Background: Immune checkpoint interaction helps tumors resist immunity-induced apoptosis. Blocking PD-1/PD-L1 leads to remarkable clinical responses; however, a large portion of patients develop acquired resistance after initial therapy. Many tumor-specific antigens (TSAs) and tumor-associated antigens (TAAs) have been identified. TSAs are restricted to cancer cells only but TAAs are elevated on tumor cells and minimally expressed on healthy cells. Therefore, co-targeting TAAs/TSAs would show benefits over monotherapy. CD55 is a TAA that regulates complement activation pathway and expressed on cells exposed to the complement system to protect against complement attack. Cancer cells escape this attack by raising CD55 expression. Additionally, CD55 acts as a virus receptor for internalization. Based on the roles of CD55, we designed PD-L1/CD55 bispecific antibody (GB262) with weaker CD55 binding arm to rule out the possibility of lethality to healthy cells but maintain its CD55 binding and internalization ability specifically to cancer cells. Moreover, co-binding to PD-L1 and CD55 leads to co-internalization of PD-L1 by CD55, enhancing the blocking of PD-1/PD-L1 interaction. Methods: Co-localization and levels of PD-L1 and CD55 on cancer cells were analyzed by microscopy and FACS. T-cell activation was studied using PD-L1+/CD55+ cells and PBMCs. CD97 and CD55 blocking was performed using FACS. Cancer killing, ADCC and CDC were performed by culturing PDL-1+/CD55+ cells with PBMCs, NK cells and human complement serum (HCS), respectively. GB262 internalization and its effects on intracellular PD-L1 were studied by pHrodo and western blot. Saporin-conjugated GB262 efficacy was evaluated by cell binding, internalization and cancer killing. In vivo study was performed by engrafting PBMCs and PANC-1 in B-NDG mice. Results: PD-L1 significantly co-localized with CD55 on cancer cells from different indications. While high PD-L1 binding was maintained, a significant reduction in CD55 binding was observed by GB262 to their receptors. GB262 also blocked the interaction between CD97 and CD55 more efficiently than monoclonals. GB262 induced stronger T-cell activation and cancer killing in presence of PBMCs. Similarly, higher level of ADCC and CDC were also evident by GB262. The degree of GB262 internalization was CD55 level dependent on cells surface and leads to intracellular PD-L1 degradation. When conjugated to saporin, GB262-saporin significantly induced cancer killing in absence of effector cells or HCS, whereas GB262 or saporin had no effect. Both GB262 and GB262-saporin suppressed tumor growth in B-NDG mice models. Conclusion: GB262 is the first bispecific antibody that not only releases cancer repression on T-cell activation, but also releases cancer repression on CDC. By conjugating to a toxin, GB262 could serve as an ADC to target multiple cancer cells. GB262 has potential to serve as a novel therapy for many cancers.

Citation Format: Amit K. Chaudhary, Jiadong Shi, Wenyan Cai, Bo Wang, Mohd S. Zaman, Cai Huang, Jun Lin, Steven Z. Kan, Joe Zhou, Jianbo Dong, Yue Liu. Development of a novel bispecific antibody targeting PD-L1 and CD55 for cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB067.