Tumor-derived lactate inhibit the efficacy of lenvatinib through regulating PD-L1 expression on neutrophil in hepatocellular carcinoma

Background Neutrophils play a controversial role in tumor development. The function of programmed cell death-1 ligand (PD-L1(+)) neutrophils, however, may inhibit the cytotoxicity of anti-tumor immunity. In this study, we elucidate the stimulators of PD-L1(+) neutrophils in tumor microenvironment (TME) and explore the optimal combination to enhance the effect of lenvatinib by inhibiting PD-L1(+) neutrophils in hepatocellular carcinoma. Methods Neutrophil infiltration after lenvatinib treatment was examined with RNA sequencing and multicolor flow cytometry analysis in patient samples, subcutaneous and orthotopic mouse models. Neutrophils and T cells were isolated from peripheral blood and tumor tissues and purified with magnetic beads for cytotoxicity assay. Metabolites and cytokines were detected by a biochemical analyzer manufactured by Yellow Springs Instrument (YSI) and proteome profiler cytokines array. In vitro screening of pathway inhibitors was used to identify possible candidates that could reduce PD-L1(+) neutrophil infiltration. Further in vivo assays were used for verification. Results Lenvatinib increased neutrophil recruitment by inducing CXCL2 and CXCL5 secretion in TME. After entering TME, neutrophils polarized toward N2 phenotype. PD-L1 expression was simultaneously upregulated. Thus, lenvatinib efficacy on tumor cells hindered. The increasing PD-L1(+) neutrophils positively corelated with a suppressive T cell phenotype. Further investigation indicated that JAK/STAT1 pathway activated by immune-cell-derived interferon gamma and MCT1/NF-kB/COX-2 pathway activated by high concentrations of tumor-derived lactate could induce PD-L1(+) neutrophils. The latter could be significantly inhibited by COX-2 inhibitor celecoxib. Further in vivo assays verified that Celecoxib decreased the survival of lactate-stimulated PD-L1(+) neutrophil and promoted the antitumor effect of lenvatinib. Conclusions PD-L1(+) neutrophils decrease T cell cytotoxicity. Tumor-derived lactate induces PD-L1 expression on neutrophils via MCT1/NF-kappa B/COX-2 pathway. Thus, COX-2 inhibitor could reduce PD-L1(+) neutrophil and restore T cell cytotoxicity. This may provide a potent addition to lenvatinib.

Automated summary

Neutrophils play a controversial role in tumor development, with PD-L1(+) neutrophils inhibiting the cytotoxicity of anti-tumor immunity. Lenvatinib was found to induce CXCL2 and CXCL5 secretion, promoting recruitment of neutrophils in the tumor microenvironment. Inhibiting PD-L1(+) neutrophils with a COX-2 inhibitor could enhance the antitumor effect of lenvatinib by restoring T cell cytotoxicity.