(592b) “Smart” Nanoparticles for Immunotherapeutic Targeting of the Sting Pathway

Immune checkpoint inhibitors are transforming cancer treatment, yet the majority of patients fail to achieve complete and durable responses. Resistance to checkpoint blockade is associated with non-immunogenic tumors that lack significant T cell infiltration and are instead characterized by a highly immunosuppressive tumor microenvironment (TME). To reprogram such “cold” TMEs to a more “hot” T cell-inflamed phenotype, we are developing pH-responsive, “smart” polymer nanoparticles (NPs) for efficient cytosolic delivery of cyclic dinucleotides (CDNs), including 2’3’-cGAMP (cyclic [G(2’,5’)pA(3,5’)p]), the natural and endogenous high affinity ligand of the stimulator of interferon genes (STING) pathway. The STING pathway is the primary mechanism by which the innate immune system senses cancer cells to stimulate a localized inflammatory response that is essential to endogenous anti-tumor T cell adaptive immunity. This critical role of STING in cancer immune surveillance provides compelling rationale for the use of cGAMP and structurally related CDN STING agonists as therapeutics for reversing immunosuppression in the TME and generating robust anti-tumor T cell responses in the setting of non-immunogenic tumors. However, as an anionic compound, cGAMP does not readily cross cellular membranes, is poorly endocytosed, and, critically, enters the cytosol where STING is localized with very low efficiency. Moreover, CDNs are rapidly cleared from the body with modest delivery to tumors and/or lymph nodes. These major drug delivery barriers limit the therapeutic potential of cGAMP and other structurally related CDNs. To solve these challenges, we have designed polymer vesicles - polymersomes - that promote efficient endosomal escape of cGAMP to the cytosol. cGAMP can be efficiently loaded into NPs (38% encapsulation efficiency) with an average diameter of 100 nm, a neutral surface charge, and low cytotoxicity. NP delivery of cGAMP dramatically enhances STING activation in vitro relative to free cGAMP, which elicits only low levels of type-I interferon (IFN-I) production even at a 500-1000x higher dose in vitro. In established subcutaneous (SC) B16F10 melanoma tumors, intratumoral (IT) administration of nanoparticulate cGAMP formulations converted tumors into an inflamed phenotype, characterized by elevated IFN-I production, dendritic cell maturation, T-cell infiltration, and polarization of immunosuppressive M2 macrophages towards an anti-tumor M1 phenotype. These shifts in the TME corresponded with an inhibition of tumor growth for mice treated with STING-activating nanoparticles, with a subset of mice demonstrating complete tumor regression, long-term survival, and systemic antitumor immunity that protected against tumor re-challenge. Collectively, these data demonstrate that endosomolytic polymer nanoparticles significantly improve the therapeutic potency of cGAMP, and establish a pre-clinical foundation for the use of STING-activating nanoparticles as a strategy for increasing tumor immunogenicity and improving responses to immune checkpoint blockade.