Abstract 3548: Systemic antitumor immunity stimulated by intratumoral CD40 agonist in combination with PD1 therapy in immune-cold melanoma

Abstract Agonistic CD40 antibodies in combination with checkpoint inhibitors have shown beneficial outcomes in clinical trials for the treatment of malignancies. We demonstrated that intratumoral injection of CD40 agonist in combination with systemic anti-PD1 therapy activates innate immune responses and promotes selective expansion of effector CD8 T cells while decreasing immunosuppressive Treg and M2 populations in melanoma tumors. However, the mechanisms of systemic antitumor immune responses driven by this combination therapy in patients are not well defined. Therefore, the aim of this study was to evaluate the impact of this combination therapy on systemic antitumor immunity using a bilateral, immunologically “cold” tumor model. Mice bearing bilateral B16-F10 melanoma tumors were injected with anti-CD40 Ab into right flank tumors, either alone or in combination with anti-PD1 Ab on day 8 post-tumor implantation. Mice were received treatment every 3 days for a total of 5 doses. Our results demonstrated that intratumoral CD40 Ab alone or in combination with anti-PD1 therapy inhibited the growth of both treated and distant (non-treated) tumors. The combination therapy showed superior efficacy compared with single Ab treatments. CD40 Ab-injected tumor weights measured at day 17, were ∼9-fold reduced in mice treated with the combination therapy as compared to PD1 Ab alone (0.09±0.02g vs 0.83±0.17g). Importantly, similar antitumor efficacy of the combination therapy was observed in non-treated tumors (0.09±0.02g vs 0.74±0.23g) on the left flanks of these mice, demonstrating an abscopal treatment effect that was associated with enlargement of spleen and TdLNs. Flow cytometry analysis showed that CD40 agonist stimulated the proliferation of CD4 and CD8 T cells, and B cells in TdLNs and spleen. Furthermore, CD40 Ab treatment increased the frequency of blood circulating effector (40% vs 7% CTR) and central memory (25% vs 15% CTR) CD8 T cells as determined at day 21. Our data indicated that intratumoral CD40 agonist in combination with anti-PD1 therapy stimulated systemic antitumor immunity resulting in inhibition of distant tumor growth in a “cold” melanoma model, which was accompanied by increased proliferation of T cells and B cells in lymphoid organs and frequency of effector and memory CD8 T cells in the circulation. The results provide new insights about the mechanisms of systemic antitumor responses induced by intratumoral CD40 agonist. Citation Format: Barbara Pazdrak, Heather M. Sonnemann, Barbara M. Nassif, Salah E. Bentebibel, Greg Lizee, Adi Diab. Systemic antitumor immunity stimulated by intratumoral CD40 agonist in combination with PD1 therapy in immune-cold melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3548.