(A) Schematics of treatment design using the CRPC GEMM model. PbPPS mice were surgically castrated, followed by treatment with enzalutamide (Enza) for three weeks. Tumor relapse was monitored by 7T-MRI. Mice were then treated with Enza in combination with B7-H3 inhibitor (MJ18, 300 μg/injection) for four weeks. Isotype IgG was used in the control group. (B,C,D) Fold changes in PbPPS CRPC tumor volume (B), representative MRI images (C), and histopathology analysis (D) after treatment. Prostate tumor regions are circled with red lines. n = 4 in Enza Control group and n = 3 in Enza+anti-B7-H3 group. Scale bars = 2mm. (E,F) Immunoprofiling of syngeneic tumors treated with Enza in combination with IgG or anti-B7-H3 using CyTOF. Colored viSNE plots of CD45+ immune cells and expression patterns of PD-1 and PD-L1 are shown (E). Quantification of tumor-infiltrating lymphocytes is presented (F). TIL: Tumor Infiltrating Leukocytes. (G) Expression of PD-L1 (Median Metal Intensity, MMI) in cancer cell and myeloid components after treatment. Data represent the mean ± SD. Student’s t-tests were performed using GraphPad Prism version 9.2.0. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. ns, not significant.