(789) Role of ADAR1 in regulating IFN and TNF mediated responses

Introduction/Rationale: Immunotherapy has achieved remarkable success in hematologic malignancies, but its efficacy in solid tumors remains limited due to tumor heterogeneity, immune suppression within the tumor microenvironment, and inefficient T cell infiltration. I hypothesize that enhancing tumor cell sensitivity to cytokine-mediated killing can potentiate paracrine killing, in which T cell–derived cytokines induce tumor cell death even without direct cell–cell contact. This mechanism can overcome antigen heterogeneity and limited T cell penetration, broadening the therapeutic reach of immunotherapy.

Methods: I propose targeting ADAR1, an RNA-editing enzyme that suppresses innate immune activation, to enhance paracrine killing. Loss of ADAR1 leads to dsRNA accumulation and activation of PKR, MDA5, and RIG-I, sensitizing tumor cells to T cell–derived cytokines such as IFNγ and TNF. To test this, I performed in vitro paracrine killing co-culture assays using T cells from healthy donor PBMCs, antigen-positive tumor targets, and GFP-labeled antigen-negative ADAR1KO tumor targets. GFP signal loss in ADAR1KO cells will indicate enhanced paracrine killing independent of antigen recognition.

Results: My data show that ADAR1 inhibition sensitizes tumor cells to cytokine-mediated cytotoxicity, though how dsRNA sensor and cytokine receptor signaling intersect remains unclear. Supernatant from antigen-positive T cell–tumor co-cultures induced cell death in ADAR1KO but not WT tumor targets, indicating selective cytokine sensitivity. Consistently, Western blot analysis revealed increased cleaved Caspase-3 and PARP in ADAR1KO cells. The paracrine killing assay is ongoing.

Conclusion: This study aims to elucidate the interplay between dsRNA sensing and cytokine signaling in the absence of ADAR1 and to define how this crosstalk shapes the efficacy of paracrine killing in solid tumors. These insights could inform new therapeutic strategies to enhance the potency of immunotherapy against resistant and heterogeneous solid tumors.