PD-1 blockade therapy has made great breakthroughs in treatment of multiple solid tumors. However, patients with microsatellite-stable (MSS) colorectal cancer (CRC) respond poorly to anti-PD-1 immunotherapy. Although CRC patients with microstatellite instability (MSI) or microsatellite instability-high (MSI-H) can benefit from PD-1 blockade therapy, there are still some problems such as tumor recurrence. Tumor-associated macrophages (TAMs), most abundant immune components in tumor microenvironment (TME), largely limit the therapeutic efficacy of anti-PD-1 against CRC. The CSF1/CSF1R pathway plays a key role in regulating macrophage polarization, and blocking CSF1R signaling transduction may be a potential strategy to effectively reprogram macrophages and remodel TME. Here, we found that increasing expression of CSF1R in macrophages predicted poor prognosis in CRC cohort. Furthermore, we discovered a novel potent CSF1R inhibitor, PXB17, which significantly reprogramed M2 macrophages to M1 phenotype. Mechanically, PXB17 significantly blocked activation of PI3K/AKT/mTORC1 signaling, resulting in inhibition of cholesterol biosynthesis. Results from 3D co-culture system suggested that PXB17-repolarized macrophages could induce infiltration of CD8⁺ T lymphocytes in tumors and improve the immunosuppressive microenvironment. In vivo, PXB17 significantly halted CRC growth, with a stronger effect than PLX3397. In particular, PXB17 potently enhanced therapeutic activity of PD-1 mAb in CT-26 (MSS) model and prevented tumor recurrence in MC-38 (MSI-H) model by promoting formation of long-term memory immunity. Our study opens a new avenue for CSF1R in tumor innate and adaptive anti-tumor immunomodulatory activity and suggests that PXB17 is a promising immunotherapy molecule for enhancing the efficacy of PD-1 mAb or reducing tumor recurrence of CRC.