccgat acctctaaaa acaccgccta cctgcagatg 780

aatagtttgc gagccgagga taccgccgtg tattattgtg cccggcgcca ttggcccggc 840

ggcttcgatt attgggggca ggggacactt gtgacagtgt ctgcagcctc taccaagggc 900

cccagcgtgt tccctctggc cccctgcagc agaagcacca gcgagtctac agccgccctg 960

ggatgcctgg tcaaggacta cttccccgag cccgtgaccg tgtcctggaa ctctggcgcc 1020

ctgaccagcg gcgtgcacac ctttccagcc gtgctgcaga gcagcggcct gtacagcctg 1080

agcagcgtcg tgaccgtgcc tagcagcagc ctgggcacca agacctacac ctgtaacgtg 1140

gaccacaagc ccagcaacac caaggtggac aagcgggtgg aatctaagta cggccctccc 1200

tgccccccct gcccagcccc tgaatttctg ggcggaccct ccgtgttcct gttcccccca 1260

aagcccaagg acaccctgat gatcagccgg acccccgaag tgacctgcgt ggtggtggac 1320

gtgtcccagg aagatcccga ggtccagttc aactggtacg tggacggcgt ggaagtgcac 1380

aacgccaaga ccaagcccag agaggaacag ttcaacagca cctaccgggt ggtgtccgtg 1440

ctgaccgtgc tgcaccagga ctggctgaac ggcaaagagt acaagtgcaa ggtctccaac 1500

aagggcctgc ccagctccat cgagaaaacc atcagcaagg ccaagggcca gccccgcgag 1560

ccccaggtgt acacactgcc ccccagccag gaagagatga ccaagaacca ggtgtccctg 1620

acctgcctcg tgaagggctt ctaccccagc gatatcgccg tggaatggga gagcaacggc 1680

cagcccgaga acaactacaa gaccaccccc cctgtgctgg acagcgacgg cagcttcttc 1740

ctgtactccc ggctgaccgt cgacaagagc cggtggcagg aaggcaacgt cttcagctgc 1800

agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag cctgagcctg 1860

gga 1863

<210> 126

<211> 621

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc重链（N-末端融合）

<400> 126

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Pro Leu Gly Leu Ala

165 170 175

Gly Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly

180 185 190

Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp

195 200 205

Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp

210 215 220

Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser

225 230 235 240

Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala

245 250 255

Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr

260 265 270

Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly

275 280 285

Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe

290 295 300

Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu

305 310 315 320

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp

325 330 335

Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu

340 345 350

Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser

355 360 365

Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro

370 375 380

Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro

385 390 395 400

Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe

405 410 415

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

420 425 430

Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val

435 440 445

Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

450 455 460

Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val

465 470 475 480

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

485 490 495

Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser

500 505 510

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

515 520 525

Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

530 535 540

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

545 550 555 560

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

565 570 575

Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp

580 585 590

Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

595 600 605

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 127

<211> 1173

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc轻链（N-末端融合）

<400> 127

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgccgctgg gcctggcggg cgacatccag 540

atgacccaat ccccatcctc cttgagcgca tctgtaggag acagagtcac catcacctgc 600

agggcttcac aggatgttag cacagctgtg gcttggtacc agcagaaacc tggaaaggcc 660

ccaaaactgc tgatatacag cgcctcattc ctgtacagcg gagtgccctc tcgctttagt 720

ggctctggga gtggcactga tttcacactg actatttcta gtctgcagcc cgaagatttc 780

gccacttact attgccagca gtatctctat caccccgcca cctttgggca gggcacaaag 840

gtcgagatta agcgtacggt ggccgctccc agcgtgttca tcttcccacc cagcgacgag 900

cagctgaagt ctggcaccgc cagcgtcgtg tgcctgctga acaacttcta cccccgcgag 960

gccaaggtgc agtggaaggt ggacaacgcc ctgcagtccg gcaacagcca ggaaagcgtc 1020

accgagcagg acagcaagga ctccacctac agcctgagca gcaccctgac cctgagcaag 1080

gccgactacg agaagcacaa ggtgtacgcc tgcgaagtga cccaccaggg cctgtccagc 1140

cccgtgacca agagcttcaa ccggggcgag tgc 1173

<210> 128

<211> 391

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-NanoLuc轻链（N-末端融合）

<400> 128

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Pro Leu Gly Leu Ala

165 170 175

Gly Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val

180 185 190

Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr

195 200 205

Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu

210 215 220

Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser

225 230 235 240

Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln

245 250 255

Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro

260 265 270

Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala

275 280 285

Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser

290 295 300

Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu

305 310 315 320

Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser

325 330 335

Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu

340 345 350

Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val

355 360 365

Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys

370 375 380

Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 129

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2重链（C-末端融合）

<400> 129

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaccgctggg cctggcgggc gcacctacct cgagttctac aaagaaaaca 1380

cagctacaac tggagcattt actgctggat ttacagatga ttttgaatgg aattaataat 1440

tacaagaatc ccaaactcac caggatgctc acatttaagt tttacatgcc caagaaggcc 1500

acagaactga aacatcttca gtgtctagaa gaagaactca aacctctgga ggaagtgcta 1560

aatttagctc aaagcaaaaa ctttcactta agacccaggg acttaatcag caatatcaac 1620

gtaatagttc tggaactaaa gggatctgaa acaacattca tgtgtgaata tgctgatgag 1680

acagcaacca ttgtagaatt tctgaacaga tggattacct tttgtcaaag catcatctca 1740

acactgact 1749

<210> 130

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2重链（C-末端融合）

<400> 130

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Pro Leu Gly Leu

435 440 445

Ala Gly Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu

450 455 460

Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn

465 470 475 480

Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met

485 490 495

Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu

500 505 510

Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe

515 520 525

His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu

530 535 540

Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu

545 550 555 560

Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln

565 570 575

Ser Ile Ile Ser Thr Leu Thr

580

<210> 131

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2轻链（C-末端融合）

<400> 131

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcccgctggg cctggcgggc 660

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 720

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 780

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 840

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 900

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 960

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 1020

tggattacct tttgtcaaag catcatctca acactgact 1059

<210> 132

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2轻链（C-末端融合）

<400> 132

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Pro Leu Gly Leu Ala Gly Ala Pro Thr Ser

210 215 220

Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp

225 230 235 240

Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu

245 250 255

Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr Glu

260 265 270

Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu Glu Glu

275 280 285

Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro Arg Asp

290 295 300

Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly Ser Glu

305 310 315 320

Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu

325 330 335

Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu

340 345 350

Thr

<210> 133

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2重链（N-末端融合）

<400> 133

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactc cgctgggcct ggcgggcgaa 420

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 480

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 540

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 600

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 660

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 720

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 780

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 840

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 900

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 960

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1020

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1080

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1140

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1200

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1260

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1320

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1380

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1440

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1500

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1560

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1620

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1680

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1740

agcctggga 1749

<210> 134

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2重链（N-末端融合）

<400> 134

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Pro Leu Gly Leu Ala Gly Glu Val Gln Leu Val

130 135 140

Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser

145 150 155 160

Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His Trp Val

165 170 175

Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile Ser Pro

180 185 190

Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr

195 200 205

Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met Asn Ser

210 215 220

Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg His Trp

225 230 235 240

Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser

245 250 255

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser

260 265 270

Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

275 280 285

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

290 295 300

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

305 310 315 320

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys

325 330 335

Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp

340 345 350

Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala

355 360 365

Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro

370 375 380

Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val

385 390 395 400

Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val

405 410 415

Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln

420 425 430

Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln

435 440 445

Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly

450 455 460

Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro

465 470 475 480

Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr

485 490 495

Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser

500 505 510

Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr

515 520 525

Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr

530 535 540

Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe

545 550 555 560

Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys

565 570 575

Ser Leu Ser Leu Ser Leu Gly

580

<210> 135

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2轻链（N-末端融合）

<400> 135

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactc cgctgggcct ggcgggcgac 420

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 480

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 540

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 600

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 660

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 720

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 780

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 840

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 900

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 960

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1020

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1059

<210> 136

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-PLGLAG-IL2轻链（N-末端融合）

<400> 136

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Pro Leu Gly Leu Ala Gly Asp Ile Gln Met Thr

130 135 140

Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile

145 150 155 160

Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp Tyr Gln

165 170 175

Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala Ser Phe

180 185 190

Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr

195 200 205

Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr

210 215 220

Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly Gln Gly

225 230 235 240

Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile

245 250 255

Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser Val Val

260 265 270

Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys

275 280 285

Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val Thr Glu

290 295 300

Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu

305 310 315 320

Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr

325 330 335

His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu

340 345 350

Cys

<210> 137

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc重链（C-末端融合）

<400> 137

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcattgt gggaccgctg ctcgagatgg tcttcacact cgaagatttc 1380

gttggggact ggcgacagac agccggctac aacctggacc aagtccttga acagggaggt 1440

gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc cgatccaaag gattgtcctg 1500

agcggtgaaa atgggctgaa gatcgacatc catgtcatca tcccgtatga aggtctgagc 1560

ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg tgtaccctgt ggatgatcat 1620

cactttaagg tgatcctgca ctatggcaca ctggtaatcg acggggttac gccgaacatg 1680

atcgactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact 1740

gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat caaccccgac 1800

ggctccctgc tgttccgagt aaccatcaac ggagtgaccg gctggcggct gtgcgaacgc 1860

attctggcg 1869

<210> 138

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc重链（C-末端融合）

<400> 138

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Ile Val Gly

435 440 445

Pro Leu Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp

450 455 460

Arg Gln Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly

465 470 475 480

Val Ser Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln

485 490 495

Arg Ile Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val

500 505 510

Ile Ile Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu

515 520 525

Lys Ile Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe Lys Val

530 535 540

Ile Leu His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met

545 550 555 560

Ile Asp Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly

565 570 575

Lys Lys Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile

580 585 590

Asp Glu Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr

595 600 605

Ile Asn Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

610 615 620

<210> 139

<211> 1179

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc轻链（C-末端融合）

<400> 139

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcattgt gggaccgctg 660

ctcgagatgg tcttcacact cgaagatttc gttggggact ggcgacagac agccggctac 720

aacctggacc aagtccttga acagggaggt gtgtccagtt tgtttcagaa tctcggggtg 780

tccgtaactc cgatccaaag gattgtcctg agcggtgaaa atgggctgaa gatcgacatc 840

catgtcatca tcccgtatga aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt 900

tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgca ctatggcaca 960

ctggtaatcg acggggttac gccgaacatg atcgactatt tcggacggcc gtatgaaggc 1020

atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa 1080

attatcgacg agcgcctgat caaccccgac ggctccctgc tgttccgagt aaccatcaac 1140

ggagtgaccg gctggcggct gtgcgaacgc attctggcg 1179

<210> 140

<211> 393

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc轻链（C-末端融合）

<400> 140

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Ile Val Gly Pro Leu Leu Glu Met Val

210 215 220

Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala Gly Tyr

225 230 235 240

Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu Phe Gln

245 250 255

Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu Ser Gly

260 265 270

Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr Glu Gly

275 280 285

Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys Val Val

290 295 300

Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr Gly Thr

305 310 315 320

Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe Gly Arg

325 330 335

Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr Val Thr

340 345 350

Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu Ile Asn

355 360 365

Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val Thr Gly

370 375 380

Trp Arg Leu Cys Glu Arg Ile Leu Ala

385 390

<210> 141

<211> 1863

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc重链（N-末端融合）

<400> 141

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcattg tgggaccgct ggaagtccaa 540

ctggtcgaaa gcggtggagg tctggtacag ccaggaggaa gtttgagact gtcatgcgct 600

gcttccgggt ttactttcag cgactcctgg attcactggg ttcgtcaggc acctggaaag 660

gggcttgagt gggttgcatg gatttcccca tacggcggct ctacatacta cgctgacagc 720

gtcaagggca ggtttacaat aagcgccgat acctctaaaa acaccgccta cctgcagatg 780

aatagtttgc gagccgagga taccgccgtg tattattgtg cccggcgcca ttggcccggc 840

ggcttcgatt attgggggca ggggacactt gtgacagtgt ctgcagcctc taccaagggc 900

cccagcgtgt tccctctggc cccctgcagc agaagcacca gcgagtctac agccgccctg 960

ggatgcctgg tcaaggacta cttccccgag cccgtgaccg tgtcctggaa ctctggcgcc 1020

ctgaccagcg gcgtgcacac ctttccagcc gtgctgcaga gcagcggcct gtacagcctg 1080

agcagcgtcg tgaccgtgcc tagcagcagc ctgggcacca agacctacac ctgtaacgtg 1140

gaccacaagc ccagcaacac caaggtggac aagcgggtgg aatctaagta cggccctccc 1200

tgccccccct gcccagcccc tgaatttctg ggcggaccct ccgtgttcct gttcccccca 1260

aagcccaagg acaccctgat gatcagccgg acccccgaag tgacctgcgt ggtggtggac 1320

gtgtcccagg aagatcccga ggtccagttc aactggtacg tggacggcgt ggaagtgcac 1380

aacgccaaga ccaagcccag agaggaacag ttcaacagca cctaccgggt ggtgtccgtg 1440

ctgaccgtgc tgcaccagga ctggctgaac ggcaaagagt acaagtgcaa ggtctccaac 1500

aagggcctgc ccagctccat cgagaaaacc atcagcaagg ccaagggcca gccccgcgag 1560

ccccaggtgt acacactgcc ccccagccag gaagagatga ccaagaacca ggtgtccctg 1620

acctgcctcg tgaagggctt ctaccccagc gatatcgccg tggaatggga gagcaacggc 1680

cagcccgaga acaactacaa gaccaccccc cctgtgctgg acagcgacgg cagcttcttc 1740

ctgtactccc ggctgaccgt cgacaagagc cggtggcagg aaggcaacgt cttcagctgc 1800

agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag cctgagcctg 1860

gga 1863

<210> 142

<211> 621

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc重链（N-末端融合）

<400> 142

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Ile Val Gly Pro

165 170 175

Leu Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly

180 185 190

Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp

195 200 205

Ser Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp

210 215 220

Val Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser

225 230 235 240

Val Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala

245 250 255

Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr

260 265 270

Cys Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly

275 280 285

Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe

290 295 300

Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu

305 310 315 320

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp

325 330 335

Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu

340 345 350

Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser

355 360 365

Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro

370 375 380

Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro

385 390 395 400

Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe

405 410 415

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

420 425 430

Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val

435 440 445

Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

450 455 460

Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val

465 470 475 480

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

485 490 495

Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser

500 505 510

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

515 520 525

Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

530 535 540

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

545 550 555 560

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

565 570 575

Gly Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp

580 585 590

Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

595 600 605

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly

610 615 620

<210> 143

<211> 1173

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc轻链（N-末端融合）

<400> 143

atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg 60

gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta 120

actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc 180

atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag 240

gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta 300

atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc 360

gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc 420

gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg 480

accggctggc ggctgtgcga acgcattctg gcgggcattg tgggaccgct ggacatccag 540

atgacccaat ccccatcctc cttgagcgca tctgtaggag acagagtcac catcacctgc 600

agggcttcac aggatgttag cacagctgtg gcttggtacc agcagaaacc tggaaaggcc 660

ccaaaactgc tgatatacag cgcctcattc ctgtacagcg gagtgccctc tcgctttagt 720

ggctctggga gtggcactga tttcacactg actatttcta gtctgcagcc cgaagatttc 780

gccacttact attgccagca gtatctctat caccccgcca cctttgggca gggcacaaag 840

gtcgagatta agcgtacggt ggccgctccc agcgtgttca tcttcccacc cagcgacgag 900

cagctgaagt ctggcaccgc cagcgtcgtg tgcctgctga acaacttcta cccccgcgag 960

gccaaggtgc agtggaaggt ggacaacgcc ctgcagtccg gcaacagcca ggaaagcgtc 1020

accgagcagg acagcaagga ctccacctac agcctgagca gcaccctgac cctgagcaag 1080

gccgactacg agaagcacaa ggtgtacgcc tgcgaagtga cccaccaggg cctgtccagc 1140

cccgtgacca agagcttcaa ccggggcgag tgc 1173

<210> 144

<211> 391

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-NanoLuc轻链（N-末端融合）

<400> 144

Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln Thr Ala

1 5 10 15

Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser Ser Leu

20 25 30

Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile Val Leu

35 40 45

Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile Pro Tyr

50 55 60

Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile Phe Lys

65 70 75 80

Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu His Tyr

85 90 95

Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp Tyr Phe

100 105 110

Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys Ile Thr

115 120 125

Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu Arg Leu

130 135 140

Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn Gly Val

145 150 155 160

Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala Gly Ile Val Gly Pro

165 170 175

Leu Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val

180 185 190

Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr

195 200 205

Ala Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu

210 215 220

Ile Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser

225 230 235 240

Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln

245 250 255

Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro

260 265 270

Ala Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala

275 280 285

Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser

290 295 300

Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu

305 310 315 320

Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser

325 330 335

Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu

340 345 350

Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val

355 360 365

Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys

370 375 380

Ser Phe Asn Arg Gly Glu Cys

385 390

<210> 145

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2重链（C-末端融合）

<400> 145

gaagtccaac tggtcgaaag cggtggaggt ctggtacagc caggaggaag tttgagactg 60

tcatgcgctg cttccgggtt tactttcagc gactcctgga ttcactgggt tcgtcaggca 120

cctggaaagg ggcttgagtg ggttgcatgg atttccccat acggcggctc tacatactac 180

gctgacagcg tcaagggcag gtttacaata agcgccgata cctctaaaaa caccgcctac 240

ctgcagatga atagtttgcg agccgaggat accgccgtgt attattgtgc ccggcgccat 300

tggcccggcg gcttcgatta ttgggggcag gggacacttg tgacagtgtc tgcagcctct 360

accaagggcc ccagcgtgtt ccctctggcc ccctgcagca gaagcaccag cgagtctaca 420

gccgccctgg gatgcctggt caaggactac ttccccgagc ccgtgaccgt gtcctggaac 480

tctggcgccc tgaccagcgg cgtgcacacc tttccagccg tgctgcagag cagcggcctg 540

tacagcctga gcagcgtcgt gaccgtgcct agcagcagcc tgggcaccaa gacctacacc 600

tgtaacgtgg accacaagcc cagcaacacc aaggtggaca agcgggtgga atctaagtac 660

ggccctccct gccccccctg cccagcccct gaatttctgg gcggaccctc cgtgttcctg 720

ttccccccaa agcccaagga caccctgatg atcagccgga cccccgaagt gacctgcgtg 780

gtggtggacg tgtcccagga agatcccgag gtccagttca actggtacgt ggacggcgtg 840

gaagtgcaca acgccaagac caagcccaga gaggaacagt tcaacagcac ctaccgggtg 900

gtgtccgtgc tgaccgtgct gcaccaggac tggctgaacg gcaaagagta caagtgcaag 960

gtctccaaca agggcctgcc cagctccatc gagaaaacca tcagcaaggc caagggccag 1020

ccccgcgagc cccaggtgta cacactgccc cccagccagg aagagatgac caagaaccag 1080

gtgtccctga cctgcctcgt gaagggcttc taccccagcg atatcgccgt ggaatgggag 1140

agcaacggcc agcccgagaa caactacaag accacccccc ctgtgctgga cagcgacggc 1200

agcttcttcc tgtactcccg gctgaccgtc gacaagagcc ggtggcagga aggcaacgtc 1260

ttcagctgca gcgtgatgca cgaggccctg cacaaccact acacccagaa gtccctgagc 1320

ctgagcctgg gaggcattgt gggaccgctg gcacctacct cgagttctac aaagaaaaca 1380

cagctacaac tggagcattt actgctggat ttacagatga ttttgaatgg aattaataat 1440

tacaagaatc ccaaactcac caggatgctc acatttaagt tttacatgcc caagaaggcc 1500

acagaactga aacatcttca gtgtctagaa gaagaactca aacctctgga ggaagtgcta 1560

aatttagctc aaagcaaaaa ctttcactta agacccaggg acttaatcag caatatcaac 1620

gtaatagttc tggaactaaa gggatctgaa acaacattca tgtgtgaata tgctgatgag 1680

acagcaacca ttgtagaatt tctgaacaga tggattacct tttgtcaaag catcatctca 1740

acactgact 1749

<210> 146

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2重链（C-末端融合）

<400> 146

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser

20 25 30

Trp Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Trp Ile Ser Pro Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Arg His Trp Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr

100 105 110

Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro

115 120 125

Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly

130 135 140

Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn

145 150 155 160

Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln

165 170 175

Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser

180 185 190

Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser

195 200 205

Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys

210 215 220

Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu

225 230 235 240

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu

245 250 255

Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln

260 265 270

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys

275 280 285

Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu

290 295 300

Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys

305 310 315 320

Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys

325 330 335

Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser

340 345 350

Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys

355 360 365

Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln

370 375 380

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly

385 390 395 400

Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln

405 410 415

Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn

420 425 430

His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Gly Ile Val Gly

435 440 445

Pro Leu Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu

450 455 460

Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn

465 470 475 480

Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met

485 490 495

Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu

500 505 510

Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe

515 520 525

His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu

530 535 540

Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu

545 550 555 560

Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln

565 570 575

Ser Ile Ile Ser Thr Leu Thr

580

<210> 147

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2轻链（C-末端融合）

<400> 147

gacatccaga tgacccaatc cccatcctcc ttgagcgcat ctgtaggaga cagagtcacc 60

atcacctgca gggcttcaca ggatgttagc acagctgtgg cttggtacca gcagaaacct 120

ggaaaggccc caaaactgct gatatacagc gcctcattcc tgtacagcgg agtgccctct 180

cgctttagtg gctctgggag tggcactgat ttcacactga ctatttctag tctgcagccc 240

gaagatttcg ccacttacta ttgccagcag tatctctatc accccgccac ctttgggcag 300

ggcacaaagg tcgagattaa gcgtacggtg gccgctccca gcgtgttcat cttcccaccc 360

agcgacgagc agctgaagtc tggcaccgcc agcgtcgtgt gcctgctgaa caacttctac 420

ccccgcgagg ccaaggtgca gtggaaggtg gacaacgccc tgcagtccgg caacagccag 480

gaaagcgtca ccgagcagga cagcaaggac tccacctaca gcctgagcag caccctgacc 540

ctgagcaagg ccgactacga gaagcacaag gtgtacgcct gcgaagtgac ccaccagggc 600

ctgtccagcc ccgtgaccaa gagcttcaac cggggcgagt gcggcattgt gggaccgctg 660

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 720

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 780

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 840

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 900

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 960

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 1020

tggattacct tttgtcaaag catcatctca acactgact 1059

<210> 148

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2轻链（C-末端融合）

<400> 148

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala

20 25 30

Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Ser Ala Ser Phe Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala

85 90 95

Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys Gly Ile Val Gly Pro Leu Ala Pro Thr Ser

210 215 220

Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp

225 230 235 240

Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu

245 250 255

Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr Glu

260 265 270

Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu Glu Glu

275 280 285

Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro Arg Asp

290 295 300

Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly Ser Glu

305 310 315 320

Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu

325 330 335

Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu

340 345 350

Thr

<210> 149

<211> 1749

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2重链（N-末端融合）

<400> 149

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcattgtggg accgctggaa 420

gtccaactgg tcgaaagcgg tggaggtctg gtacagccag gaggaagttt gagactgtca 480

tgcgctgctt ccgggtttac tttcagcgac tcctggattc actgggttcg tcaggcacct 540

ggaaaggggc ttgagtgggt tgcatggatt tccccatacg gcggctctac atactacgct 600

gacagcgtca agggcaggtt tacaataagc gccgatacct ctaaaaacac cgcctacctg 660

cagatgaata gtttgcgagc cgaggatacc gccgtgtatt attgtgcccg gcgccattgg 720

cccggcggct tcgattattg ggggcagggg acacttgtga cagtgtctgc agcctctacc 780

aagggcccca gcgtgttccc tctggccccc tgcagcagaa gcaccagcga gtctacagcc 840

gccctgggat gcctggtcaa ggactacttc cccgagcccg tgaccgtgtc ctggaactct 900

ggcgccctga ccagcggcgt gcacaccttt ccagccgtgc tgcagagcag cggcctgtac 960

agcctgagca gcgtcgtgac cgtgcctagc agcagcctgg gcaccaagac ctacacctgt 1020

aacgtggacc acaagcccag caacaccaag gtggacaagc gggtggaatc taagtacggc 1080

cctccctgcc ccccctgccc agcccctgaa tttctgggcg gaccctccgt gttcctgttc 1140

cccccaaagc ccaaggacac cctgatgatc agccggaccc ccgaagtgac ctgcgtggtg 1200

gtggacgtgt cccaggaaga tcccgaggtc cagttcaact ggtacgtgga cggcgtggaa 1260

gtgcacaacg ccaagaccaa gcccagagag gaacagttca acagcaccta ccgggtggtg 1320

tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagagtacaa gtgcaaggtc 1380

tccaacaagg gcctgcccag ctccatcgag aaaaccatca gcaaggccaa gggccagccc 1440

cgcgagcccc aggtgtacac actgcccccc agccaggaag agatgaccaa gaaccaggtg 1500

tccctgacct gcctcgtgaa gggcttctac cccagcgata tcgccgtgga atgggagagc 1560

aacggccagc ccgagaacaa ctacaagacc accccccctg tgctggacag cgacggcagc 1620

ttcttcctgt actcccggct gaccgtcgac aagagccggt ggcaggaagg caacgtcttc 1680

agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg 1740

agcctggga 1749

<210> 150

<211> 583

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2重链（N-末端融合）

<400> 150

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Ile Val Gly Pro Leu Glu Val Gln Leu Val

130 135 140

Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser

145 150 155 160

Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Ser Trp Ile His Trp Val

165 170 175

Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Trp Ile Ser Pro

180 185 190

Tyr Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr

195 200 205

Ile Ser Ala Asp Thr Ser Lys Asn Thr Ala Tyr Leu Gln Met Asn Ser

210 215 220

Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Arg His Trp

225 230 235 240

Pro Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser

245 250 255

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser

260 265 270

Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

275 280 285

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

290 295 300

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

305 310 315 320

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys

325 330 335

Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp

340 345 350

Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala

355 360 365

Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro

370 375 380

Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val

385 390 395 400

Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val

405 410 415

Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln

420 425 430

Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln

435 440 445

Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly

450 455 460

Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro

465 470 475 480

Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr

485 490 495

Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser

500 505 510

Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr

515 520 525

Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr

530 535 540

Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe

545 550 555 560

Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys

565 570 575

Ser Leu Ser Leu Ser Leu Gly

580

<210> 151

<211> 1059

<212> DNA

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2轻链（N-末端融合）

<400> 151

gcacctacct cgagttctac aaagaaaaca cagctacaac tggagcattt actgctggat 60

ttacagatga ttttgaatgg aattaataat tacaagaatc ccaaactcac caggatgctc 120

acatttaagt tttacatgcc caagaaggcc acagaactga aacatcttca gtgtctagaa 180

gaagaactca aacctctgga ggaagtgcta aatttagctc aaagcaaaaa ctttcactta 240

agacccaggg acttaatcag caatatcaac gtaatagttc tggaactaaa gggatctgaa 300

acaacattca tgtgtgaata tgctgatgag acagcaacca ttgtagaatt tctgaacaga 360

tggattacct tttgtcaaag catcatctca acactgactg gcattgtggg accgctggac 420

atccagatga cccaatcccc atcctccttg agcgcatctg taggagacag agtcaccatc 480

acctgcaggg cttcacagga tgttagcaca gctgtggctt ggtaccagca gaaacctgga 540

aaggccccaa aactgctgat atacagcgcc tcattcctgt acagcggagt gccctctcgc 600

tttagtggct ctgggagtgg cactgatttc acactgacta tttctagtct gcagcccgaa 660

gatttcgcca cttactattg ccagcagtat ctctatcacc ccgccacctt tgggcagggc 720

acaaaggtcg agattaagcg tacggtggcc gctcccagcg tgttcatctt cccacccagc 780

gacgagcagc tgaagtctgg caccgccagc gtcgtgtgcc tgctgaacaa cttctacccc 840

cgcgaggcca aggtgcagtg gaaggtggac aacgccctgc agtccggcaa cagccaggaa 900

agcgtcaccg agcaggacag caaggactcc acctacagcc tgagcagcac cctgaccctg 960

agcaaggccg actacgagaa gcacaaggtg tacgcctgcg aagtgaccca ccagggcctg 1020

tccagccccg tgaccaagag cttcaaccgg ggcgagtgc 1059

<210> 152

<211> 353

<212> PRT

<213> 人工序列

<220>

<223> PD-L1-GIVGPL-IL2轻链（N-末端融合）

<400> 152

Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu His

1 5 10 15

Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys

20 25 30

Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys

35 40 45

Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys

50 55 60

Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu

65 70 75 80

Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu

85 90 95

Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala

100 105 110

Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile

115 120 125

Ile Ser Thr Leu Thr Gly Ile Val Gly Pro Leu Asp Ile Gln Met Thr

130 135 140

Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile

145 150 155 160

Thr Cys Arg Ala Ser Gln Asp Val Ser Thr Ala Val Ala Trp Tyr Gln

165 170 175

Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Ser Ala Ser Phe

180 185 190

Leu Tyr Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr

195 200 205

Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr

210 215 220

Tyr Tyr Cys Gln Gln Tyr Leu Tyr His Pro Ala Thr Phe Gly Gln Gly

225 230 235 240

Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile

245 250 255

Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser Val Val

260 265 270

Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys

275 280 285

Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val Thr Glu

290 295 300

Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu

305 310 315 320

Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr

325 330 335

His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu

340 345 350

Cys

<210> 153

<211> 1863

<212> DNA

<213> 人工序列

<220>

<223> 9G4-PVGLIG-NanoLuc重链（C-末端融合）

<400> 153

gaagtgatgt tggtggagtc tgggggaggc ttagtgaagc ctggagggtc cctgaaactc 60

tcctgtacag cctctggatt cactttcagt acctatgaca tgtcttgggt tcgccagact 120

ccggagaaga ggctggagtg ggtctcaacc attagtagta gtcgtactta cacctactat 180

tcagccagtg tgaaggggcg attcaccatc tccagagaca atgccaggaa catcctgtac 240

ctgcaaatga gcagtctgag gtctgaggac acggccatgt attactgtgc aagacaattt 300

tctcactggg gccaagggac tctggtcact gtctctgcag cgtccacaaa gggcccaagc 360

gtgttcccgc tagcccccag cagcaagagc accagcggcg gcacagccgc cctgggctgc 420

ctggtgaagg actacttccc cgagcccgtg accgtgtcct ggaacagcgg agccctgacc 480

tccggcgtgc acaccttccc cgccgtgctg cagagcagcg gcctgtacag cctgagcagc 540

gtggtgaccg tgcccagcag cagcctgggc acccagacct acatctgtaa cgtgaaccac 600

aagcccagca acaccaaggt ggacaagaga gtggagccca agagctgtga caagacccac 660

acctgtcccc cctgcccagc ccccgagctg ctgggcggac ccagcgtgtt cctgttcccc 720

cccaagccca aggacaccct gatgatcagc agaacccccg aggtgacctg tgtggtggtg 780

gacgtgtccc acgaggaccc agaggtgaag ttcaactggt acgtggacgg cgtggaggtg 840

cacaacgcca agaccaagcc cagagaggag cagtacaaca gcacctacag ggtggtgtcc 900

gtgctgaccg tgctgcacca ggactggctg aacggcaagg agtacaagtg taaggtgtcc 960

aacaaggccc tgccagcccc aatcgaaaag accatcagca aggccaaggg ccagccaaga 1020

gagccccagg tgtacaccct gccacccagc agggaggaga tgaccaagaa ccaggtgtcc 1080

ctgacctgtc tggtgaaggg cttctaccca agcgacatcg ccgtggagtg ggagagcaac 1140

ggccagcccg agaacaacta caagaccacc cccccagtgc tggacagcga cggcagcttc 1200

ttcctgtaca gcaagctgac cgtggacaag agcagatggc agcagggcaa cgtgttcagc 1260

tgctccgtga tgcacgaggc cctgcacaac cactacaccc agaagagcct gagcctgtcc 1320

ccaggcccgg tgggcctgat tggcctcgag atggtcttca cactcgaaga tttcgttggg 1380

gactggcgac agacagccgg ctacaacctg gaccaagtcc ttgaacaggg aggtgtgtcc 1440

agtttgtttc agaatctcgg ggtgtccgta actccgatcc aaaggattgt cctgagcggt 1500

gaaaatgggc tgaagatcga catccatgtc atcatcccgt atgaaggtct gagcggcgac 1560

caaatgggcc agatcgaaaa aatttttaag gtggtgtacc ctgtggatga tcatcacttt 1620

aaggtgatcc tgcactatgg cacactggta atcgacgggg ttacgccgaa catgatcgac 1680

tatttcggac ggccgtatga aggcatcgcc gtgttcgacg gcaaaaagat cactgtaaca 1740

gggaccctgt ggaacggcaa caaaattatc gacgagcgcc tgatcaaccc cgacggctcc 1800

ctgctgttcc gagtaaccat caacggagtg accggctggc ggctgtgcga acgcattctg 1860

gcg 1863

<210> 154

<211> 621

<212> PRT

<213> 人工序列

<220>

<223> 9G4-PVGLIG-NanoLuc重链（C-末端融合）

<400> 154

Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly

1 5 10 15

Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Thr Tyr

20 25 30

Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val

35 40 45

Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser Ala Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr

65 70 75 80

Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val Thr Val Ser

100 105 110

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser

115 120 125

Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

130 135 140

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

145 150 155 160

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

165 170 175

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln

180 185 190

Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp

195 200 205

Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro

210 215 220

Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro

225 230 235 240

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr

245 250 255

Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn

260 265 270

Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg

275 280 285

Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val

290 295 300

Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser

305 310 315 320

Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys

325 330 335

Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu

340 345 350

Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe

355 360 365

Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu

370 375 380

Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe

385 390 395 400

Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly

405 410 415

Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr

420 425 430

Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val Gly Leu Ile Gly

435 440 445

Leu Glu Met Val Phe Thr Leu Glu Asp Phe Val Gly Asp Trp Arg Gln

450 455 460

Thr Ala Gly Tyr Asn Leu Asp Gln Val Leu Glu Gln Gly Gly Val Ser

465 470 475 480

Ser Leu Phe Gln Asn Leu Gly Val Ser Val Thr Pro Ile Gln Arg Ile

485 490 495

Val Leu Ser Gly Glu Asn Gly Leu Lys Ile Asp Ile His Val Ile Ile

500 505 510

Pro Tyr Glu Gly Leu Ser Gly Asp Gln Met Gly Gln Ile Glu Lys Ile

515 520 525

Phe Lys Val Val Tyr Pro Val Asp Asp His His Phe Lys Val Ile Leu

530 535 540

His Tyr Gly Thr Leu Val Ile Asp Gly Val Thr Pro Asn Met Ile Asp

545 550 555 560

Tyr Phe Gly Arg Pro Tyr Glu Gly Ile Ala Val Phe Asp Gly Lys Lys

565 570 575

Ile Thr Val Thr Gly Thr Leu Trp Asn Gly Asn Lys Ile Ile Asp Glu

580 585 590

Arg Leu Ile Asn Pro Asp Gly Ser Leu Leu Phe Arg Val Thr Ile Asn

595 600 605

Gly Val Thr Gly Trp Arg Leu Cys Glu Arg Ile Leu Ala

610 615 620

<210> 155

<211> 1551

<212> DNA

<213> 人工序列

<220>

<223> 9G4-PVGLIG-CCL4重链（C-末端融合）

<400> 155

gaagtgatgt tggtggagtc tgggggaggc ttagtgaagc ctggagggtc cctgaaactc 60

tcctgtacag cctctggatt cactttcagt acctatgaca tgtcttgggt tcgccagact 120

ccggagaaga ggctggagtg ggtctcaacc attagtagta gtcgtactta cacctactat 180

tcagccagtg tgaaggggcg attcaccatc tccagagaca atgccaggaa catcctgtac 240

ctgcaaatga gcagtctgag gtctgaggac acggccatgt attactgtgc aagacaattt 300

tctcactggg gccaagggac tctggtcact gtctctgcag cgtccacaaa gggcccaagc 360

gtgttcccgc tagcccccag cagcaagagc accagcggcg gcacagccgc cctgggctgc 420

ctggtgaagg actacttccc cgagcccgtg accgtgtcct ggaacagcgg agccctgacc 480

tccggcgtgc acaccttccc cgccgtgctg cagagcagcg gcctgtacag cctgagcagc 540

gtggtgaccg tgcccagcag cagcctgggc acccagacct acatctgtaa cgtgaaccac 600

aagcccagca acaccaaggt ggacaagaga gtggagccca agagctgtga caagacccac 660

acctgtcccc cctgcccagc ccccgagctg ctgggcggac ccagcgtgtt cctgttcccc 720

cccaagccca aggacaccct gatgatcagc agaacccccg aggtgacctg tgtggtggtg 780

gacgtgtccc acgaggaccc agaggtgaag ttcaactggt acgtggacgg cgtggaggtg 840

cacaacgcca agaccaagcc cagagaggag cagtacaaca gcacctacag ggtggtgtcc 900

gtgctgaccg tgctgcacca ggactggctg aacggcaagg agtacaagtg taaggtgtcc 960

aacaaggccc tgccagcccc aatcgaaaag accatcagca aggccaaggg ccagccaaga 1020

gagccccagg tgtacaccct gccacccagc agggaggaga tgaccaagaa ccaggtgtcc 1080

ctgacctgtc tggtgaaggg cttctaccca agcgacatcg ccgtggagtg ggagagcaac 1140

ggccagcccg agaacaacta caagaccacc cccccagtgc tggacagcga cggcagcttc 1200

ttcctgtaca gcaagctgac cgtggacaag agcagatggc agcagggcaa cgtgttcagc 1260

tgctccgtga tgcacgaggc cctgcacaac cactacaccc agaagagcct gagcctgtcc 1320

ccaggcccgg tgggcctgat tggcgcacca atgggctcag accctcccac cgcctgctgc 1380

ttttcttaca ccgcgaggaa acttcctcgc aactttgtgg tagattacta tgagaccagc 1440

agcctctgct cccagccagc tgtggtattc caaaccaaaa gaagcaagca agtctgtgct 1500

gatcccagtg aatcctgggt ccaggagtac gtgtatgacc tggaactgaa c 1551

<210> 156

<211> 517

<212> PRT

<213> 人工序列

<220>

<223> 9G4-PVGLIG-CCL4重链（C-末端融合）

<400> 156

Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly

1 5 10 15

Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Thr Tyr

20 25 30

Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val

35 40 45

Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser Ala Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr

65 70 75 80

Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val Thr Val Ser

100 105 110

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser

115 120 125

Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

130 135 140

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

145 150 155 160

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

165 170 175

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln

180 185 190

Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp

195 200 205

Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro

210 215 220

Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro

225 230 235 240

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr

245 250 255

Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn

260 265 270

Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg

275 280 285

Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val

290 295 300

Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser

305 310 315 320

Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys

325 330 335

Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu

340 345 350

Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe

355 360 365

Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu

370 375 380

Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe

385 390 395 400

Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly

405 410 415

Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr

420 425 430

Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val Gly Leu Ile Gly

435 440 445

Ala Pro Met Gly Ser Asp Pro Pro Thr Ala Cys Cys Phe Ser Tyr Thr

450 455 460

Ala Arg Lys Leu Pro Arg Asn Phe Val Val Asp Tyr Tyr Glu Thr Ser

465 470 475 480

Ser Leu Cys Ser Gln Pro Ala Val Val Phe Gln Thr Lys Arg Ser Lys

485 490 495

Gln Val Cys Ala Asp Pro Ser Glu Ser Trp Val Gln Glu Tyr Val Tyr

500 505 510

Asp Leu Glu Leu Asn

515

<210> 157

<211> 1686

<212> DNA

<213> 人工序列

<220>

<223> 9G4-PVGLIG-IL15重链（C-末端融合）

<400> 157

gaagtgatgt tggtggagtc tgggggaggc ttagtgaagc ctggagggtc cctgaaactc 60

tcctgtacag cctctggatt cactttcagt acctatgaca tgtcttgggt tcgccagact 120

ccggagaaga ggctggagtg ggtctcaacc attagtagta gtcgtactta cacctactat 180

tcagccagtg tgaaggggcg attcaccatc tccagagaca atgccaggaa catcctgtac 240

ctgcaaatga gcagtctgag gtctgaggac acggccatgt attactgtgc aagacaattt 300

tctcactggg gccaagggac tctggtcact gtctctgcag cgtccacaaa gggcccaagc 360

gtgttcccgc tagcccccag cagcaagagc accagcggcg gcacagccgc cctgggctgc 420

ctggtgaagg actacttccc cgagcccgtg accgtgtcct ggaacagcgg agccctgacc 480

tccggcgtgc acaccttccc cgccgtgctg cagagcagcg gcctgtacag cctgagcagc 540

gtggtgaccg tgcccagcag cagcctgggc acccagacct acatctgtaa cgtgaaccac 600

aagcccagca acaccaaggt ggacaagaga gtggagccca agagctgtga caagacccac 660

acctgtcccc cctgcccagc ccccgagctg ctgggcggac ccagcgtgtt cctgttcccc 720

cccaagccca aggacaccct gatgatcagc agaacccccg aggtgacctg tgtggtggtg 780

gacgtgtccc acgaggaccc agaggtgaag ttcaactggt acgtggacgg cgtggaggtg 840

cacaacgcca agaccaagcc cagagaggag cagtacaaca gcacctacag ggtggtgtcc 900

gtgctgaccg tgctgcacca ggactggctg aacggcaagg agtacaagtg taaggtgtcc 960

aacaaggccc tgccagcccc aatcgaaaag accatcagca aggccaaggg ccagccaaga 1020

gagccccagg tgtacaccct gccacccagc agggaggaga tgaccaagaa ccaggtgtcc 1080

ctgacctgtc tggtgaaggg cttctaccca agcgacatcg ccgtggagtg ggagagcaac 1140

ggccagcccg agaacaacta caagaccacc cccccagtgc tggacagcga cggcagcttc 1200

ttcctgtaca gcaagctgac cgtggacaag agcagatggc agcagggcaa cgtgttcagc 1260

tgctccgtga tgcacgaggc cctgcacaac cactacaccc agaagagcct gagcctgtcc 1320

ccaggcccgg tgggcctgat tggcaactgg gtgaatgtaa taagtgattt gaaaaaaatt 1380

gaagatctta ttcaatctat gcatattgat gctactttat atacggaaag tgatgttcac 1440

cccagttgca aagtaacagc aatgaagtgc tttctcttgg agttacaagt tatttcactt 1500

gagtccggag atgcaagtat tcatgataca gtagaaaatc tgatcatcct agcaaacaac 1560

agtttgtctt ctaatgggaa tgtaacagaa tctggatgca aagaatgtga ggaactggag 1620

gaaaaaaata ttaaagaatt tttgcagagt tttgtacata ttgtccaaat gttcatcaac 1680

acttct 1686

<210> 158

<211> 562

<212> PRT

<213> 人工序列

<220>

<223> 9G4-PVGLIG-IL15重链（C-末端融合）

<400> 158

Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly

1 5 10 15

Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Thr Tyr

20 25 30

Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val

35 40 45

Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser Ala Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr

65 70 75 80

Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val Thr Val Ser

100 105 110

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser

115 120 125

Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

130 135 140

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

145 150 155 160

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

165 170 175

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln

180 185 190

Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp

195 200 205

Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro

210 215 220

Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro

225 230 235 240

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr

245 250 255

Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn

260 265 270

Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg

275 280 285

Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val

290 295 300

Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser

305 310 315 320

Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys

325 330 335

Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu

340 345 350

Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe

355 360 365

Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu

370 375 380

Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe

385 390 395 400

Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly

405 410 415

Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr

420 425 430

Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val Gly Leu Ile Gly

435 440 445

Asn Trp Val Asn Val Ile Ser Asp Leu Lys Lys Ile Glu Asp Leu Ile

450 455 460

Gln Ser Met His Ile Asp Ala Thr Leu Tyr Thr Glu Ser Asp Val His

465 470 475 480

Pro Ser Cys Lys Val Thr Ala Met Lys Cys Phe Leu Leu Glu Leu Gln

485 490 495

Val Ile Ser Leu Glu Ser Gly Asp Ala Ser Ile His Asp Thr Val Glu

500 505 510

Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu Ser Ser Asn Gly Asn Val

515 520 525

Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu Leu Glu Glu Lys Asn Ile

530 535 540

Lys Glu Phe Leu Gln Ser Phe Val His Ile Val Gln Met Phe Ile Asn

545 550 555 560

Thr Ser

<210> 159

<211> 1839

<212> DNA

<213> 人工序列

<220>

<223> 9G4-PVGLIG-IFNalpha重链（C-末端融合）

<400> 159

gaagtgatgt tggtggagtc tgggggaggc ttagtgaagc ctggagggtc cctgaaactc 60

tcctgtacag cctctggatt cactttcagt acctatgaca tgtcttgggt tcgccagact 120

ccggagaaga ggctggagtg ggtctcaacc attagtagta gtcgtactta cacctactat 180

tcagccagtg tgaaggggcg attcaccatc tccagagaca atgccaggaa catcctgtac 240

ctgcaaatga gcagtctgag gtctgaggac acggccatgt attactgtgc aagacaattt 300

tctcactggg gccaagggac tctggtcact gtctctgcag cgtccacaaa gggcccaagc 360

gtgttcccgc tagcccccag cagcaagagc accagcggcg gcacagccgc cctgggctgc 420

ctggtgaagg actacttccc cgagcccgtg accgtgtcct ggaacagcgg agccctgacc 480

tccggcgtgc acaccttccc cgccgtgctg cagagcagcg gcctgtacag cctgagcagc 540

gtggtgaccg tgcccagcag cagcctgggc acccagacct acatctgtaa cgtgaaccac 600

aagcccagca acaccaaggt ggacaagaga gtggagccca agagctgtga caagacccac 660

acctgtcccc cctgcccagc ccccgagctg ctgggcggac ccagcgtgtt cctgttcccc 720

cccaagccca aggacaccct gatgatcagc agaacccccg aggtgacctg tgtggtggtg 780

gacgtgtccc acgaggaccc agaggtgaag ttcaactggt acgtggacgg cgtggaggtg 840

cacaacgcca agaccaagcc cagagaggag cagtacaaca gcacctacag ggtggtgtcc 900

gtgctgaccg tgctgcacca ggactggctg aacggcaagg agtacaagtg taaggtgtcc 960

aacaaggccc tgccagcccc aatcgaaaag accatcagca aggccaaggg ccagccaaga 1020

gagccccagg tgtacaccct gccacccagc agggaggaga tgaccaagaa ccaggtgtcc 1080

ctgacctgtc tggtgaaggg cttctaccca agcgacatcg ccgtggagtg ggagagcaac 1140

ggccagcccg agaacaacta caagaccacc cccccagtgc tggacagcga cggcagcttc 1200

ttcctgtaca gcaagctgac cgtggacaag agcagatggc agcagggcaa cgtgttcagc 1260

tgctccgtga tgcacgaggc cctgcacaac cactacaccc agaagagcct gagcctgtcc 1320

ccaggcccgg tgggcctgat tggctgtgat ctgcctcaaa cccacagcct gggtagcagg 1380

aggaccttga tgctcctggc acagatgagg aaaatctctc ttttctcctg cttgaaggac 1440

agacatgact ttggatttcc ccaggaggag tttggcaacc agttccaaaa ggctgaaacc 1500

atccctgtcc tccatgagat gatccagcag atcttcaatc tcttcagcac aaaggactca 1560

tctgctgctt gggatgagac cctcctagac aaattctaca ctgaactcta ccagcagctg 1620

aatgacctgg aagcctgtgt gatacagggg gtgggggtga cagagactcc cctgatgaag 1680

gaggactcca ttctggctgt gaggaaatac ttccaaagaa tcactctcta tctgaaagag 1740

aagaaataca gcccttgtgc ctgggaggtt gtcagagcag aaatcatgag atctttttct 1800

ttgtcaacaa acttgcaaga aagtttaaga agtaaggaa 1839

<210> 160

<211> 613

<212> PRT

<213> 人工序列

<220>

<223> 9G4-PVGLIG-IFNalpha重链（C-末端融合）

<400> 160

Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly

1 5 10 15

Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Thr Tyr

20 25 30

Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val

35 40 45

Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser Ala Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr

65 70 75 80

Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val Thr Val Ser

100 105 110

Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser

115 120 125

Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp

130 135 140

Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr

145 150 155 160

Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr

165 170 175

Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln

180 185 190

Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp

195 200 205

Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro

210 215 220

Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro

225 230 235 240

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr

245 250 255

Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn

260 265 270

Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg

275 280 285

Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val

290 295 300

Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser

305 310 315 320

Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys

325 330 335

Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu

340 345 350

Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe

355 360 365

Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu

370 375 380

Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe

385 390 395 400

Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly

405 410 415

Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr

420 425 430

Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val Gly Leu Ile Gly

435 440 445

Cys Asp Leu Pro Gln Thr His Ser Leu Gly Ser Arg Arg Thr Leu Met

450 455 460

Leu Leu Ala Gln Met Arg Lys Ile Ser Leu Phe Ser Cys Leu Lys Asp

465 470 475 480

Arg His Asp Phe Gly Phe Pro Gln Glu Glu Phe Gly Asn Gln Phe Gln

485 490 495

Lys Ala Glu Thr Ile Pro Val Leu His Glu Met Ile Gln Gln Ile Phe

500 505 510

Asn Leu Phe Ser Thr Lys Asp Ser Ser Ala Ala Trp Asp Glu Thr Leu

515 520 525

Leu Asp Lys Phe Tyr Thr Glu Leu Tyr Gln Gln Leu Asn Asp Leu Glu

530 535 540

Ala Cys Val Ile Gln Gly Val Gly Val Thr Glu Thr Pro Leu Met Lys

545 550 555 560

Glu Asp Ser Ile Leu Ala Val Arg Lys Tyr Phe Gln Arg Ile Thr Leu

565 570 575

Tyr Leu Lys Glu Lys Lys Tyr Ser Pro Cys Ala Trp Glu Val Val Arg

580 585 590

Ala Glu Ile Met Arg Ser Phe Ser Leu Ser Thr Asn Leu Gln Glu Ser

595 600 605

Leu Arg Ser Lys Glu

610

<210> 161

<211> 1698

<212> DNA

<213> 人工序列

<220>

<223> NHS76-PVGLIG-IL15重链（C-末端融合）

<400> 161

caggtgcagc tgcaggagtc cggcccagga ctggtgaagc cttcggagac cctgtccctc 60

acctgcgctg tctctggtta ctccatcagc agtggttact actggggctg gattcggcag 120

cccccaggga aggggctgga gtggattggg agtatctatc atagtgggag cacctactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca cgtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcagac acggccgtgt attactgtgc aagagggaag 300

tggtcgaagt ttgactattg gggccaaggc accctggtca ccgtctcttc agcgtccaca 360

aagggcccaa gcgtgttccc gctagccccc agcagcaaga gcaccagcgg cggcacagcc 420

gccctgggct gcctggtgaa ggactacttc cccgagcccg tgaccgtgtc ctggaacagc 480

ggagccctga cctccggcgt gcacaccttc cccgccgtgc tgcagagcag cggcctgtac 540

agcctgagca gcgtggtgac cgtgcccagc agcagcctgg gcacccagac ctacatctgt 600

aacgtgaacc acaagcccag caacaccaag gtggacaaga gagtggagcc caagagctgt 660

gacaagaccc acacctgtcc cccctgccca gcccccgagc tgctgggcgg acccagcgtg 720

ttcctgttcc cccccaagcc caaggacacc ctgatgatca gcagaacccc cgaggtgacc 780

tgtgtggtgg tggacgtgtc ccacgaggac ccagaggtga agttcaactg gtacgtggac 840

ggcgtggagg tgcacaacgc caagaccaag cccagagagg agcagtacaa cagcacctac 900

agggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa ggagtacaag 960

tgtaaggtgt ccaacaaggc cctgccagcc ccaatcgaaa agaccatcag caaggccaag 1020

ggccagccaa gagagcccca ggtgtacacc ctgccaccca gcagggagga gatgaccaag 1080

aaccaggtgt ccctgacctg tctggtgaag ggcttctacc caagcgacat cgccgtggag 1140

tgggagagca acggccagcc cgagaacaac tacaagacca cccccccagt gctggacagc 1200

gacggcagct tcttcctgta cagcaagctg accgtggaca agagcagatg gcagcagggc 1260

aacgtgttca gctgctccgt gatgcacgag gccctgcaca accactacac ccagaagagc 1320

ctgagcctgt ccccaggccc ggtgggcctg attggcaact gggtgaatgt aataagtgat 1380

ttgaaaaaaa ttgaagatct tattcaatct atgcatattg atgctacttt atatacggaa 1440

agtgatgttc accccagttg caaagtaaca gcaatgaagt gctttctctt ggagttacaa 1500

gttatttcac ttgagtccgg agatgcaagt attcatgata cagtagaaaa tctgatcatc 1560

ctagcaaaca acagtttgtc ttctaatggg aatgtaacag aatctggatg caaagaatgt 1620

gaggaactgg aggaaaaaaa tattaaagaa tttttgcaga gttttgtaca tattgtccaa 1680

atgttcatca acacttct 1698

<210> 162

<211> 566

<212> PRT

<213> 人工序列

<220>

<223> NHS76-PVGLIG-IL15重链（C-末端融合）

<400> 162

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly

20 25 30

Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu

100 105 110

Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu

115 120 125

Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys

130 135 140

Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser

145 150 155 160

Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser

165 170 175

Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser

180 185 190

Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn

195 200 205

Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His

210 215 220

Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val

225 230 235 240

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr

245 250 255

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu

260 265 270

Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys

275 280 285

Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser

290 295 300

Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys

305 310 315 320

Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile

325 330 335

Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro

340 345 350

Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu

355 360 365

Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn

370 375 380

Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser

385 390 395 400

Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg

405 410 415

Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu

420 425 430

His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Pro Val

435 440 445

Gly Leu Ile Gly Asn Trp Val Asn Val Ile Ser Asp Leu Lys Lys Ile

450 455 460

Glu Asp Leu Ile Gln Ser Met His Ile Asp Ala Thr Leu Tyr Thr Glu

465 470 475 480

Ser Asp Val His Pro Ser Cys Lys Val Thr Ala Met Lys Cys Phe Leu

485 490 495

Leu Glu Leu Gln Val Ile Ser Leu Glu Ser Gly Asp Ala Ser Ile His

500 505 510

Asp Thr Val Glu Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu Ser Ser

515 520 525

Asn Gly Asn Val Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu Leu Glu

530 535 540

Glu Lys Asn Ile Lys Glu Phe Leu Gln Ser Phe Val His Ile Val Gln

545 550 555 560

Met Phe Ile Asn Thr Ser

565

<210> 163

<211> 1704

<212> DNA

<213> 人工序列

<220>

<223> H16/L16-PVGLIG-IL15重链（C-末端融合）

<400> 163

caggtgcagc tgcaggagtc gggcccagga ctggtgaagc cttcggggac cctgtccctc 60

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 120

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 180

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 240

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 300

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 360

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 420

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 480

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 540

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 600

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 660

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 720

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 780

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 840

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 900

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 960

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1020

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1080

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1140

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1200

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1260

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1320

aagagcctga gcctgtcccc aggcccggtg ggcctgattg gcaactgggt gaatgtaata 1380

agtgatttga aaaaaattga agatcttatt caatctatgc atattgatgc tactttatat 1440

acggaaagtg atgttcaccc cagttgcaaa gtaacagcaa tgaagtgctt tctcttggag 1500

ttacaagtta tttcacttga gtccggagat gcaagtattc atgatacagt agaaaatctg 1560

atcatcctag caaacaacag tttgtcttct aatgggaatg taacagaatc tggatgcaaa 1620

gaatgtgagg aactggagga aaaaaatatt aaagaatttt tgcagagttt tgtacatatt 1680

gtccaaatgt tcatcaacac ttct 1704

<210> 164

<211> 568

<212> PRT

<213> 人工序列

<220>

<223> H16/L16-PVGLIG-IL15重链（C-末端融合）

<400> 164

Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Gly

1 5 10 15

Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser Ile Ser Ser Ser

20 25 30

Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp

35 40 45

Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu

50 55 60

Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys Asn Gln Phe Ser

65 70 75 80

Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile Trp Gly Gln Gly

100 105 110

Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe

115 120 125

Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu

130 135 140

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp

145 150 155 160

Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu

165 170 175

Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser

180 185 190

Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro

195 200 205

Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys

210 215 220

Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro

225 230 235 240

Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser

245 250 255

Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp

260 265 270

Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn

275 280 285

Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val

290 295 300

Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu

305 310 315 320

Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys

325 330 335

Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr

340 345 350

Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr

355 360 365

Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu

370 375 380

Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu

385 390 395 400

Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys

405 410 415

Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu

420 425 430

Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

435 440 445

Pro Val Gly Leu Ile Gly Asn Trp Val Asn Val Ile Ser Asp Leu Lys

450 455 460

Lys Ile Glu Asp Leu Ile Gln Ser Met His Ile Asp Ala Thr Leu Tyr

465 470 475 480

Thr Glu Ser Asp Val His Pro Ser Cys Lys Val Thr Ala Met Lys Cys

485 490 495

Phe Leu Leu Glu Leu Gln Val Ile Ser Leu Glu Ser Gly Asp Ala Ser

500 505 510

Ile His Asp Thr Val Glu Asn Leu Ile Ile Leu Ala Asn Asn Ser Leu

515 520 525

Ser Ser Asn Gly Asn Val Thr Glu Ser Gly Cys Lys Glu Cys Glu Glu

530 535 540

Leu Glu Glu Lys Asn Ile Lys Glu Phe Leu Gln Ser Phe Val His Ile

545 550 555 560

Val Gln Met Phe Ile Asn Thr Ser

565

<210> 165

<211> 12

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 165

tcaggcagat ct 12

<210> 166

<211> 4

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 166

Ser Gly Arg Ser

1

<210> 167

<211> 15

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 167

tcaggcagat ctgcc 15

<210> 168

<211> 5

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 168

Ser Gly Arg Ser Ala

1 5

<210> 169

<211> 24

<212> DNA

<213> 人工序列

<220>

<223> 连接子

<400> 169

ccgagcagcc ggagaagagt gaac 24

<210> 170

<211> 8

<212> PRT

<213> 人工序列

<220>

<223> 连接子

<400> 170

Pro Ser Ser Arg Arg Arg Val Asn

1 5

<210> 171

<211> 447

<212> DNA

<213> 人

<400> 171

aaacccatca ccggcaccat caacgacctg aaccagcaag tgtggacact gcagggccag 60

aatctggtgg ccgtgcctag aagcgatagc gtgacacctg tgaccgtggc cgtgatcaca 120

tgcaagtacc ccgaggctct tgagcaaggc agaggcgatc ctatctacct gggcattcag 180

aaccccgaga tgtgcctgta ctgcgagaaa gtgggagagc agcccacact gcagctgaaa 240

gaacagaaaa tcatggacct gtacggccag cctgagcctg tgaagccctt cctgttttac 300

agagccaaga ccggccggac cagcacactg gaatctgtgg cctttcctga ctggtttatc 360

gccagcagca agcgggacca gcctatcatc ctgacaagcg agctgggcaa gagctacaac 420

accgccttcg agctgaacat caacgat 447

<210> 172

<211> 149

<212> PRT

<213> 人

<400> 172

Lys Pro Ile Thr Gly Thr Ile Asn Asp Leu Asn Gln Gln Val Trp Thr

1 5 10 15

Leu Gln Gly Gln Asn Leu Val Ala Val Pro Arg Ser Asp Ser Val Thr

20 25 30

Pro Val Thr Val Ala Val Ile Thr Cys Lys Tyr Pro Glu Ala Leu Glu

35 40 45

Gln Gly Arg Gly Asp Pro Ile Tyr Leu Gly Ile Gln Asn Pro Glu Met

50 55 60

Cys Leu Tyr Cys Glu Lys Val Gly Glu Gln Pro Thr Leu Gln Leu Lys

65 70 75 80

Glu Gln Lys Ile Met Asp Leu Tyr Gly Gln Pro Glu Pro Val Lys Pro

85 90 95

Phe Leu Phe Tyr Arg Ala Lys Thr Gly Arg Thr Ser Thr Leu Glu Ser

100 105 110

Val Ala Phe Pro Asp Trp Phe Ile Ala Ser Ser Lys Arg Asp Gln Pro

115 120 125

Ile Ile Leu Thr Ser Glu Leu Gly Lys Ser Tyr Asn Thr Ala Phe Glu

130 135 140

Leu Asn Ile Asn Asp

145

<210> 173

<211> 1869

<212> DNA

<213> 人工序列

<220>

<223> H16/L16-PVGLIG-IL36G重链

<400> 173

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 120

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 180

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 360

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 420

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 480

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 540

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 600

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 660

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 720

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 780

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 840

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 900

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 960

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 1020

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1080

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1140

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1200

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1260

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1320

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1380

aagagcctga gcctgtcccc aggcccggtg ggcctgattg gcaaacccat caccggcacc 1440

atcaacgacc tgaaccagca agtgtggaca ctgcagggcc agaatctggt ggccgtgcct 1500

agaagcgata gcgtgacacc tgtgaccgtg gccgtgatca catgcaagta ccccgaggct 1560

cttgagcaag gcagaggcga tcctatctac ctgggcattc agaaccccga gatgtgcctg 1620

tactgcgaga aagtgggaga gcagcccaca ctgcagctga aagaacagaa aatcatggac 1680

ctgtacggcc agcctgagcc tgtgaagccc ttcctgtttt acagagccaa gaccggccgg 1740

accagcacac tggaatctgt ggcctttcct gactggttta tcgccagcag caagcgggac 1800

cagcctatca tcctgacaag cgagctgggc aagagctaca acaccgcctt cgagctgaac 1860

atcaacgat 1869

<210> 174

<211> 623

<212> PRT

<213> 人工序列

<220>

<223> H16/L16-PVGLIG-IL36G重链

<400> 174

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Gly Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser

35 40 45

Ile Ser Ser Ser Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile

115 120 125

Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly

130 135 140

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

145 150 155 160

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

165 170 175

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

180 185 190

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val

195 200 205

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

210 215 220

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys

225 230 235 240

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

245 250 255

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

275 280 285

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

290 295 300

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

305 310 315 320

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

355 360 365

Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln

370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

405 410 415

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

420 425 430

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

435 440 445

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

450 455 460

Leu Ser Pro Gly Pro Val Gly Leu Ile Gly Lys Pro Ile Thr Gly Thr

465 470 475 480

Ile Asn Asp Leu Asn Gln Gln Val Trp Thr Leu Gln Gly Gln Asn Leu

485 490 495

Val Ala Val Pro Arg Ser Asp Ser Val Thr Pro Val Thr Val Ala Val

500 505 510

Ile Thr Cys Lys Tyr Pro Glu Ala Leu Glu Gln Gly Arg Gly Asp Pro

515 520 525

Ile Tyr Leu Gly Ile Gln Asn Pro Glu Met Cys Leu Tyr Cys Glu Lys

530 535 540

Val Gly Glu Gln Pro Thr Leu Gln Leu Lys Glu Gln Lys Ile Met Asp

545 550 555 560

Leu Tyr Gly Gln Pro Glu Pro Val Lys Pro Phe Leu Phe Tyr Arg Ala

565 570 575

Lys Thr Gly Arg Thr Ser Thr Leu Glu Ser Val Ala Phe Pro Asp Trp

580 585 590

Phe Ile Ala Ser Ser Lys Arg Asp Gln Pro Ile Ile Leu Thr Ser Glu

595 600 605

Leu Gly Lys Ser Tyr Asn Thr Ala Phe Glu Leu Asn Ile Asn Asp

610 615 620

<210> 175

<211> 1626

<212> DNA

<213> 人工序列

<220>

<223> 9G4-SGRS-hCXCL10重链

<400> 175

atgaacttcg gactctccct gatcttcctt gttctggtgc ttaagggcgt gcagtgcgaa 60

gtgatgttgg tagaaagcgg tggtgggctg gtgaaacctg gcgggtccct gaaactctcc 120

tgtactgcta gtggattcac attcagcact tacgacatgt catgggtccg gcagacccca 180

gagaagagac tggagtgggt cagcactatc tcctcttctc ggacctatac gtattacagt 240

gccagcgtca agggacgctt taccattagc agggataatg ccagaaacat actgtacctg 300

cagatgtcaa gcctccgaag tgaggacacc gccatgtatt actgcgctag gcaattttca 360

cactggggcc aggggacatt ggttaccgtg tctgcagcgt ccacaaaggg cccaagcgtg 420

ttcccgctag cccccagcag caagagcacc agcggcggca cagccgccct gggctgcctg 480

gtgaaggact acttccccga gcccgtgacc gtgtcctgga acagcggagc cctgacctcc 540

ggcgtgcaca ccttccccgc cgtgctgcag agcagcggcc tgtacagcct gagcagcgtg 600

gtgaccgtgc ccagcagcag cctgggcacc cagacctaca tctgtaacgt gaaccacaag 660

cccagcaaca ccaaggtgga caagagagtg gagcccaaga gctgtgacaa gacccacacc 720

tgtcccccct gcccagcccc cgagctgctg ggcggaccca gcgtgttcct gttccccccc 780

aagcccaagg acaccctgat gatcagcaga acccccgagg tgacctgtgt ggtggtggac 840

gtgtcccacg aggacccaga ggtgaagttc aactggtacg tggacggcgt ggaggtgcac 900

aacgccaaga ccaagcccag agaggagcag tacaacagca cctacagggt ggtgtccgtg 960

ctgaccgtgc tgcaccagga ctggctgaac ggcaaggagt acaagtgtaa ggtgtccaac 1020

aaggccctgc cagccccaat cgaaaagacc atcagcaagg ccaagggcca gccaagagag 1080

ccccaggtgt acaccctgcc acccagcagg gaggagatga ccaagaacca ggtgtccctg 1140

acctgtctgg tgaagggctt ctacccaagc gacatcgccg tggagtggga gagcaacggc 1200

cagcccgaga acaactacaa gaccaccccc ccagtgctgg acagcgacgg cagcttcttc 1260

ctgtacagca agctgaccgt ggacaagagc agatggcagc agggcaacgt gttcagctgc 1320

tccgtgatgc acgaggccct gcacaaccac tacacccaga agagcctgag cctgtcccca 1380

ggctcaggca gatctgtacc tctctctaga actgtacgct gtacctgcat cagcattagt 1440

aatcaacctg ttaatccaag gtctttagaa aaacttgaaa ttattcctgc aagccaattt 1500

tgtccacgtg ttgagatcat tgctacaatg aaaaagaagg gtgagaagag atgtctgaat 1560

ccagaatcga aggccatcaa gaatttactg aaagcagtta gcaaggaaag gtctaaaaga 1620

tctcct 1626

<210> 176

<211> 542

<212> PRT

<213> 人工序列

<220>

<223> 9G4-SGRS-hCXCL10重链

<400> 176

Met Asn Phe Gly Leu Ser Leu Ile Phe Leu Val Leu Val Leu Lys Gly

1 5 10 15

Val Gln Cys Glu Val Met Leu Val Glu Ser Gly Gly Gly Leu Val Lys

20 25 30

Pro Gly Gly Ser Leu Lys Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe

35 40 45

Ser Thr Tyr Asp Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu

50 55 60

Glu Trp Val Ser Thr Ile Ser Ser Ser Arg Thr Tyr Thr Tyr Tyr Ser

65 70 75 80

Ala Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn

85 90 95

Ile Leu Tyr Leu Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met

100 105 110

Tyr Tyr Cys Ala Arg Gln Phe Ser His Trp Gly Gln Gly Thr Leu Val

115 120 125

Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala

130 135 140

Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu

145 150 155 160

Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly

165 170 175

Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser

180 185 190

Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu

195 200 205

Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr

210 215 220

Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr

225 230 235 240

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

245 250 255

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

260 265 270

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

275 280 285

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

290 295 300

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

305 310 315 320

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

325 330 335

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

340 345 350

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

355 360 365

Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

370 375 380

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

385 390 395 400

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

405 410 415

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

420 425 430

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

435 440 445

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Ser Gly Arg

450 455 460

Ser Val Pro Leu Ser Arg Thr Val Arg Cys Thr Cys Ile Ser Ile Ser

465 470 475 480

Asn Gln Pro Val Asn Pro Arg Ser Leu Glu Lys Leu Glu Ile Ile Pro

485 490 495

Ala Ser Gln Phe Cys Pro Arg Val Glu Ile Ile Ala Thr Met Lys Lys

500 505 510

Lys Gly Glu Lys Arg Cys Leu Asn Pro Glu Ser Lys Ala Ile Lys Asn

515 520 525

Leu Leu Lys Ala Val Ser Lys Glu Arg Ser Lys Arg Ser Pro

530 535 540

<210> 177

<211> 1641

<212> DNA

<213> 人工序列

<220>

<223> NHS76-SGRS-hCXCL10重链

<400> 177

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc cggcccagga ctggtgaagc cttcggagac cctgtccctc 120

acctgcgctg tctctggtta ctccatcagc agtggttact actggggctg gattcggcag 180

cccccaggga aggggctgga gtggattggg agtatctatc atagtgggag cacctactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca cgtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcagac acggccgtgt attactgtgc aagagggaag 360

tggtcgaagt ttgactattg gggccaaggc accctggtca ccgtctcttc agcgtccaca 420

aagggcccaa gcgtgttccc gctagccccc agcagcaaga gcaccagcgg cggcacagcc 480

gccctgggct gcctggtgaa ggactacttc cccgagcccg tgaccgtgtc ctggaacagc 540

ggagccctga cctccggcgt gcacaccttc cccgccgtgc tgcagagcag cggcctgtac 600

agcctgagca gcgtggtgac cgtgcccagc agcagcctgg gcacccagac ctacatctgt 660

aacgtgaacc acaagcccag caacaccaag gtggacaaga gagtggagcc caagagctgt 720

gacaagaccc acacctgtcc cccctgccca gcccccgagc tgctgggcgg acccagcgtg 780

ttcctgttcc cccccaagcc caaggacacc ctgatgatca gcagaacccc cgaggtgacc 840

tgtgtggtgg tggacgtgtc ccacgaggac ccagaggtga agttcaactg gtacgtggac 900

ggcgtggagg tgcacaacgc caagaccaag cccagagagg agcagtacaa cagcacctac 960

agggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa ggagtacaag 1020

tgtaaggtgt ccaacaaggc cctgccagcc ccaatcgaaa agaccatcag caaggccaag 1080

ggccagccaa gagagcccca ggtgtacacc ctgccaccca gcagggagga gatgaccaag 1140

aaccaggtgt ccctgacctg tctggtgaag ggcttctacc caagcgacat cgccgtggag 1200

tgggagagca acggccagcc cgagaacaac tacaagacca cccccccagt gctggacagc 1260

gacggcagct tcttcctgta cagcaagctg accgtggaca agagcagatg gcagcagggc 1320

aacgtgttca gctgctccgt gatgcacgag gccctgcaca accactacac ccagaagagc 1380

ctgagcctgt ccccaggctc aggcagatct gtacctctct ctagaactgt acgctgtacc 1440

tgcatcagca ttagtaatca acctgttaat ccaaggtctt tagaaaaact tgaaattatt 1500

cctgcaagcc aattttgtcc acgtgttgag atcattgcta caatgaaaaa gaagggtgag 1560

aagagatgtc tgaatccaga atcgaaggcc atcaagaatt tactgaaagc agttagcaag 1620

gaaaggtcta aaagatctcc t 1641

<210> 178

<211> 547

<212> PRT

<213> 人工序列

<220>

<223> NHS76-SGRS-hCXCL10重链

<400> 178

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser

35 40 45

Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly

115 120 125

Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser

130 135 140

Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala

145 150 155 160

Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val

165 170 175

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala

180 185 190

Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val

195 200 205

Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His

210 215 220

Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys

225 230 235 240

Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly

245 250 255

Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met

260 265 270

Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His

275 280 285

Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val

290 295 300

His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr

305 310 315 320

Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly

325 330 335

Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile

340 345 350

Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val

355 360 365

Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser

370 375 380

Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu

385 390 395 400

Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro

405 410 415

Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val

420 425 430

Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met

435 440 445

His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser

450 455 460

Pro Gly Ser Gly Arg Ser Val Pro Leu Ser Arg Thr Val Arg Cys Thr

465 470 475 480

Cys Ile Ser Ile Ser Asn Gln Pro Val Asn Pro Arg Ser Leu Glu Lys

485 490 495

Leu Glu Ile Ile Pro Ala Ser Gln Phe Cys Pro Arg Val Glu Ile Ile

500 505 510

Ala Thr Met Lys Lys Lys Gly Glu Lys Arg Cys Leu Asn Pro Glu Ser

515 520 525

Lys Ala Ile Lys Asn Leu Leu Lys Ala Val Ser Lys Glu Arg Ser Lys

530 535 540

Arg Ser Pro

545

<210> 179

<211> 1647

<212> DNA

<213> 人工序列

<220>

<223> H16L16-SGRS-hCXCL10重链

<400> 179

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 120

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 180

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 360

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 420

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 480

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 540

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 600

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 660

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 720

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 780

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 840

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 900

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 960

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 1020

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1080

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1140

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1200

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1260

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1320

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1380

aagagcctga gcctgtcccc aggctcaggc agatctgtac ctctctctag aactgtacgc 1440

tgtacctgca tcagcattag taatcaacct gttaatccaa ggtctttaga aaaacttgaa 1500

attattcctg caagccaatt ttgtccacgt gttgagatca ttgctacaat gaaaaagaag 1560

ggtgagaaga gatgtctgaa tccagaatcg aaggccatca agaatttact gaaagcagtt 1620

agcaaggaaa ggtctaaaag atctcct 1647

<210> 180

<211> 549

<212> PRT

<213> 人工序列

<220>

<223> H16L16-SGRS-hCXCL10重链

<400> 180

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Gly Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser

35 40 45

Ile Ser Ser Ser Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile

115 120 125

Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly

130 135 140

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

145 150 155 160

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

165 170 175

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

180 185 190

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val

195 200 205

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

210 215 220

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys

225 230 235 240

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

245 250 255

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

275 280 285

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

290 295 300

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

305 310 315 320

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

355 360 365

Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln

370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

405 410 415

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

420 425 430

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

435 440 445

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

450 455 460

Leu Ser Pro Gly Ser Gly Arg Ser Val Pro Leu Ser Arg Thr Val Arg

465 470 475 480

Cys Thr Cys Ile Ser Ile Ser Asn Gln Pro Val Asn Pro Arg Ser Leu

485 490 495

Glu Lys Leu Glu Ile Ile Pro Ala Ser Gln Phe Cys Pro Arg Val Glu

500 505 510

Ile Ile Ala Thr Met Lys Lys Lys Gly Glu Lys Arg Cys Leu Asn Pro

515 520 525

Glu Ser Lys Ala Ile Lys Asn Leu Leu Lys Ala Val Ser Lys Glu Arg

530 535 540

Ser Lys Arg Ser Pro

545

<210> 181

<211> 1761

<212> DNA

<213> 人工序列

<220>

<223> H16L16-SGRSA-hIL15重链

<400> 181

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 120

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 180

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 360

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 420

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 480

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 540

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 600

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 660

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 720

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 780

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 840

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 900

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 960

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 1020

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1080

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1140

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1200

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1260

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1320

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1380

aagagcctga gcctgtcccc aggctcaggc agatctgcca actgggtgaa tgtaataagt 1440

gatttgaaaa aaattgaaga tcttattcaa tctatgcata ttgatgctac tttatatacg 1500

gaaagtgatg ttcaccccag ttgcaaagta acagcaatga agtgctttct cttggagtta 1560

caagttattt cacttgagtc cggagatgca agtattcatg atacagtaga aaatctgatc 1620

atcctagcaa acaacagttt gtcttctaat gggaatgtaa cagaatctgg atgcaaagaa 1680

tgtgaggaac tggaggaaaa aaatattaaa gaatttttgc agagttttgt acatattgtc 1740

caaatgttca tcaacacttc t 1761

<210> 182

<211> 587

<212> PRT

<213> 人工序列

<220>

<223> H16L16-SGRSA-hIL15重链

<400> 182

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Gly Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser

35 40 45

Ile Ser Ser Ser Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile

115 120 125

Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly

130 135 140

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

145 150 155 160

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

165 170 175

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

180 185 190

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val

195 200 205

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

210 215 220

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys

225 230 235 240

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

245 250 255

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

275 280 285

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

290 295 300

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

305 310 315 320

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

355 360 365

Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln

370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

405 410 415

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

420 425 430

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

435 440 445

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

450 455 460

Leu Ser Pro Gly Ser Gly Arg Ser Ala Asn Trp Val Asn Val Ile Ser

465 470 475 480

Asp Leu Lys Lys Ile Glu Asp Leu Ile Gln Ser Met His Ile Asp Ala

485 490 495

Thr Leu Tyr Thr Glu Ser Asp Val His Pro Ser Cys Lys Val Thr Ala

500 505 510

Met Lys Cys Phe Leu Leu Glu Leu Gln Val Ile Ser Leu Glu Ser Gly

515 520 525

Asp Ala Ser Ile His Asp Thr Val Glu Asn Leu Ile Ile Leu Ala Asn

530 535 540

Asn Ser Leu Ser Ser Asn Gly Asn Val Thr Glu Ser Gly Cys Lys Glu

545 550 555 560

Cys Glu Glu Leu Glu Glu Lys Asn Ile Lys Glu Phe Leu Gln Ser Phe

565 570 575

Val His Ile Val Gln Met Phe Ile Asn Thr Ser

580 585

<210> 183

<211> 1914

<212> DNA

<213> 人工序列

<220>

<223> H16L16-SGRSA-hIFNa重链

<400> 183

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 120

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 180

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 360

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 420

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 480

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 540

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 600

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 660

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 720

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 780

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 840

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 900

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 960

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 1020

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1080

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1140

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1200

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1260

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1320

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1380

aagagcctga gcctgtcccc aggctcaggc agatctgcct gtgatctgcc tcaaacccac 1440

agcctgggta gcaggaggac cttgatgctc ctggcacaga tgaggaaaat ctctcttttc 1500

tcctgcttga aggacagaca tgactttgga tttccccagg aggagtttgg caaccagttc 1560

caaaaggctg aaaccatccc tgtcctccat gagatgatcc agcagatctt caatctcttc 1620

agcacaaagg actcatctgc tgcttgggat gagaccctcc tagacaaatt ctacactgaa 1680

ctctaccagc agctgaatga cctggaagcc tgtgtgatac agggggtggg ggtgacagag 1740

actcccctga tgaaggagga ctccattctg gctgtgagga aatacttcca aagaatcact 1800

ctctatctga aagagaagaa atacagccct tgtgcctggg aggttgtcag agcagaaatc 1860

atgagatctt tttctttgtc aacaaacttg caagaaagtt taagaagtaa ggaa 1914

<210> 184

<211> 638

<212> PRT

<213> 人工序列

<220>

<223> H16L16-SGRSA-hIFNa重链

<400> 184

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Gly Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser

35 40 45

Ile Ser Ser Ser Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile

115 120 125

Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly

130 135 140

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

145 150 155 160

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

165 170 175

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

180 185 190

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val

195 200 205

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

210 215 220

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys

225 230 235 240

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

245 250 255

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

275 280 285

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

290 295 300

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

305 310 315 320

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

355 360 365

Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln

370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

405 410 415

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

420 425 430

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

435 440 445

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

450 455 460

Leu Ser Pro Gly Ser Gly Arg Ser Ala Cys Asp Leu Pro Gln Thr His

465 470 475 480

Ser Leu Gly Ser Arg Arg Thr Leu Met Leu Leu Ala Gln Met Arg Lys

485 490 495

Ile Ser Leu Phe Ser Cys Leu Lys Asp Arg His Asp Phe Gly Phe Pro

500 505 510

Gln Glu Glu Phe Gly Asn Gln Phe Gln Lys Ala Glu Thr Ile Pro Val

515 520 525

Leu His Glu Met Ile Gln Gln Ile Phe Asn Leu Phe Ser Thr Lys Asp

530 535 540

Ser Ser Ala Ala Trp Asp Glu Thr Leu Leu Asp Lys Phe Tyr Thr Glu

545 550 555 560

Leu Tyr Gln Gln Leu Asn Asp Leu Glu Ala Cys Val Ile Gln Gly Val

565 570 575

Gly Val Thr Glu Thr Pro Leu Met Lys Glu Asp Ser Ile Leu Ala Val

580 585 590

Arg Lys Tyr Phe Gln Arg Ile Thr Leu Tyr Leu Lys Glu Lys Lys Tyr

595 600 605

Ser Pro Cys Ala Trp Glu Val Val Arg Ala Glu Ile Met Arg Ser Phe

610 615 620

Ser Leu Ser Thr Asn Leu Gln Glu Ser Leu Arg Ser Lys Glu

625 630 635

<210> 185

<211> 1923

<212> DNA

<213> 人工序列

<220>

<223> H16L16-PSSRRRVN-hIFNa重链

<400> 185

atggaaaccg acaccctgct gctgtgggtg ctgctgctct gggtcccagg ctctaccggc 60

caggtgcagc tgcaggagtc gggaccagga ctggtgaagc cttcggggac cctgtccctc 120

acctgcgctg tctctggtgg ctccatcagc agtagtaact ggtggagttg ggtccgccag 180

cccccaggga aggggctgga gtggattggg gaaatctatc atagtgggag caccaactac 240

aacccgtccc tcaagagtcg agtcaccata tcagtagaca agtccaagaa ccagttctcc 300

ctgaagctga gctctgtgac cgccgcggac acggccgtgt attactgtgc gagatggacc 360

gggcgtactg atgcttttga tatctggggc caagggacaa tggtcaccgt ctcaagcgcg 420

tccacaaagg gcccaagcgt gttcccgcta gcccccagca gcaagagcac cagcggcggc 480

acagccgccc tgggctgcct ggtgaaggac tacttccccg agcccgtgac cgtgtcctgg 540

aacagcggag ccctgacctc cggcgtgcac accttccccg ccgtgctgca gagcagcggc 600

ctgtacagcc tgagcagcgt ggtgaccgtg cccagcagca gcctgggcac ccagacctac 660

atctgtaacg tgaaccacaa gcccagcaac accaaggtgg acaagagagt ggagcccaag 720

agctgtgaca agacccacac ctgtcccccc tgcccagccc ccgagctgct gggcggaccc 780

agcgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag aacccccgag 840

gtgacctgtg tggtggtgga cgtgtcccac gaggacccag aggtgaagtt caactggtac 900

gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc 960

acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaggag 1020

tacaagtgta aggtgtccaa caaggccctg ccagccccaa tcgaaaagac catcagcaag 1080

gccaagggcc agccaagaga gccccaggtg tacaccctgc cacccagcag ggaggagatg 1140

accaagaacc aggtgtccct gacctgtctg gtgaagggct tctacccaag cgacatcgcc 1200

gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg 1260

gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagag cagatggcag 1320

cagggcaacg tgttcagctg ctccgtgatg cacgaggccc tgcacaacca ctacacccag 1380

aagagcctga gcctgtcccc aggcccgagc agccggagaa gagtgaactg tgatctgcct 1440

caaacccaca gcctgggtag caggaggacc ttgatgctcc tggcacagat gaggaaaatc 1500

tctcttttct cctgcttgaa ggacagacat gactttggat ttccccagga ggagtttggc 1560

aaccagttcc aaaaggctga aaccatccct gtcctccatg agatgatcca gcagatcttc 1620

aatctcttca gcacaaagga ctcatctgct gcttgggatg agaccctcct agacaaattc 1680

tacactgaac tctaccagca gctgaatgac ctggaagcct gtgtgataca gggggtgggg 1740

gtgacagaga ctcccctgat gaaggaggac tccattctgg ctgtgaggaa atacttccaa 1800

agaatcactc tctatctgaa agagaagaaa tacagccctt gtgcctggga ggttgtcaga 1860

gcagaaatca tgagatcttt ttctttgtca acaaacttgc aagaaagttt aagaagtaag 1920

gaa 1923

<210> 186

<211> 641

<212> PRT

<213> 人工序列

<220>

<223> H16L16-PSSRRRVN-hIFNa重链

<400> 186

Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro

1 5 10 15

Gly Ser Thr Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val

20 25 30

Lys Pro Ser Gly Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser

35 40 45

Ile Ser Ser Ser Asn Trp Trp Ser Trp Val Arg Gln Pro Pro Gly Lys

50 55 60

Gly Leu Glu Trp Ile Gly Glu Ile Tyr His Ser Gly Ser Thr Asn Tyr

65 70 75 80

Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Lys Ser Lys

85 90 95

Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala

100 105 110

Val Tyr Tyr Cys Ala Arg Trp Thr Gly Arg Thr Asp Ala Phe Asp Ile

115 120 125

Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly

130 135 140

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

145 150 155 160

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

165 170 175

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

180 185 190

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val

195 200 205

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

210 215 220

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys

225 230 235 240

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

245 250 255

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

275 280 285

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

290 295 300

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

305 310 315 320

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

355 360 365

Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln

370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

405 410 415

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

420 425 430

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

435 440 445

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

450 455 460

Leu Ser Pro Gly Pro Ser Ser Arg Arg Arg Val Asn Cys Asp Leu Pro

465 470 475 480

Gln Thr His Ser Leu Gly Ser Arg Arg Thr Leu Met Leu Leu Ala Gln

485 490 495

Met Arg Lys Ile Ser Leu Phe Ser Cys Leu Lys Asp Arg His Asp Phe

500 505 510

Gly Phe Pro Gln Glu Glu Phe Gly Asn Gln Phe Gln Lys Ala Glu Thr

515 520 525

Ile Pro Val Leu His Glu Met Ile Gln Gln Ile Phe Asn Leu Phe Ser

530 535 540

Thr Lys Asp Ser Ser Ala Ala Trp Asp Glu Thr Leu Leu Asp Lys Phe

545 550 555 560

Tyr Thr Glu Leu Tyr Gln Gln Leu Asn Asp Leu Glu Ala Cys Val Ile

565 570 575

Gln Gly Val Gly Val Thr Glu Thr Pro Leu Met Lys Glu Asp Ser Ile

580 585 590

Leu Ala Val Arg Lys Tyr Phe Gln Arg Ile Thr Leu Tyr Leu Lys Glu

595 600 605

Lys Lys Tyr Ser Pro Cys Ala Trp Glu Val Val Arg Ala Glu Ile Met

610 615 620

Arg Ser Phe Ser Leu Ser Thr Asn Leu Gln Glu Ser Leu Arg Ser Lys

625 630 635 640

Glu