Targeting PID1 generates oxysterols to switch macrophage cell fates for improved antitumor immunity.

Disordered cholesterol-oxysterol profiles are observed in the tumor microenvironment, yet their roles in tumor-associated macrophages (TAMs) remain underexplored. This study reveals that TAMs across human pan-cancers exhibit elevated phosphotyrosine interaction domain-containing protein 1 (PID1) expression and prominent immunosuppressive gene signatures. PID1 deficiency in myeloid cells upregulates low-density lipoprotein (LDL) receptor expression, thereby promoting LDL uptake and intracellular accumulation of free cholesterol and reactive oxygen species (ROS). Increased ROS drives cholesterol oxidation to generate the oxysterols 5α,6α-epoxycholesterol (5α,6α-EC) and 7β-hydroxycholesterol (7β-OHC), which inhibit mTOR-STAT6 signaling in macrophages. Pid1 deletion switches immunosuppressive macrophages toward an antitumor subtype that downregulates arginase 1 expression while upregulating proinflammatory cytokines, thereby potentiating CD8+ T cell-mediated immunosurveillance across multiple tumor types. Moreover, combination treatment with the oxysterol and chemotherapeutic agent 5-fluorouracil produces synergistically enhanced antitumor effects. Previous studies showed that cholesterol-derived oxysterols differentially regulate macrophage cell fates, with 25-OHC promoting the protumor and immunosuppressive phenotype of TAMs. Targeting PID1 reroutes cholesterol and ROS metabolism toward the production of 5α,6α-EC and 7β-OHC in TAMs, representing a promising immunometabolic strategy to restore antitumor immunosurveillance.