Abstract 2366: Balancing efficacy and toxicity of a fibroblast activation protein dependent tetravalent CD40 agonist antibody for cancer therapy

Abstract Background: CD40 agonist antibodies have been found to augment tumor antigen presentation and have shown potential as anti-tumor treatments in human clinical trials. The clinical success of CD40 agonist antibodies has been restricted due to their limited efficacy and on-target, off-tumor toxicity. We hypothesized that toxicity associated with CD40 agonist antibodies could be overcome by activating CD40 specifically through tumor-specific antigens. Additionally, limited efficacy can be improved by rational design of CD40 bispecific antibodies (BsAbs). Herein, we present FAPxCD40, a trivalent CD40 agonist BsAb targeting fibroblast activation protein (FAP) possessing an optimized therapeutic window. Method: We generated a series of FAPxCD40 bispecific antibodies with 2, 4, and 6 CD40 valences by using a high-affinity anti-FAP antibody and a crosslink-dependent CD40 agonist VHH antibody. These BsAbs were evaluated using multiple immune cell functional assays, including assays for dendritic cell (DC) maturation, B cell maturation, and CMV-reactive T cell recall. The in vivo therapeutic window of the FAPxCD40 bispecifics was compared in CD40 humanized mice with subcutaneously transplanted FAP-positive MC38 tumor xenografts. Results: We developed FAPxCD40 BsAbs with varying valences, including bivalent FAPxCD40-2, tetravalent FAPxCD40-4, and hexavalent FAPxCD40-6. FAPxCD40-2 showed strict FAP-dependent CD40 agonist activity in both DC and B cell maturation assays, while its CD40 agonist activity upon FAP-crosslinking was modest. FAPxCD40-4 exhibited efficient CD40 agonist activity upon FAP-crosslinking, but however, it also exhibited some undesired FAP-independent CD40 agonist activity. FAPxCD40-6 efficiently clustered CD40 but marginally benefited from FAP binding, making it unsuitable as a choice. In the humanized CD40 mice model, FAPxCD40-2 effectively eliminated systemic toxicity at all tested doses. Its antitumor activity, however, is modest with TGI (tumor growth inhibition) rate of 32. 8 % at a dose of 6. 96 mpk. Further escalation of the FAPxCD40-2 dosage to 13. 92 mpk did not result in enhanced antitumor activity with TGI rate of 46. 9 %, indicating that its CD40 agonist activity may have reached a plateau. In contrast, FAPxCD40-4 broke this plateau and induce a promising antitumor response at a dose of 1. 3 mpk, resulting in a 92% TGI rate. Furthermore, no liver enzyme upregulation or platelet count decrease was observed under these conditions, indicating an ideal therapeutic window. In addition, during nonclinical toxicology studies conducted on cynomolgus monkeys, it was found that doses of FAPxCD40-4 up to 99 mg/kg administered weekly for a total of four doses were well-tolerated. Conclusion: Our study demonstrates that FAPxCD40-4, a tetravalent FAPxCD40 BsAb, exhibits efficient CD40 agonist activity with an optimized therapeutic window. Citation Format: Simeng Chen, Yuan Lin, Xiaoru Zhou, Xing Sun, Changyong Yang, Cheng Liao. Balancing efficacy and toxicity of a fibroblast activation protein dependent tetravalent CD40 agonist antibody for cancer therapy abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 2366.