Cancer cell-derived exosomal circUSP7 induces CD8(+) T cell dysfunction and anti-PD1 resistance by regulating the miR-934/SHP2 axis in NSCLC.

CD8+ T cells play a critical role in the innate antitumour immune response. Recently, CD8+ T cell dysfunction has been verified in various malignant cancers, including non-small cell lung cancer (NSCLC). However, the molecular biological mechanisms of CD8+ T cell dysfunction in human NSCLC are still unclear. The expression of circular ubiquitin-specific protease-7 (circUSP7) in NSCLC tissues, exosomes, and cell lines was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Exosomes were isolated from the culture medium of NSCLC cells and the plasma of NSCLC patients using an ultracentrifugation method and the ExoQuick Exosome Precipitation Solution kit. The exosomes were then characterized by transmission electronic microscopy (TEM), NanoSight and western blotting. The role of circUSP7 in CD8+ T cell dysfunction was assessed by enzyme-linked immunosorbent assay (ELISA). In vivo circular RNA (circRNA) precipitation (circRIP), RNA immunoprecipitation (RIP), and luciferase reporter assays were performed to explore the molecular mechanisms of circUSP7 in CD8+ T cells. In a retrospective study, the clinical characteristics and prognostic significance of circUSP7 in NSCLC tissues were determined. The expression levels of circUSP7 were higher in human NSCLC tissues than in matched adjacent nontumour tissues. Increased levels of circUSP7 indicate poor clinical prognosis and CD8+ T cell dysfunction in patients with NSCLC. The circUSP7 found in NSCLC patient plasma is predominantly secreted by NSCLC cells in an exosomal manner, and circUSP7 inhibits IFN-γ, TNF-α, Granzyme-B and Perforin secretion by CD8+ T cells. Furthermore, circUSP7 inhibits CD8+ T cell function by upregulating the expression of Src homology region 2 (SH2)-containing protein tyrosine phosphatase 2 (SHP2) via sponging miR-934. Finally, we show that circUSP7 may promote resistance to anti-PD1 immunotherapy in NSCLC patients. Exosomal circUSP7 is predominantly secreted by NSCLC cells and contributes to immunosuppression by promoting CD8+ T cell dysfunction in NSCLC. CircUSP7 induces resistance to anti-PD1 immunotherapy, providing a potential therapeutic strategy for NSCLC patients.

Chen SW ，Zhu SQ ，Pei X ，Qiu BQ ，Xiong D ，Long X ，Lin K ，Lu F ，Xu JJ ，Wu YB ... -  《Molecular Cancer》

Cancer cell-derived exosomal circUHRF1 induces natural killer cell exhaustion and may cause resistance to anti-PD1 therapy in hepatocellular carcinoma.

Zhang PF ，Gao C ，Huang XY ，Lu JC ，Guo XJ ，Shi GM ，Cai JB ，Ke AW ... -  《Molecular Cancer》

Exosome-derived circCCAR1 promotes CD8 + T-cell dysfunction and anti-PD1 resistance in hepatocellular carcinoma.

Circular RNAs (circRNAs) can be encapsulated into exosomes to participate in intercellular communication, affecting the malignant progression of a variety of tumors. Dysfunction of CD8 + T cells is the main factor in immune escape from hepatocellular carcinoma (HCC). Nevertheless, the effect of exosome-derived circRNAs on CD8 + T-cell dysfunction needs further exploration. The effect of circCCAR1 on the tumorigenesis and metastasis of HCC was assessed by in vitro and in vivo functional experiments. The function of circCCAR1 in CD8 + T-cell dysfunction was measured by enzyme-linked immunosorbent assay (ELISA), western blotting and flow cytometry. Chromatin immunoprecipitation, biotinylated RNA pull-down, RNA immunoprecipitation, and MS2 pull-down assays were used to the exploration of mechanism. A mouse model with reconstituted human immune system components (huNSG mice) was constructed to explore the role of exosomal circCCAR1 in the resistance to anti-PD1 therapy in HCC. Increased circCCAR1 levels existed in tumor tissues and exosomes in the plasma of HCC patients, in the culture supernatant and HCC cells. CircCCAR1 accelerated the growth and metastasis of HCC in vitro and in vivo. E1A binding protein p300 (EP300) and eukaryotic translation initiation factor 4A3 (EIF4A3) promoted the biogenesis of circCCAR1, and Wilms tumor 1-associated protein (WTAP)-mediated m6A modification enhanced circCCAR1 stability by binding insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3). CircCCAR1 acted as a sponge for miR-127-5p to upregulate its target WTAP and a feedback loop comprising circCCAR1/miR-127-5p/WTAP axis was formed. CircCCAR1 is secreted by HCC cells in a heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2B1)-dependent manner. Exosomal circCCAR1 was taken in by CD8 + T cells and caused dysfunction of CD8 + T cells by stabilizing the PD-1 protein. CircCCAR1 promoted resistance to anti-PD1 immunotherapy. Furthermore, increased cell division cycle and apoptosis regulator 1 (CCAR1) induced by EP300 promoted the binding of CCAR1 and β-catenin protein, which further enhanced the transcription of PD-L1. The circCCAR1/miR-127-5p/WTAP feedback loop enhances the growth and metastasis of HCC. Exosomal circCCAR1 released by HCC cells contributes to immunosuppression by facilitating CD8 + T-cell dysfunction in HCC. CircCCAR1 induces resistance to anti-PD1 immunotherapy, providing a potential therapeutic strategy for HCC patients.

Hu Z ，Chen G ，Zhao Y ，Gao H ，Li L ，Yin Y ，Jiang J ，Wang L ，Mang Y ，Gao Y ，Zhang S ，Ran J ，Li L ... -  《Molecular Cancer》

Exosomal circ_PIP5K1A regulates the progression of non-small cell lung cancer and cisplatin sensitivity by miR-101/ABCC1 axis.

Shao N ，Song L ，Sun X 《-》

N(6)-methyladenosine-modified circIGF2BP3 inhibits CD8(+) T-cell responses to facilitate tumor immune evasion by promoting the deubiquitination of PD-L1 in non-small cell lung cancer.

An in-depth understanding of immune evasion mechanisms in tumors is crucial to overcome resistance and enable innovative advances in immunotherapy. Circular RNAs (circRNAs) have been implicated in cancer progression. However, much remains unknown regarding whether circRNAs impact immune escape in non-small-cell lung carcinoma (NSCLC). We performed bioinformatics analysis to profile and identify the circRNAs mediating immune evasion in NSCLC. A luciferase reporter assay, RNA immunoprecipitation (RIP), RNA pulldown assays and fluorescence in situ hybridization were performed to identify the interactions among circIGF2BP3, miR-328-3p, miR-3173-5p and plakophilin 3 (PKP3). In vitro T cell-mediated killing assays and in vivo syngeneic mouse models were used to investigate the functional roles of circIGF2BP3 and its downstream target PKP3 in antitumor immunity in NSCLC. The molecular mechanism of PKP3-induced PD-L1 upregulation was explored by immunoprecipitation, RIP, and ubiquitination assays. We demonstrated that circIGF2BP3 (hsa_circ_0079587) expression was increased in NSCLC and negatively correlated with CD8+ T cell infiltration. Functionally, elevated circIGF2BP3 inactivated cocultured T cells in vitro and compromised antitumor immunity in an immunocompetent mouse model, and this effect was dependent on CD8+ T cells. Mechanistically, METTL3 mediates the N6-methyladenosine (m6A) modification of circIGF2BP3 and promotes its circularization in a manner dependent on the m6A reader protein YTHDC1. circIGF2BP3 competitively upregulates PKP3 expression by sponging miR-328-3p and miR-3173-5p to compromise the cancer immune response. Furthermore, PKP3 engages with the RNA-binding protein FXR1 to stabilize OTUB1 mRNA, and OTUB1 elevates PD-L1 abundance by facilitating its deubiquitination. Tumor PD-L1 deletion completely blocked the impact of the circIGF2BP3/PKP3 axis on the CD8+ T cell response. The inhibition of circIGF2BP3/PKP3 enhanced the treatment efficacy of anti-PD-1 therapy in a Lewis lung carcinoma mouse model. Collectively, the PKP3/PD-L1 signature and the infiltrating CD8+ T cell status stratified NSCLC patients into different risk groups. Our results reveal the function of circIGF2BP3 in causing immune escape from CD8+ T cell-mediated killing through a decrease in PD-L1 ubiquitination and subsequent proteasomal degradation by stabilizing OTUB1 mRNA in a PKP3-dependent manner. This work sheds light on a novel mechanism of PD-L1 regulation in NSCLC and provides a rationale to enhance the efficacy of anti-PD-1 treatment in NSCLC.

Liu Z ，Wang T ，She Y ，Wu K ，Gu S ，Li L ，Dong C ，Chen C ，Zhou Y ... -  《Molecular Cancer》