Abstract LB360: Type-1 interferon sensing is critical for CD8+ resident memory (TRM) cell generation in tumor-draining lymph nodes

CD8+ tissue resident memory T cells (TRM) are potent mediators of anti-tumor immunity. We have previously identified a population of lymph node resident memory T cells (LN TRM) that are critical for protection against melanoma growth in tumor-draining lymph nodes (TDLN). However, the precursor population that gives rise to these cells, and mechanisms supporting their generation are not understood. Here we sought to define the features of tumor-specific T cells that govern their fate as TRM cells in TDLNs. We employed our previously described model wherein depletion of CD4+ T cells induces the generation of protective melanoma Ag-specific TRM cells throughout skin and LNs of B16 tumor-excised mice. To map the fate of TRM precursor cells (pre-TRM), we implemented paired scTCR/RNA sequencing of antigen-experienced (CD44+ CD8+) T cells from surgically harvested TDLNs of individual mice and identified clonotypes that matched to differentiated TRM cells in LNs and skin of the same mice 30 days later. Additionally, we tracked gp100-specific CD8+ transgenic T cells (pmel cells) in vivo to define requirements for LN TRM generation. Clonal tracing revealed that expanded clonotypes in TDLNs differentiated into TRM cells in LNs and skin of tumor-excised mice. Interestingly, there were no apparent differences in the transcriptional properties of pre-TRM clonotypes (in early TDLNs) that acquired a TRM fate in the skin versus the LN. These pre-TRM clonotypes in TDLNs clustered into three major subsets, one of which appeared terminally differentiated with high expression of cell cycle genes, and two of which had high expression of genes associated with memory (Il7r, Tcf7, Slamf6) and tissue residency (Cd69, Itgae, Cxcr6, Cxcr3). One of the two memory-like precursor clusters additionally expressed high levels of interferon responsive genes (Isg15, Isg20, Ifit1, Ifi47). Experiments involving pmel cell adoptive transfer and FTY720 treatment showed that pre-TRM cells seeded LNs early, underscoring the existence of a committed TRM precursor population. Based on the pronounced IFN-sensing signature in pre-TRM clonotypes, we sought to define a role for IFN in TRM formation. Using Mx-1-GFP reporter pmel cells, we found that type-1 IFN sensing reached peak levels in TDLNs during tumor growth. To test whether type-1 IFN sensing was required for TRM generation, responses were assessed in IFNAR1 KO mice and additionally in mice that received wild-type vs. IFNAR1 KO pmel T cells. Interestingly, in both cases the absence of IFNAR1 resulted in a reduction in TRM generation TDLNs but not in skin. Overall, these results reveal the features of tumor-specific pre-TRM cells in TDLNs, and demonstrate that early type 1 IFN sensing dictates tissue-specific TRM fate decisions, predisposing to TRM generation in LNs. Citation Format: Nikhil Khatwani, Aleksey Molodtsov, Cameron Messier, Jichang Han, Tyler Glenn Searles, Aaron R Hawkes, Fred W Kolling IV, Mary Jo Turk. Type-1 interferon sensing is critical for CD8+ resident memory (TRM) cell generation in tumor-draining lymph nodes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB360.