Tumor-derived ILT3 inhibition to reshape the immunosuppressive microenvironment and potentiate the PD-L1 blockade immunotherapy in lung adenocarcinoma.

Abstract

e14647 Background: The low response of immune checkpoint inhibitors against PD-1/PD-L1 in cancer patients calls for further development of new immune targets. Immunoglobulin-like transcript (ILT)3 and its ligand ApoE have been demonstrated to be highly expressed in a variety of malignant tumor cells and involved in tumor biological behaviors, including invasion and angiogenesis. ILT3 targeting drugs are undergoing increasing preclinical and clinical trials. However, the role of tumor-derived ILT3 in immune regulation is unclear. The aim of this study was to investigate the immunomodulatory effect of tumor-derived ILT3 and its immunotherapy therapeutic efficacy that combination with PD-L1 blockade in NSCLC. Methods: The correlations between ILT3/ApoE expression and prognosis of NSCLC patients were analyzed basing on public databases. Multiplex immunofluorescence was performed to analyze the correlations between tumor-derived ILT3/ApoE expression and intratumoral immune cells infiltration in human adenocarcinoma tissues. The impact of ILT3 and ApoE on tumor-associated macrophage (TAM) recruitment and polarization were evaluated by transwell migration assay, flow cytometry, and RT-PCR, while their direct impact on T cell survival and cytotoxicity was detected by flow cytometry. Using lung cancer xenograft murine models, we further confirmed the pro-tumoural activity of tumor-derived gp49B and analyzed its correlations with immune cells in vivo. Tumor immunotherapy models targeting at gp49B and/or PD-L1 were established in C57BL/6 mice inoculated with LLC cells. Gp49b inhibition was implemented by infection of specific knockdown lentivirus and PD-L1 was blocked using mouse neutralizing antibodies. Flow cytometry was performed to detect the frequency and phenotype of macrophages and T cells in the tumors, spleen, and blood of the mice. Results: Patients with high expression of ILT3 had significantly worse prognosis in LUAD, but not in LUSC. In vitro assays demonstrated that tumor-derived ILT3/ApoE promoted recruitment and M2-like polarization of TAMs and directly inhibited the proliferation and cytotoxicity of T cells, which had been further confirmed in vivo models. Moreover, ILT3 (gp49B) inhibition enhanced anti-tumor immunity and suppressed tumor progression by counteracting TAM- and dysfunctional T cell- induced immunosuppressive TME; the combined inhibition of gp49B and PD-L1 showed the most dramatic tumor retraction in LLC xenograft models. Conclusions: Tumor-derived ILT3 overexpression suppressed anti-tumor immunity by recruiting M2-like TAMs and directly impairing T cell activities, while ILT3 inhibition counteracted above immunosuppression and potentiated the efficacy of PD-L1 blockade in LUAD. Our study verified that ILT3 may be a promising novel immunotherapeutic target for LUAD patients.