人类激动型CD40抗体Fc段的活性分析及突变体筛选

Targeted therapy based on monoclonal antibody is one of the important means of cancer immunotherapy. CD40 and other TNFR superfamily members control many key signaling pathways involved in antitumor immune responses, and are hot targets for the development of antitumor therapies. Agonistic antibodies can effectively activate these signaling pathways and are of significant antitumor activities, and are therefore of great potential as antitumor therapies. So far there has been no successful agonistic antibody drug which recognize CD40 or other TNFR superfamily members developed. Recent studies based on murine antibodies have found that the in vivo activity of the agonistic anti-TNFR superfamily antibodies depend on the interaction with the inhibitory FcγR, whereas the interaction with the activated FcγRs will decrease its activity. For example, in mice improved specificity antibody binding ability to inhibitory receptors, can significantly increase its anti-tumor activity. Therefore, this project is planned to study the effect of different IgG subtypes Fc on the immune activation and anti-tumor activity of the agonistic antibodies by using the human anti-CD40 antibody as the research object. Based on this, we plan to screen the stronger activity human agonistic anti-CD40 mutant antibody by Fc engineering. This will provide favorable conditions for development of such human anti-tumor antibody drugs successfully.

以单克隆抗体为基础的靶向治疗是肿瘤免疫治疗的重要手段之一。CD40等TNFR超家族成员控制抗肿瘤免疫应答的许多关键信号通路，是抗肿瘤药物研发的热门靶点。激动型抗体能够有效的激活这些信号通路因而具有显著的抗肿瘤效果和广阔的应用前景。然而，到目前为止还没有一个识别CD40或其它TNFR超家族成员的激动型抗体药物研制成功。以鼠类抗体为基础的最新研究发现激动型抗TNFR超家族成员抗体的体内活性依赖于与抑制性FcγR的相互作用，而与活化性FcγRs的相互作用反而会降低其活性。例如，在小鼠体内特异性提高抗体与抑制性FcγR的结合能力，能够显著提高其抗肿瘤活性。因此本项目计划以人类激动型CD40抗体为研究对象，研究不同IgG亚型Fc对人类激动型抗体的免疫激活与抗肿瘤活性的影响。并以此为基础通过定向改造筛选出具有更强活性的人类激动型抗CD40突变抗体，为该类抗肿瘤抗体药物的研发成功提供有利条件。

以单克隆抗体为基础的靶向治疗是肿瘤免疫治疗的重要手段之一。CD40等TNFR超家族成员控制抗肿瘤免疫应答的许多关键信号通路，是抗肿瘤药物研发的热门靶点。激动型抗体能够有效的激活这些信号通路因而具有显著的抗肿瘤效果和广阔的应用前景。然而，到目前为止还没有一个识别CD40或其它TNFR超家族成员的激动型抗体药物研制成功。本研究以人类激动型CD40抗体为研究对象，研究不同IgG亚型Fc对人类激动型抗体的免疫激活与抗肿瘤活性的影响，发现四种IgG激动型抗体中，IgG2具有最强的免疫激活活性。本研究建立了哺乳动物表面展示系统用于抗体恒定区的筛选平台：利用PCR技术构建了大容量的IgG2 Fc突变文库并转染进入细胞，流式细胞分选与抑制型FcγRIIB高亲和力的突变体，然后通过高通量测序技术分析获得具有与FcγRIIB结合有利的突变。结合亲和力和活性分析，本项目筛选到多个与抑制性Fcγ受体分子结合能力增强的IgG2恒定区变体，并且这些抗体的体外和体内免疫激活活性明显增强。该项目获得的候选恒定区突变序列以及建立的高通量突变体筛选方法均可用于后续抗肿瘤抗体药物的研发，为优化激动型抗体活性提供了新的思路和有利条件。我们已经就项目建立的激动型抗体恒定区优化策略和序列申请专利一项（李福彬、张燕等，FcγRIIB亲和力增强的抗体Fc区，中国专利申请号：201911223227.1；PCT/CN2020/133685）；同时以这些研究为基础的成果目前处于数据整理阶段，计划近期以学术论文的形式发表。目前已经和药物研发企业合作进行激动型抗体药物的研发，签署技术开发合同一项。