MiR-526b-3p inhibits lung cancer cisplatin-resistance and metastasis by inhibiting STAT3-promoted PD-L1.
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ABSTRACT:
SUBMITTER: Chen KB
PROVIDER: S-EPMC8319181 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:ACT001, which is derived from an ancient anti-inflammatory drug, has been shown to cross the blood-brain barrier in preclinical studies and has demonstrated anti-glioblastoma (GBM) activity in clinical trials. However, its pharmacological potential for anti-GBM immune response modulation remains unclear. The chemical structure of ACT001 indicates that it may bind to STAT3 and thus modulate antitumor immune response. Methods: Bioinformatics and immunohistochemistry (IHC) were used to assess STAT3 and PD-L1 expression in gliomas. Western blotting, RT-PCR and immunofluorescence were used to detect PD-L1 and p-STAT3 expression in glioma cells exposed to ACT001. Chromatin immunoprecipitation, an ACT001-Biotin probe, and a dual-luciferase reporter assay were used to detect direct modulation. The in vivo efficacy of ACT001 was evaluated in GL261 murine glioma model. Survival analyses were conducted using the log-rank (Mantel-Cox) test. Results: Bioinformatic analysis of 1,837 samples from 4 public glioma datasets showed that STAT3 mRNA expression was correlated with the degree of malignancy and therapeutic resistance and that STAT3 mRNA expression was related to immunosuppression, leukocyte infiltration, and PD-L1 expression. IHC staining of 53 tissue samples confirmed that relatively high phosphorylated STAT3 and PD-L1 protein expression was associated with a relatively advanced World Health Organization (WHO) glioma grade. Next, we confirmed that ACT001 treatment reduced PD-L1 expression and STAT3 phosphorylation. An ACT001-biotin probe was used to verify that ACT001 bound to STAT3. We also demonstrated that STAT3 bound to the PD-L1 promoter. The inhibition of PD-L1 expression and STAT3 phosphorylation by ACT001 could be rescued by STAT3 overexpression. Additionally, ACT001 inhibited GBM growth and decreased PD-L1 expression in vivo. The expression of the M2 markers CD206 and CD163 was decreased, while that of the antitumor immune markers iNOS and IFNγ was increased by ACT001 in vivo. Conclusion: Our results demonstrate that STAT3 plays a key role in immunosuppression of glioma and is inhibited by ACT001. ACT001 inhibits PD-L1 transcription and modulates anti-tumor immune response in glioma bearing mice. These findings will help us to understand the mechanism of ACT001 in GBM therapy.
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Project description:BACKGROUND:Our previous work showed that some Rho GTPases, including Rho, Rac1 and Cdc42, play critical roles in gastric cancer (GC); however, how they are regulated in GC remains largely unknown. In this study, we aimed to investigate the roles and molecular mechanisms of Dock6, an atypical Rho guanine nucleotide exchange factor (GEF), in GC metastasis. METHODS:The expression levels of Dock6 and miR-148b-3p in GC tissues and paired nontumor tissues were determined by immunohistochemistry (IHC) and in situ hybridization (ISH), respectively. The correlation between Dock6/miR-148b-3p expression and the overall survival of GC patients was calculated by the Kaplan-Meier method and log-rank test. The roles of Dock6 and miR-148b-3p in GC were investigated by in vitro and in vivo functional studies. Rac1 and Cdc42 activation was investigated by GST pull-down assays. The inhibition of Dock6 transcription by miR-148b-3p was determined by luciferase reporter assays. RESULTS:A significant increase in Dock6 expression was found in GC tissues compared with nontumor tissues, and its positive expression was associated with lymph node metastasis and a higher TNM stage. Patients with positive Dock6 expression exhibited shorter overall survival periods than patients with negative Dock6 expression. Dock6 promoted GC migration and invasion by increasing the activation of Rac1 and Cdc42. miR-148b-3p expression was negatively correlated with Dock6 expression in GC, and it decreased the motility of GC cells by inhibiting the Dock6/Rac1/Cdc42 axis. CONCLUSIONS:Dock6 was over-expressed in GC tissues, and its positive expression was associated with GC metastasis and indicated poor prognosis of GC patients. Targeting of Dock6 by miR-148b-3p could activate Rac1 and Cdc42, directly affecting the motility of GC cells. Targeting the Dock6-Rac1/Cdc42 axis could serve as a new therapeutic strategy for GC treatment.
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Project description:Head and neck cancer is one of the most prevalent cancers around the world. Head and neck squamous cell carcinoma (HNSCC) accounts for nearly 90% of head and neck cancer. In recent years, significant advances have been made in immunotherapy for HNSCC. Although some clinical trials targeting immune checkpoints have shown success, the molecular mechanism for regulation of programmed death 1 (PD-1) and its ligand (PD-L1) is partially understood. In an effort to explore the effect of activation of signal transducers and activators of transcriptions (STAT3) on PD-1/PD-L1, the expression and correlation between phosphorylation of STAT3 and PD-1/PD-L1 were determined with immunostaining of human and mouse HNSCC tissue sections. PD-1/PD-L1 overexpression was found to be significantly associated with p-STAT3 in human and mouse HNSCC. Targeting STAT3 by a small molecule effectively inhibited the expression of PD-L1 in the CAL27 cell line. Furthermore, we found that blockade of STAT3 signaling downregulated PD-1/PD-L1 in a Tgfbr1/Pten 2cKO HNSCC mouse model. These findings suggest that STAT3 signaling plays an important role in PD-1/PD-L1 regulation and the antitumor immune response of HNSCC.
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Project description:Diffuse large B cell lymphoma (DLBCL) is the commonest disorder derived from the B-lymphocytes. Inhibiting the immune checkpoint through naturalizing programmed death-1 (PD-1) and programmed death ligand 1 (PD-L1) is proved to be a successful therapeutic regime for lymphoma. Long non-coding RNAs (lncRNAs) are unceasingly reported to be promising biological targets for the cancer therapies. This study planned to explore the regulation of small nucleolar RNA host gene 14 (SNHG14) on DLBCL. SNHG14 level in DLBCL samples and cell lines was analyzed by GEPIA bioinformatics tool and RT-qPCR. Biological functions of SNHG14 in DLBCL were detected by CCK-8, colony formation, and transwell invasion assays. Molecular interaction was determined by RNA immunoprecipitation (RIP) and luciferase reporter assays. MiR-5590-3p-related pathway was identified through KEGG pathway analysis applying DAVID6.8 online bioinformatics tool. Effect of SNHG14 on CD8+ T cells was detected by flow cytometry. Results depicted that SNHG14 was upregulated in DLBCL and its depletion retarded proliferation, migration and epithelial-to-mesenchymal transition (EMT). Mechanistically, SNHG14 sponged miR-5590-3p to upregulate Zinc finger E-box binding homeobox 1 (ZEB1), and ZEB1 transcriptionally activated SNHG14 and PD-L1 to promote the immune evasion of DLBCL cells. In conclusion, we firstly showed that SNHG14/miR-5590-3p/ZEB1 positive feedback loop promoted diffuse large B cell lymphoma progression and immune evasion through regulating PD-1/PD-L1 checkpoint, indicating that targeting SNHG14 was a potential approach to improve the efficacy of immunotherapy in DLBCL.
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Project description:BACKGROUND:Angiogenesis plays an important role in the progress of glioma. RNA-binding proteins (RBPs) and circular RNAs (circRNAs), dysregulated in various tumors, have been verified to mediate diverse biological behaviors including angiogenesis. METHODS:Quantitative real-time PCR (qRT-PCR) and western blot were performed to detect the expression of SRSF10, circ-ATXN1, miR-526b-3p, and MMP2/VEGFA. The potential function of SRSF10/circ-ATXN1/miR-526b-3p axis in glioma-associated endothelial cells (GECs) angiogenesis was further studied. RESULTS:SRSF10 and circ-ATXN1 were significantly upregulated in GECs compared with astrocyte-associated endothelial cells (AECs). Knockdown of SRSF10 or circ-ATXN1 significantly inhibited cell viability, migration and tube formation of GECs where knockdown of SRSF10 exerted its function by inhibiting the formation of circ-ATXN1. Moreover, the combined knockdown of SRSF10 and circ-ATXN1 significantly enhanced the inhibitory effects on cell viability, migration and tube formation of GECs, compared with knockdown of SRSF10 and circ-ATXN1, respectively. MiR-526b-3p was downregulated in GECs. Circ-ATXN1 functionally targeted miR-526b-3p in an RNA-induced silencing complex. Up-regulation of miR-526b-3p inhibited cell viability, migration and tube formation of GECs. Furthermore, miR-526b-3p affected the angiogenesis of GECs via negatively regulating the expression of MMP2/VEGFA. CONCLUSION:SRSF10/circ-ATXN1/miR-526b-3p axis played a crucial role in regulating the angiogenesis of GECs. The above findings provided new targets for anti-angiogenic therapy in glioma.
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Project description:BackgroundThe cisplatin-resistance is still a main course for chemotherapy failure of lung cancer patients. Cisplatin-resistant cancer cells own higher malignance and exhibited increased metastatic ability, but the mechanism is not clear. In this study, we investigated the effects of Ataxia Telangiectasia Mutated (ATM) on lung cancer metastasis.Materials and methodsCisplatin-resistant A549CisR and H157CisR cell line were generated by long-term treating parental A549 and H157 cells (A549P and H157P) with cisplatin. Cell growth, cell migration and cell invasion were determined. Gene expressions were determined by Western Blot and qPCR. Tumor metastasis was investigated using a xenograft mouse model.ResultsThe IC50 of the cisplatin-resistant cells (A549CisR and H157CisR cells) to cisplatin was 6-8 higher than parental cells. The A549CisR and H157CisR cells expressed lower level of E-cadherin and higher levels of N-cadherin, Vimentin and Snail compared to the parental A549P and H157P cells, and exhibited stronger capabilities of metastatic potential compared to the parental cells. The ATM expression was upregulated in A549CisR and H157CisR cells and cisplatin treatment also upregulated expression of ATM in parental cells, The inhibition of ATM by using specific ATM inhibitor CP466722 or knock-down ATM by siRNA suppressed Epithelial-to-Mesenchymal transition (EMT) and metastatic potential of A549CisR and H157CisR cells. These data suggest that ATM mediates the cisplatin-resistance in lung cancer cells. Expressions of JAK1,2,、 STAT3 、PD-L1 and ATM were increased in A549CisR and H157CisR cells and could by induced by cisplatin in parental lung cancer cells. Interestedly, ATM upregulated PD-L1 expression via JAK1,2/STAT3 pathway and inhibition of ATM decreased JAK/STAT3 signaling and decreased PD-L1 expression. The treatment of PD-L1 neutralizing Ab reduced EMT and cell invasion. Inhibition of JAK1,2/STAT3 signaling by specific inhibitors suppressed ATM-induced PD-L1 expression, EMT and cell invasion. Importantly, inhibition of ATM suppressed EMT and tumor metastasis in cisplatin-resistant lung cancer cells in an orthotopic xenograft mouse model.ConclusionsOur results show that ATM regulates PD-L1 expression through activation of JAK/STAT3 signaling in cisplatin-resistant cells. Overexpression of ATM contributes to cisplatin-resistance in lung cancer cells. Inhibition of ATM reversed EMT and inhibited cell invasion and tumor metastasis. Thus, ATM may be a potential target for the treatment of cisplatin-resistant lung cancer.
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Project description:Increasing bodies of evidence support the involvement of tumor-intrinsic action in PD-L1-mediated cancer progression. However, the mechanisms underlying the tumor-intrinsic function of PD-L1 are less well understood. In the present study, we found a positive correlation between PD-L1 expression and MET phosphorylation in lung cancer and melanoma cell lines. PD-L1 inhibition led to a decrease in MET phosphorylation, while PD-L1 induction by IFN-γ resulted in a PD-L1-dependent increase of MET phosphorylation both in vitro and in vivo. The results indicated that MET phosphorylation can be positively regulated by PD-L1. Furthermore, we identified PTP1B as a mediator contributing to the regulation of MET phosphorylation by PD-L1. In agreement with the induction of MET phosphorylation by PD-L1, inhibition of PD-L1 caused reduced phosphorylation of ERKs, a known downstream kinase of MET, and inhibited cell proliferation. Collectively, the present study demonstrated for the first time that the MET pathway, as a downstream of PD-L1, contributed to its tumor-intrinsic effect, and provided a novel mechanistic explanation for the tumor-intrinsic function of PD-L1 and a rationale for the combination of immunotherapy and MET-targeted therapy in cancer treatment.
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Project description:The anti-PD-L1 monoclonal antibody (mAb) targeting PD-1/PD-L1 immune checkpoint has achieved outstanding results in clinical application and has become one of the most popular anti-cancer drugs. The mechanism of molecular recognition and inhibition of PD-L1 mAbs is not yet clear, which hinders the subsequent antibody design and modification. In this work, the trajectories of PD-1/PD-L1 and nanobody/PD-L1 complexes were obtained via comparative molecular dynamics simulations. Then, a series of physicochemical parameters including hydrogen bond, dihedral angle distribution, pKa value and binding free energy, and so forth, were all comparatively analyzed to investigate the recognition difference between PD-L1 and PD-1 and nanobody. Both LR113 (the amino acid residues in PD-L1 are represented by the lower left sign of L) and LR125 residues of PD-L1 undergo significant conformational change after association with mAbs, which dominates a strong electrostatic interaction. Solvation effect analysis revealed that solvent-water enhanced molecular recognition between PD-L1 and nanobody. By combining the analyses of the time-dependent root mean squared fluctuation (RMSF), free energy landscape, clustering and energy decomposition, the potential inhibition mechanism was proposed that the nanobody competitively and specifically bound to the ?-sheet groups of PD-L1, reduced the PD-L1’s flexibility and finally blocked the formation of PD-1/PD-L1 complex. Based on the simulation results, site-directed mutagenesis of ND99 (the amino acid residues in Nano are displayed by the lower left sign of N) and NQ116 in the nanobody may be beneficial for improving antibody activity. This work offers some structural guidance for the design and modification of anticancer mAbs based on the structure of the PD-1/PD-L1 complex.
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