PD-1/PD-L1 inhibition after cryoablation is associated with improved tumor control and immunosuppressive microenvironment remodeling in Lewis lung adenocarcinoma.

Cryoablation (Cryo) is used for local tumor control in lung cancer patients who are not candidates for surgical resection, but its effects on distant tumor growth remain incompletely understood. The present study evaluated the effects of Cryo on distant Lewis lung tumors, explored whether Cryo-induced PD-1/PD-L1 axis upregulation may represent a treatment-limiting pathway, and assessed whether PD-1/PD-L1 inhibition in Cryo-treated mice was associated with remodeling of the immunosuppressive tumor microenvironment and reduced tumor proliferation. In the experiments, mice bearing bilateral Lewis tumors were assigned to Untreated, Cryo, In-PD-1, and Cryo + In-PD-1 groups. Immunofluorescence was used to assess PD-L1 and Ki-67 expression. Quantitative PCR, enzyme-linked immunosorbent assay, and flow cytometry were used to evaluate immunosuppressive cytokines and immune-cell populations, while western blotting was performed to analyze signaling-related proteins. Results showed that Cryo mildly inhibited distant tumor growth, increased tumor-infiltrating T-cell levels, and upregulated the PD-1/PD-L1 axis. In-PD-1 was associated with prolonged survival and further inhibition of distant tumor growth in Cryo-treated tumor-bearing mice. The Cryo + In-PD-1 group showed reduced regulatory T cells, M2-type tumor-associated macrophages, myeloid-derived suppressor cells, and the immunosuppressive cytokines interleukin-6 and VEGF, together with decreased Ki-67 expression and p-ERK/ERK ratio in tumor tissues. These findings suggest that Cryo-induced upregulation of the PD-1/PD-L1 axis may be associated with limited efficacy of Cryo monotherapy, and that PD-1/PD-L1 inhibition in Cryo-treated mice may contribute to broader remodeling of the immunosuppressive tumor microenvironment and improved tumor control, accompanied by a less proliferative tumor phenotype and decreased ERK pathway activation.