Neoadjuvant PD-1 Inhibition prior to Partial Cryoablation of Murine Hepatocellular Carcinoma Modulates the Tumor Microenvironment toward Favorable Immunological Profiles.

PURPOSE: To evaluate the impact of neoadjuvant systemic PD-1 immune checkpoint inhibition on the local immune response in residual tumors following partial cryoablation in a TIB-75 murine hepatocellular carcinoma (HCC) model. MATERIALS AND METHODS: Forty-eight BALB/c mice (6-12 weeks) were orthotopically implanted with TIB-75 cells to induce a single lesion of HCC. Mice were randomized into 4 treatment groups: (a) control, (b) anti-PD-1, (c) partial cryoablation, and (d) anti-PD-1 and partial cryoablation. Anti-PD-1 was administered on Days 7, 9, and 11 after inoculation, followed by partial cryoablation on Day 13 and tumor harvest on Day 18. The presence of T cell subsets (CD3(+), CD4(+), and CD8(+)), tumor-associated macrophages (CD68(+) and CD206(+)), PD-1, and PD-L1 were assessed by histopathological analysis of immunohistochemistry. The percentage of positively stained cells within the tumor was determined using QuPath. RESULTS: Mice treated with anti-PD-1 (n = 12) had greater infiltration of CD3(+), CD4(+), and CD8(+) T cells into residual tumors than control (CD3(+): median, 22.4% vs 5.5% [P < .001]; CD4(+): median, 19.8% vs 5.1% [P < .001]; CD8(+): median, 8.2% vs 3.1% [P = .007]). Partial cryoablation alone (n = 12) increased CD206(+) M2-like macrophages (median, 36.6% vs 14.7%; P = .03). Partial cryoablation combined with neoadjuvant anti-PD-1 (n = 12) showed significantly higher infiltration of CD3(+) T cells (median, 14.3% vs 4.5%; P = .048) than partial cryoablation alone (n = 12) and significantly lower PD-1 expression than anti-PD-1 alone (median, 2.9% vs 7.3%; P = .004). CONCLUSIONS: In a mouse model of HCC, neoadjuvant PD-1 immune checkpoint inhibition can modulate the immunosuppressive tumor microenvironment observed after cryoablation. This highlights the potential of a combination therapy to treat both early- and advanced-stage HCCs.