Therapeutic Induction of Tertiary Lymphoid Structures in Melanoma using STING Agonists

Abstract

Abstract Tertiary lymphoid structures (TLS) are non-encapsulated functional aggregates of T cells, B cells, dendritic cells (DC) and high endothelial venules (HEV) located in peripheral sites of chronic inflammation. TLS may serve as sites of local antigen presentation and immune priming that may protect against tumor progression. Homeostatic chemokines CCL19, CCL21 and CXCL13 produced by DCs and/or stromal cells are known to support secondary lymphoid organogenesis and also play key roles in TLS formation. Other DC-associated pro-inflammatory cytokines including lymphotoxin α, IL-36γ and type I interferons also appear to aid in organizing TLS. Hence, we hypothesize that appropriate therapeutic activation of DCs within the tumor microenvironment may serve to nucleate TLS in vivo, leading to slowed tumor progression. We found that activation of the cytosolic DNA sensor STING using ML-RR-S2-CDA, a murine and human STING agonist, leads to not only an expected increased production of type I interferons by CD11c+ DCs in vitro but also leads to increased transcript levels of TLS-associated cytokines/chemokines via activation of pIRF3 and pIRF7. In mice harboring B16.F10 melanomas, provision of STING agonists slows tumor growth in a host STING dependent manner. Further using immunofluorescence microscopy, we observed increased infiltration and clustering of CD3+ T cells and CD11c+ DCs and the induction of HEVs in treated tumors within tumor-associated TLS as early as 5 days post-treatment. Future studies will determine how TLS shape the local versus systemic protective T cell repertoire in treated animals.