Abstract B116: Myeloid cell co-activation via Dectin-1 and CD40 agonists coordinates non-classical mechanisms of T cell cytotoxicity in Pancreatic Ductal Adenocarcinoma

Abstract

Immunotherapy efficacy in pancreatic ductal adenocarcinoma (PDA) remains limited due to immune evasion mechanisms like MHCI downregulation which impair T cell–based responses. Strategies engaging alternative cytotoxic pathways are needed to overcome such resistance. We previously demonstrated that coactivation of myeloid cells by targeting the pattern recognition receptor Dectin-1 and activating receptor CD40 drives T cell infiltration into tumors and durable tumor regressions in PDA mouse models. Although CD8+ T cells were necessary for tumor regressions, neither MHCI expression by tumor cells nor perforin were required. Here, we investigated the Fas/Fas-ligand (FasL) axis activated by Dectin-1/CD40 coactivation. Mice were subcutaneously implanted with KPC PDA tumors and treated 14 days later with an injection of Dectin-1 (odetiglucan) and CD40 agonist (FGK45), followed by blocking antibodies every 3 days for 14 days. We found increased frequencies of FasL+ CD8+ T cells in the peripheral blood 7 days after coactivation via flow cytometry (control mean 13.3% CD8+ cells, coactivation 32.5%, p=0.038). Further, scRNAseq revealed a 113% increase in mean Fasl expression by tumor infiltrating CD8+ T cells. IHC demonstrated increased Fas expression on tumor cells after coactivation. This increased Fas expression was partially dependent on IFNg, as aIFNg reduced Fas expression, though not to control levels (mean control Fas+ 2.4% of CK19+ area, coactivation 24.3%, coactivation + aIFNg 10.53%, control vs coactivation p=0.003, coactivation +/- aIFNg p=0.004). In vitro assays demonstrated that the highest levels of Fas expression on PDA cells were induced by the combination of IFNg and IL1B (Fas MFI control 120.9, IL1B 123.2, IFNg 198, IL1B + IFNg 332.3). Notably, scRNAseq revealed a 29.8% increase in mean Il1b expression by tumor macrophages after coactivation, suggesting a role for activated macrophages in sensitizing tumor cells to Fas-induced cell death. FasL blockade abrogated the survival benefit of coactivation therapy in MHCI-knockout tumors (median survival: control 26 days, coactivation undefined, coactivation + aFasL 28 days; control vs coactivation p=0.024, coactivation vs coactivation + aFasL p=0.049). In MHCI-competent tumors, aFasL impaired tumor regressions, as percent change in tumor volume between day 33 and day 0 post-treatment was higher with aFasL (coactivation mean tumor volume percent change -90.86, coactivation + aFasL 61.84, p=0.049). While coactivation therapy induced durable antitumor memory that allowed rejection of an MHCI-knockout tumor rechallenge, FasL blockade during rechallenge impaired this memory response and significantly shortened survival (naïve control median survival 41 days post-tumor implantation, rechallenge control undefined, rechallenge + aFasL 66 days; rechallenge +/- aFasL p=0.005). These studies demonstrate that Dectin-1/CD40 coactivation overcomes PDA immune evasion by engaging FasL-dependent pathways of T cell cytotoxicity, offering a promising immunotherapeutic strategy for refractory tumors.