Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

Cytokines dimerize two receptor chains to activate Janus kinases and STAT transcription factors that regulate immune cells but have therapeutic liabilities. We engineered “Trikines” to compel cis formation of three-chain cytokine receptor complexes at the cell surface that induce bespoke STAT transcriptional signaling programs optimized for therapeutic efficacy. Designed Trikines co-activated pSTAT5 and pSTAT3 signatures distinct from any natural cytokines, by assembling trimeric combinations of Interleukin-2 (IL-2), Interleukin-10 (IL-10), and Interleukin-21 (IL-21) receptors. An IL-2-based-Trikine restrained terminal differentiation of T cells, promoted stemness, and enhanced durability of tumor control without toxicity. Unexpectedly, an IL-10-based Trikine induced immune infiltration into poorly immunogenic tumors, showing striking efficacy in small cell lung cancer and pancreatic cancer models. Trikines obviate the need for cell engineering to customize STAT signatures for immunotherapy Overall design: Mice were inoculated s.c. with 2.5 × 105 6694c2vTRP1 cells and received i.p. injections of PBS, mono-IL-10 (26 µg/mouse, n = 5), or IL-10/2-Trikine (30.6 µg/mouse, n = 5) every other day starting on day 8, for a total of two doses. Mice were sacrificed on day 14. CD45? TILs were bead-isolated, individually labeled with hashtag antibodies, pooled by treatment group, and subjected to Single Cell 5' RNA and cell surface protein sequencing.