Immunomodulatory Effect And Its Mechanism Of The Combination Of Microwave Ablation And OK-432 On Breast Cancer

Backgrounds:Minimally invasive therapies,such as microwave ablation(MWA),are widely used for the treatment of solid tumors.Previous studies suggest that MWA is feasible for the treatment of small breast cancer,and thermal ablation may induce antitumor T-cell response.However,the induced immune responses are mostly weak,and the immunomodulation effects of MWA in breast cancer are unclear.Immunostimulant OK-432 was approved as an imnunotherapeutic agent in cancers.Therefore,OK-432 may augment the immunity induced by MWA.Objective:In this study,we evaluated the local and systemic changes in T lymphocyte and its subtypes,the specificity of immune response,the cytokine concentrations,and survival of breast cancer mouse models after MWA and MWA plus OK-432.Materials and methods:We treated 4T1 breast cancer bearing BALB/c mice with MWA,OK-432,MWA plus OK-432,or left without treatment.Survival time was evaluated with the Kaplan?Meyer method comparing survival curves by log-rank test.On day 25 after ablation,surviving mice received tumor rechallenge,and the rechallenged tumor volumes were calculated.Flow cytometry were used to evaluate the proportions of T-cell and its subtypes in spleens.And iununohistochemistry was used to examine the CD4+and CD8+T cell inflltrating into tumor issues.The tumor-specific immunity was assessed by enzyme-linked inununospot assays.Besides,the cytokine patterns were identified from enzyme-linked immunosorbent assay.The significance of differences between four experimental groups was assessed by statistical analysis.Results:Survival prolongation was observed in MWA treated mice,when compared with mice in OK-432 group and untreated group(P?0.006 and 0.001,respectively).The eombination therapy led to longer survival than that of other groups(all P<0.001)and complete rejection of the second tumor in 83%mice in re-challenge test.Moreover,the number of intratumoral CD4+and CD8+T cells was significantly higher in MWA alone than in control group(both P<0.001,Fig.2B).the combination of MWA and OK-432 further enhanced CD8+T-cell infiltration into tumors compared with tumors treated with single treatments(both P<0.001).Besides,the percentage of splenic CD4+and CD8+T cells of mice treated with MWA alone were significantly increased when compared with those of untreated mice(P?0.004 and<0.001,respectively).Subsequent administration of OK-432 after MWA further increased the levels of CD4+and CD8+T cells in the spleen as compared with those of MWA ouly treated mice(P=0.011 and?0.004,respectively).The combination treatment group also had higher number of 4T1 specific IFN-? secreting cells when compared with MWA alone or OK-432 alone treatment groups(P=0.031,<0.001 and<0.001,respectively).In addition,The levels of T helper type1(Th1)-type cytokines IL-2,IL-12 and IL-18 in MWA plus OK-432 group were significantly higher than those in MWA group(P=0.027,0.008,and 0.007,respectively).While plasma level of T helper type 2(Th2)-type cytokines IL-4 and IL-10 were not significantly affected by these treatments.Finally,MWA plus OK-432 treated mice had higher percentage of IFN-? producing Thl cells in the spleen when compared with MWA or OK-432 alone treated mice(P?0.004 and<0.001,respectively)and lower percentage of Th2 cells when compared with untreated mice(P=0.05).Therefore,the combination treatment group showed markedly increased Thl to Th2 ratio as comparing with MWA or OK-432 treatment alone groups(P=0.022,<0.001 and<0.001,respectively).Conclusions:T-cell responses was induced by microwave ablation in a murine breast cancer model,and the combination of microwave ablation and immunostimulant OK-432 synergistically polarized the T-cell responses to Thl dominance and augmented specific antitumor immunity.Thus,the combination of microwave ablation and OK-432 might be a promising treatment option for breast cancer.