Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics [scRNAseq of IL21 Trikine]

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. IL-2-based-Trikines restrain terminal differentiation of T cells, promote stemness, and enhance durability of tumor control. 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: For the low dose regimen of super IL-2(2), sIL-2 + IL-21 (2Pos21), or IL-2/21-trikine(2-trikine) treatment in the B16F10 +pmel ACT model, tumors from mice were harvested on day 18 following tumor injection. Tumorswere minced and dissociated using a gentleMACS Octo Dissociator. TILs were collected forsequencing by sorting for live, single cell, Thy1.1+CD8+ cells on a Sony Cell Sorter SH800S.