Experimental Study Of Cryoablation Combined With PD-1 And CTLA-4 Inhibitors In The Treatment Of Hepatocellular Carcinoma Model Mice

Background:Cryoablation(Cryo)is a minimally invasive treatment technique for solid tumors,and clinical trials have confirmed that Cryo is safe and effective in the treatment of hepatocellular carcinoma(HCC).Strategies for physical ablation of tumors,including Cryo,can activate the body to generate a T-cell-mediated antitumor immune response as an alternative way to kill tumor cells.However,this immune effect is usually weak,and the immune response induced by Cryo in HCC is poorly understood.Both PD-1 and CTLA-4 monoclonal antibodys(mAbs)are immune checkpoint inhibitors that are clinically used in the immunotherapy of malignant tumors and can enhance the specific antitumor immunity produced by the body.Therefore,the combined application of PD-1 and CTLA-4 mAbs may enhance the immune effect of Cryo.Methods:A Balb/c mouse model of HCC was established and treated with Cryo,immune checkpoint blockade(ICB),or Cryo combined with ICB.The growth trend of contralateral tumors and survival time of mice in each group were determined.The expression of apoptosis-related proteins was detected by WB assay.The percentages of immune cells and immunosuppressive cells in contralateral tumor tissues were analyzed by flow cytometry.The numbers of infiltrating T lymphocytes in contralateral tumor tissues were checked by immunohistochemistry,the levels of T-cell-associated cytokines was detected by qRT-PCR assays and ELISA assays.Results:Cryo combined with ICB inhibited the growth of contralateral tumors,increased the expression of apoptosis related proteins,and prolonged the survival time of mice.Local T-cell infiltration in distant tumor tissues increased after the combination therapy,while the number of immunosuppressive cells was significantly reduced.In addition,the combination therapy may induce the production of multiple Thl-type cytokines but reduce the production of Th2-type cytokines.We also verified the antiangiogenic effect of the combination therapy.Conclusions:Cryo can activate certain T-cell immune responses.Cryo combined with ICB can improve the immunosuppressive tumor microenvironment and shift the Thl/Th2 balance towards Thl dominance,further enhancing the Cryo-induced T-cell immune response and resulting in a stronger antitumor immune response.Therefore.Cryo combined with ICB may be an effective strategy for immunotherapy in HCC.