Biogel Delivery of Sting Ligands is Effective in a Murine Model of Locally Advanced Pancreatic Adenocarcinoma, and is Potentiated by Local CCR2 Inhibition

Background: STING (Stimulator of INterferon Genes) ligands represent a novel class of immunotherapy compounds with potential application in locally advance pancreatic cancer (LAPC). We have previously generated a preclinical model, and performed some initial experiments evaluating its safety and use in a preclinical vascular encasement model. Here we demonstrate their use as a bioresorbable depot PLGA-PEG-PLGA (P3), through combination with an injectable thermo-gel in margin accentuation and direct injection strategies. In addition, we identify synergy with regional CCR2 blockade using a small molecule inhibitor. Using knockout mice, we characterize the role of key inflammatory mediators in this response. Methods: A previously described model of vascular encasement was established in the femoral triangle of C57BL/6 mice using the PANC-02 cell line. ML-RR-S2-CDA (S100), is a modified STING agonist was used in conjunction with P3 to create an injectable depot using a 15% W/V solution containing S100. This was applied to the resection bed of partially resected flank tumors. Tumors established within the femoral triangle were injected by palpation, or ultrasound guidance. Combination with local CCR2 inhibition was evaluated in a subcutaneous flank model using a sub-therapeutic dose of S100 (10ug), combined with a titration of PF-4136309, a small molecule inhibitor of CCR2. Finally, the mechanism of combined STING activation and CCR2 inhibition was interrogated using CD8 depletion, and mouse knockouts of TNFa, STING and IFNAR. Results: Within partially resected flank tumors, application of S100 in P3resulted in durable cure, with an increased fraction of tumor clearance as the dose increased (Survival at 60 days, PBS control 0%, S100-25ug: 14%, S100- 50ug: 60% p = 0.0008). When subtherapeutic doses of S100 (10ug) were combined with CCR2 inhibition, improved survival was noted (Survival at 60 days, S100/PF-4136309 0ug/0ug: 0%, 10 ug/0 ug: 0%, 0ug/100ug 0%, 10 ug/25 ug 33%, P = <0.0001). When injected into femoral tumors, serial injection of S100 delayed tumor outgrowth, while injection of S100 - 400 ug in P3, or 200 ug with 100 ug PF-4136309 resulted in tumor regression, but only when performed under ultrasound guidance. When injections of S100 10 ug, and PF-4136309 25 ug were used with PLGA-PEG-PLGA in CD8a depletion, or knockouts, survival was markedly reduced in IFNAR-/-, TNFa-/-, STING-/-, and less dramatically in mice treated with CD8 depletion (P < 0.0001). Conclusion: STING ligands can be used in conjunction with thermogel delivery systems to produce durable tumor regression in both direct injection and margin accentuation applications. This effect is potentiated by CCR2 inhibition. These results suggest that STING activation is a flexible approach that can be potentiated through biomaterial delivery strategies, and CCR2 inhibition to achieve high response rates even with advanced tumors.