The synergy of TLR and STING in cancer immunity

Abstract Adjuvants are essential components of cancer vaccine formulations to promote effective immune responses. We initially screened multiple immune-activating targets, including TLR and STING adjuvants using in-vitro assays to test their ability to activate bone marrow DC (BMDC) and showed STING had the most potent activation (i.e., MHC and co-stimulatory molecules) on DCs. Since STING and TLR signaling is non-redundant, TLR and STING adjuvants were combined to enhance DC activation. We utilized the multidimensional synergy of combinations (MuSyc), a novel synergy algorithm, to assess molecular synergy in combining STING and TLR adjuvants, and we noted that the combination of R848 (TLR7-8) plus STING agonists provided a synergistic efficacious and potent response on BMDCs. From this data-set, we generated a MuSYC synergy strategy where we will utilize max signal dose for STING adjuvant and ten times less of max signal dose for R848 (TLR7-8) to determine synergy for other assays. We tested this same MuSYC strategy for activation of human monocytic cell line THP-1 and determined that the synergy strategy induced similar or better activation effects compared to the max signal dose for both adjuvants. Finally, we tested the in-vivo anti-tumor immune response for the B16 melanoma and MOC2 head/neck tumor-bearing vaccination models that resulted in an improved anti-tumor response for the combinatorial STING-TLR adjuvanted vaccines compared to the single agents.