Tumor-infiltrating CD36+CD8+T cells determine exhausted tumor microenvironment and correlate with inferior response to chemotherapy in non-small cell lung cancer.

BACKGROUND: The scavenger receptor CD36 was reported to be highly expressed on tumor-infiltrating CD8+ T cells, but the clinical role remains obscure. This study aims to explore the infiltration and clinical value of CD36+CD8+ T cells in NSCLC. METHODS: Immunohistochemistry and immunofluorescence were conducted for survival analyses and immunological evaluation in 232 NSCLC patients in Zhongshan Hospital. Flow cytometry analyses were carried out to assess the immune cells from fresh tumor samples, non-tumor tissues and peripheral blood. In vitro tumor infiltrating lymphocytes cultures were conducted to test the effect of CD36 blockage. RESULTS: Accumulation of CD36+CD8+ T cells in tumor tissues was correlated with more advanced stage (p < 0.001), larger tumor size (p < 0.01), and lymph node metastasis (p < 0.0001) in NSCLC. Moreover, high infiltration of CD36+CD8+ T cells indicated poor prognosis in terms of both overall survival (OS) and recurrence-free survival (RFS) and inferior chemotherapy response. CD36+CD8+ T cells showed decreased GZMB (p < 0.0001) and IFN-γ (p < 0.001) with elevated PD-1 (p < 0.0001) and TIGIT (p < 0.0001). Analysis of tumor-infiltrating immune cell landscape revealed a positive correlation between CD36+CD8+ T cells and Tregs (p < 0.01) and M2-polarized macrophages (p < 0.01) but a negative correlation with Th1 (p < 0.05). Notably, inhibition of CD36 partially restored the cytotoxic function of CD8+ T cells by producing more GZMB and IFN-γ. CONCLUSION: CD36+CD8+ T cells exhibit impaired immune function and high infiltration of CD36+CD8+ T cells indicated poor prognosis and inferior chemotherapy response in NSCLC patients. CD36 could be a therapeutic target in combination with chemotherapy in NSCLC patients.

TTI1 promotes non-small-cell lung cancer progression by regulating the mTOR signaling pathway.

The role of TELO2-interacting protein 1 (TTI1) in the progression of several types of cancer has been reported recently. The aim of this study was to estimate the expression and potential value of TTI1 in non-small-cell lung cancer (NSCLC) patients. The expression of TTI1 and its prognostic value in NSCLC from The Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database were analyzed. To verify the bioinformatics findings, a tissue microarray containing 160 NSCLC and paired peritumoral tissues from NSCLC patients was analyzed by immunohistochemistry for TTI1. Subsequently, the roles of TTI1 in NSCLC cells were investigated in vivo by establishing xenograft models in nude mice and in vitro by transwell, CCK-8, wound healing, and colony formation assays. In addition, quantitative real-time polymerase chain reaction and western blot were applied to explore the underlying mechanism by which TTI1 promotes tumor progression. Finally, the relationship between TTI1 and Ki67 expression level in NSCLC was probed, and Kaplan-Meier and Cox analyses were performed to assess the prognostic merit of TTI1 and Ki67 in NSCLC patients. We found that the expression of TTI1 was significantly upregulated in NSCLC tissues compared to paired peritumoral tissues, which coincides with the bioinformatics findings from the TCGA and GEO databases. TTI1 was highly expressed in NSCLC patients with large tumors, advanced tumor stage, and lymphatic metastasis. In addition, the prognostic analysis identified TTI1 as an independent indication for poor prognosis of NSCLC patients. In vitro, upregulation of TTI1 in NSCLC cells could facilitate cell invasion, metastasis, viability, and proliferation. Mechanistically, our study verified that TTI1 could regulate mTOR activity, which has a pivotal role in human cancer. Consistently, the expressions of TTI1 and Ki67 had a positive relationship in NSCLC cells and tissues. Notably, patients with overexpression of TTI1 or Ki67 had a shorter overall survival rate and a higher disease-free survival rate compared to patients with low expression of TTI1 or Ki67, and the combination of TTI1 and Ki67 was an independent parameter predicting the prognosis and recurrence of NSCLC patients. We conclude that TTI1 promotes NSCLC cell proliferation, metastasis, and invasion by regulating mTOR activity, and the combination of TTI1 and Ki67 is a valuable molecular biomarker for the survival and recurrence of NSCLC patients.

Exosomal circZNF451 restrains anti-PD1 treatment in lung adenocarcinoma via polarizing macrophages by complexing with TRIM56 and FXR1.

BACKGROUND: Although success was achieved in the therapy for a minority of advanced lung adenocarcinoma (LUAD) patients, anti-programmed death 1 (PD1) resistance was found in most LUAD patients. Here, we aimed to uncover a potential role of exosomal circular RNAs (circRNAs) in LUAD refractory to PD1 blockade.  METHODS: circRNA sequencing and qRT-PCR were performed to determine the level of exosomal circRNAs in LUAD patients subsequently treated with anti-PD1. Then, the RNA pulldown, RNA immunoprecipitation, mass spectrometry, chromatin immunoprecipitation, luciferase reporter assays, flow cytometry, RNA sequencing, and in vitro and in vivo models were used to uncover the biological functions and underlying mechanism of circZNF451 in LUAD anti-PD1 treatment resistance. RESULTS: circRNA sequencing and qRT-PCR identified the up-regulation of exosomal circZNF451 from LUAD patients with progressive disease (PD) compared to those with partial remission (PR) after PD1 blockade therapy. Furthermore, elevated circZNF451 was revealed to be associated with poor prognosis of LUAD patients. Additionally, exosomal circZNF451 was demonstrated to induce an anti-inflammatory phenotype in macrophages and exhaustion of cytotoxic CD8+ T cells, and enhanced TRIM56-mediated degradation of FXR1 to activate the ELF4-IRF4 pathway in macrophages. By transgenic mice, knockout of ELF4 in macrophages was found to rescue immunotherapy efficacy in tumors with high level of exosomal circZNF451. CONCLUSION: Exosomal circZNF451 reshapes the tumor immune microenvironment by inducing macrophages polarization via the FXR1- ELF4-IRF4 axis and is a novel biomarker for predicting the sensitivity of PD1 blockade in LUAD.

Elevated circASCC3 limits antitumor immunity by sponging miR-432-5p to upregulate C5a in non-small cell lung cancer.

Although anti-programmed cell death 1 (PD1) treatment has become a first-line therapy for advanced non-small cell lung cancer (NSCLC), most NSCLC patients are refractory to anti-PD1. Here, we aimed to investigate the mechanism of dysregulated circular RNAs (circRNAs) related to anti-PD1 resistance in NSCLC. The expression of circASCC3 (hsa_circ_0077,495) in NSCLC tissues and cell lines was evaluated by fluorescence in situ hybridization and quantitative reverse transcription-polymerase chain reaction. The functions and mechanisms of circASCC3 in NSCLC progression and anti-PD1 resistance were uncovered in vitro and in vivo. The circASCC3 level was upregulated in NSCLC compared with that in paired normal tissues. Specifically, circASCC3 expression was higher in tissues from NSCLC patients with anti-PD1 refractory than in those from patients who sensitive to anti-PD1. Overexpression of circASCC3 enhanced the malignant phenotype of NSCLC cells and led to an immunosuppressive microenvironment. Mechanistically, circASCC3 sponged miR-432-5p to increase complement C5a levels, which enhanced the progression and dysfunctional immune status of NSCLC. Thus, circASCC3 overexpression reshapes the tumor microenvironment by impacting the complement system in NSCLC and provides a potential strategy to overcome anti-PD1 resistance.

The circular RNA circHMGB2 drives immunosuppression and anti-PD-1 resistance in lung adenocarcinomas and squamous cell carcinomas via the miR-181a-5p/CARM1 axis.

BACKGROUND: Previous studies have confirmed the oncogenic role of HMGB2 in various cancers, but the biological functions of HMGB2-derived circRNAs remain unknown. Thus, we intended to investigate the potential role of HMGB2-derived circRNAs in lung adenocarcinomas (LUAD) and squamous cell carcinomas (LUSC). METHODS: The expression profiles of HMGB2-derived circRNAs in LUAD and LUSC tissues and matched normal tissues were assessed using qRT-PCR. The role of circHMGB2 in the progression of the LUAD and LUSC was determined in vitro by Transwell, CCK-8, flow cytometry and immunohistochemistry assays, as well as in vivo in an immunocompetent mouse model and a humanized mouse model. In addition, in vivo circRNA precipitation assays, luciferase reporter assays and RNA pulldown assays were performed to explore the underlying mechanism by which circHMGB2 promotes anti-PD-1 resistance in the LUAD and LUSC. RESULTS: The expression of circHMGB2 (hsa_circ_0071452) was significantly upregulated in NSCLC tissues, and survival analysis identified circHMGB2 as an independent indicator of poor prognosis in the LUAD and LUSC patients. We found that circHMGB2 exerted a mild effect on the proliferation of the LUAD and LUSC cells, but circHMGB2 substantially reshaped the tumor microenvironment by contributing to the exhaustion of antitumor immunity in an immunocompetent mouse model and a humanized mouse model. Mechanistically, circHMGB2 relieves the inhibition of downstream CARM1 by sponging miR-181a-5p, thus inactivating the type 1 interferon response in the LUAD and LUSC. Moreover, we found that the upregulation of circHMGB2 expression decreased the efficacy of anti-PD-1 therapy, and we revealed that the combination of the CARM1 inhibitor EZM2302 and an anti-PD-1 antibody exerted promising synergistic effects in a preclinical model. CONCLUSION: circHMGB2 overexpression promotes the LUAD and LUSC progression mainly by reshaping the tumor microenvironment and regulating anti-PD-1 resistance in the LUAD and LUSC patients. This study provides a new strategy for the LUAD and LUSC treatment.