<p>Gasdermin D (GSDMD), an effector molecule of cell pyroptosis, is known to be activated in various cells during inflammation. However, the patterns of GSDMD activation in immune regulatory cells such as myeloid-derived suppressor cells (MDSCs) remain unclear. In this study, we found that neutrophils in colorectal cancer (CRC) tissues exhibited reduced GSDMD transcription, as evidenced by a single-cell RNA sequencing result. Consistent with this, cleaved GSDMD expression is negatively correlated with S100A8 in CRC tissues. Additionally, CD15⁺CD14⁻LOX1⁺ cells (G-MDSCs) from the peripheral blood of CRC patients exhibited a significant reduction in GSDMD activation. Mice with ubiquitous GSDMD deficiency bred in a clean environment exhibited a notable increase in G-MDSCs. These GSDMD^−/− MDSCs enhanced immunosuppressive activity by both inhibiting effector T-cell activity and promoting regulatory T-cell induction. This enhancement was also observed in GSDMD^flox/flox-S100A8^Cre mice, in which GSDMD was specifically deleted in MDSCs. The tumor-promoting effects in the GSDMD^−/− and GSDMD^flox/flox-S100A8^Cre mice were abrogated following MDSC depletion, as shown by the use of an anti-DR5 antibody. In the absence of GSDMD, G-MDSCs showed reduced inflammasome activation and decreased production of IL-1β and IL-18. Furthermore, a significant reduction in interferon-related factor 8/7 (IRF8/7) was observed in GSDMD^−/− G-MDSCs via bulk RNA sequencing analysis. After treatment with LPS/nigericin, these cells maintained mitochondrial integrity, thus impairing the mtDNA release and the downstream cGAS/STING/TBK1/IRF8/7 signaling axis activation. Reduced IRF8/7 levels were responsible for increased differentiation of GSDMD^−/− G-MDSCs. Finally, treatment with a GSDMD recombinant lentivirus injected into in situ tumors significantly inhibited tumor growth and reduced G-MDSC levels, suggesting that a GSDMD-based vaccine could simultaneously exert anti-carcinoma and anti-MDSC effects.</p>