Intratumoral Delivery of a PD-1-blocking scFv encoded in Oncolytic HSV-1 Promotes Antitumor Immunity and Synergizes with TIGIT Blockade

恶性肿瘤已严重威胁人类健康和生命，现有的治疗手段远远未能满足临床需求。厦门大学研究团队联合浙江养生堂生物科技有限公司、养生堂有限公司进行协同攻关，研制出新一代肿瘤免疫治疗药物——“注射用重组人PD-1抗体单纯疱疹病毒”。研究发现，重组表达PD-1单链抗体的溶瘤病毒具有“双药合一”抗肿瘤的突出优势，提高肿瘤治愈率。相关成果于2020年3月3日以研究论文形式在线发表于Cancer Immunology Research（《癌症免疫学研究》）期刊，为指导新型溶瘤病毒的升级改造和突破肿瘤免疫耐受提供了新的思路，为重组表达PD-1单链抗体的溶瘤病毒药物运用于肿瘤临床治疗奠定了坚实的理论基础。 厦门大学公共卫生学院夏宁邵教授和黄承浩助理教授为该论文的共同通讯作者，博士生林超龙和任文峰为该论文的共同第一作者。【Abstract】Oncolytic virotherapy can lead to systemic antitumor immunity, but the therapeutic potential of oncolytic viruses (OVs) in humans is limited due to their insufficient ability to overcome the immunosuppressive tumor microenvironment (TME). Here, we showed that locoregional oncolytic virotherapy upregulated the expression of PD-L1 in the TME, which was mediated by virus-induced type I and type II interferons (IFNs). To explore PD-1/PD-L1 signaling as a direct target in tumor tissue, we developed a novel immunotherapeutic herpes simplex virus (HSV), OVH-aMPD-1, that expressed a single-chain variable fragment (scFv) against PD-1 (aMPD-1 scFv). The virus was designed to locally deliver aMPD-1 scFv in the TME to achieve enhanced antitumor effects. This virus effectively modified the TME by releasing damage associated molecular patterns (DAMPs), promoting antigen cross-presentation by dendritic cells, and enhancing the infiltration of activated T cells; these alterations resulted antitumor T cell activity which led to reduced tumor burdens in a liver cancer model. Compared with OVH, OVH-aMPD-1 promoted the infiltration of myeloid-derived suppressor cells (MDSCs),resulting in significantly higher percentages of CD155+ G-MDSCs and M-MDSCs in tumors. In combination with TIGIT blockade, this virus enhanced tumor-specific immune responses in mice with implanted subcutaneous tumors or invasive tumors. These findings highlighted that intratumoral immunomodulation with an OV expressing aMPD-1 scFv could be an effective standalone strategy to treat cancers or drive maximal efficacy of a combination therapy with other immune checkpoint inhibitors.This work was supported by grant 2018ZX10301404-001-002 from the National Science and Technology Major Project and grant 81571990 from the National Natural Science Foundation of China.该研究获得了国家自然科学基金、国家科技重大专项的资助

A novel approach for rapid high-throughput selection of recombinant functional rat monoclonal antibodies

Abstract Background Most monoclonal antibodies against mouse antigens have been derived from rat spleen-mouse myeloma fusions, which are valuable tools for purposes ranging from general laboratory reagents to therapeutic drugs, and yet selecting and expressing them remains a time-consuming and inefficient process. Here, we report a novel approach for the rapid high-throughput selection and expression of recombinant functional rat monoclonal antibodies with different isotypes. Results We have developed a robust system for generating rat monoclonal antibodies through several processes involving simultaneously immunizing rats with three different antigens expressing in a mixed cell pools, preparing hybridoma cell pools, in vitro screening and subsequent cloning of the rearranged light and heavy chains into a single expression plasmid using a highly efficient assembly method, which can decrease the time and effort required by multiple immunizations and fusions, traditional clonal selection and expression methods. Using this system, we successfully selected several rat monoclonal antibodies with different IgG isotypes specifically targeting the mouse PD-1, LAG-3 or AFP protein from a single fusion. We applied these recombinant anti-PD-1 monoclonal antibodies (32D6) in immunotherapy for therapeutic purposes that produced the expected results. Conclusions This method can be used to facilitate an increased throughput of the entire process from multiplex immunization to acquisition of functional rat monoclonal antibodies and facilitate their expression and feasibility using a single plasmid