Anlotinib Combined with Programmed Death-1/Programmed Death-Ligand 1 Inhibitor 9 Hydrochloride Suppresses Colorectal Cancer Progression by Inducing Neovascularization and Reprogramming the Tumor Immune Microenvironment in MSS Murine Model.

Background: Anlotinib, a novel multi-target tyrosine kinase inhibitor, has demonstrated promising antitumor efficacy by inhibiting angiogenesis. However, the potential therapeutic benefits and underlying mechanisms of combining anlotinib with immune checkpoint inhibitors in colorectal cancer remain unclear. Methods: Syngeneic models of colorectal cancer were established and treated with anlotinib, PD-1/PD-L1-IN-9 hydrochloride, or their combination. Tumor tissues were analyzed using immunohistochemistry, and Western blotting to assess angiogenesis-associated markers, tissue hypoxia, and key molecular markers. Flow cytometry, ELISA, and immunostaining were performed to evaluate immune cell infiltration, cytokine expression, and the tumor immune microenvironment. Results: The combination of anlotinib and PD-1/PD-L1-IN-9 hydrochloride significantly inhibited tumor growth compared to monotherapy, associated with improved neovascularization and alleviated hypoxia. Combined therapy increased CD8+ T cell infiltration, reduced immunosuppressive cell populations, and partially modulated cytokine profiles, thereby enhancing the antitumor immune response. Conclusion: Anlotinib combined with PD-1/PD-L1-IN-9 hydrochloride exhibits superior antitumor efficacy in colorectal cancer compared to either agent alone, potentially by reshaping the tumor microenvironment. These findings support further exploration of combined anti-angiogenic and immune checkpoint therapy as a promising strategy for colorectal cancer treatment.