Pyroptosis Modulates Multiple Immune Cell Populations in Targeted Therapy-Treated Melanoma.

Treatment of melanoma with BRAF inhibitors plus MEK inhibitors (BRAFi + MEKi) stimulates an intratumoral immune response, in part through pyroptosis mediated by the pore-forming protein gasdermin E (GSDME/Gsdme). How GSDME mediates effects on tumoral immunity is not well characterized. Using single-cell RNA sequencing and flow cytometry in BRAFi + MEKi-treated melanoma, we show herein that isogenic Gsdme knockout (KO) tumors show decreased infiltration with T cells, natural killer (NK) cells, and regulatory T cells (Treg) compared with control tumors. Infiltrated Tregs in Gsdme KO tumors displayed decreased expression of the IL2 receptor and phenotypic markers associated with suppressive function. Furthermore, intratumoral frequency of phenotypically suppressive Tregs was decreased after BRAFi + MEKi treatment in Gsdme KO tumors engineered to express a pyroptosis-defective mutant form of Gsdme (T6E) compared with Gsdme KO tumors engineered to reexpress wild-type Gsdme. Combining BRAFi + MEKi with a TLR9 agonist limited the regrowth of Gsdme-deficient tumors, and this was associated with a further reduction in intratumoral Tregs. Overall, we show a critical role of GSDME in the modulation of intratumoral immune cells in BRAFi + MEKi-treated melanoma.