Hsa_circ_0125356 promotes gemcitabine resistance by modulating WNT canonical and non-canonical pathways via miR-582-5p/FGF9 axis in non-small cell lung cancer.
Non-small cell lung cancer (NSCLC) is the leading cause of cancer morbidity and mortality worldwide. The prognosis of patients has been significantly improved by chemotherapy, but acquired drug resistance remains a major obstacle to NSCLC treatment. Circular RNAs (circRNAs), which act as miRNA or protein sponges, are critically associated with the development and chemotherapy resistance of NSCLC.
CircRNA sequencing was performed to analyze the differential expression of circRNAs between A549 and A549-GR cells. Chromogenic in situ hybridization (CISH) and immunohistochemistry (IHC) technologies were used to detect the expression of hsa_circ_0125356, miR-582-5p,and FGF9 in NSCLC tissues and para-carcinoma tissues. Fluorescence in situ hybridization (FISH), dual-luciferase reporter assays and RNA immunoprecipitation (RIP) were conducted to evaluate the expression and regulation of hsa_circ_0125356, miR-582-5p, and FGF9. Furthermore, the regulation of hsa_circ_0125356/miR-582-5p/FGF9 on gemcitabine sensitivity was confirmed by TUNEL, Transwell, EdU, CCK8 and immunohistochemistry.
We identified a novel hsa_circ_0125356 as a therapeutic target against gemcitabine resistance. Hsa_circ_0125356 was significantly elevated in clinical samples of patients with NSCLC. Moreover, hsa_circ_0125356 overexpression promoted gemcitabine resistance to NSCLC by upregulating FGF9 via sponging miR-582-5p in vivo and in vitro. Notably, WNT canonical (ERK/GSK3β/β-catenin) and non-canonical (Daam1/RhoA/ROCK2) signaling pathways were activated due to hsa_circ_0125356 acting as an endogenous miR-582-5p sponge to regulate the expression of FGF9, and thereby enhancing gemcitabine resistance via promoting DNA damage repair and inhibition of apoptosis. The results were further confirmed by two small molecule antagonists, WAY 316606 and XAV-939,which could inhibit the activation of WNT signaling pathway induced by hsa_circ_0125356.
We first demonstrated that hsa_circ_0125356 was significantly upregulated and served as a biomarker for gemcitabine resistance in NSCLC by sponging miR-582-5p/FGF9 axis to regulate the WNT canonical and non-canonical signaling pathways, which provided a new direction for identification of therapeutic targets for the treatment of gemcitabine resistance of NSCLC.
Du X
，Luo W
，Li H
，Gu Q
，Huang P
，Wang C
，Li N
，Liu F
，Xia C
... -
《Molecular Cancer》
Inhibition of id-1 reduces osteosarcoma growth and metastasis through mediation of snail.
Osteosarcoma (OS) is a highly invasive bone tumor that frequently metastasizes to the lungs. This study aims to investigate the role of the Id-1 gene in OS invasion and metastasis, and its relationship with the Snail gene.
This study included tissue samples from 12 non-metastatic osteosarcomas and 9 metastatic osteosarcoma patients to examine the expression of Id-1 and Snail using RT-qPCR and analyze their correlation. In cell-based experiments, four osteosarcoma cell lines (Saos-2, U2OS, MG-63, and 143B) and the human osteoblast cell line hFOB 1.19 were cultured. The expression of Id-1 and Snail was evaluated by RT-qPCR and Western blotting.Cells were randomly divided into the Control group, sh-NC group, and sh-Id-1 group using lentiviral infection. Transwell invasion and scratch assays were used to assess cell migration and invasion. WB was employed to detect the expression of Id-1, Snail, and epithelial-mesenchymal transition (EMT)-related proteins (E-cadherin, vimentin, and N-cadherin) in the OS cells of each group. In animal experiments, Tumor formation in each group was evaluated by injecting cells subcutaneously into mice. An osteosarcoma lung metastasis model was established by injecting infected cells into the tibia of mice. Tumor growth and lung metastasis were observed using HE staining. The expression of Id-1, Snail, and EMT-related proteins in osteosarcoma and lung tissues from each group of mice was assessed using Western blot and immunohistochemistry.
The expression of Id-1 and Snail was significantly higher in osteosarcoma tissues than in normal bone tissues, and the expression of Id-1 was positively correlated with that of Snail. In cell experiments, downregulation of Id-1 reduced Snail expression and significantly inhibited EMT, as well as the migration and invasion of OS cells (P < 0.05). In animal experiments, compared to the Control group, the sh-Id-1 group mice was no significant change in body weight, but the tumor volume was significantly reduced, and fewer lung metastatic nodules (P < 0.05). HE staining indicated decreased nuclear atypia, reduced invasion and destruction, fewer new blood vessels, and less calcification in the sh-Id-1 group tumors. Immunohistochemistry and WB results showed upregulation of E-cadherin and downregulation of vimentin, N-cadherin, Id-1, and Snail in the sh-Id-1 group (P < 0.05).
Downregulation of Id-1 inhibits the EMT process by reducing Snail expression, effectively suppressing the growth, invasion, and lung metastasis of OS.
Shu R
，Yu Z
，Wu J
，Cheng Q
，Peng Z
，Zhou H
，Zhao M
... -
《Journal of Orthopaedic Surgery and Research》
Intervening Non-Small-Cell Lung Cancer Progression by Cell Membrane Coated Platycodin D via Regulating Hsa-miR-1246/FUT9/GSK3β Pathway.
Metastatic non-small cell lung cancer (NSCLC) remains a global health threat, with patients facing inevitable disease progression despite standard-of-care therapy. Prior studies showed Platycodin D (PD)-induced cell cycle arrest and apoptosis in NSCLC via RNA regulatory network, yet elucidating PD's mechanisms in NSCLC progression is challenging in the real world.
Biological effects of PD on NSCLC cell lines A549 and PC-9 were assessed through in vitro assays, encompassing apoptosis, proliferation, colony formation, migration and invasion. MicroRNAs (miRNAs) expression was profiled, and their roles were investigated using miRNA mimics or inhibitors. Predicted miRNA targets were validated via dual-luciferase reporter assays and Western blotting following bioinformatic prediction. PD's metastatic inhibitory potential in NSCLC was evaluated in an in vivo lung cancer metastasis model. Furthermore, a homologous cell membrane-based PD delivery system was established to improve the biosafety and efficacy of PD in vivo.
Hsa-miR-1246 was upregulated by PD treatment, and functional experiments demonstrated that the miR-1246-mimic enhanced PD's suppressive effects on NSCLC cell proliferation, colony formation, migration, and invasion, while the miR-1246-inhibitor abrogated these effects. Notably, dual-luciferase assays confirmed that hsa-miR-1246 directly targeted the 3' untranslated regions (3' UTRs) of Fucosyltransferase 9 (FUT9), modulating its expression. Moreover, the hsa-miR-1246/FUT9 axis regulated the phosphorylation level and expression of GSK3β protein. In vivo, PD encapsulated in homologous cell membranes mitigated tumor growth and migration in metastatic NSCLC mice with minimal side effects.
The application of PD prompted an increase in the expression levels of hsa-miR-1246 and a concurrent decrease in FUT9. Importantly, the therapeutic efficacy of PD in vivo was markedly enhanced through homologous cell delivery system. Collectively, this study revealed the potential utility of PD in the treatment of NSCLC progression.
Zheng S
，Xie Z
，Zhou Z
，Wang S
，Xin Y
，Lin J
，Cheng K
，Lu T
，Qi R
，Guo Y
... -
《International Journal of Nanomedicine》
Exosomal miR-20b-5p Induces EMT and Enhances Progression in Non-Small Cell Lung Cancer Via TGFBR2 Downregulation.
The mechanism by which specific miRNAs in NSCLC exosomes regulate NSCLC progression remains unclear. First, exosomes were isolated and identified. Exosomes were labeled with PKH26 for cell tracking studies. Subsequently, BEAS-2B cells and BEAS-2B cell exosomes were transfected with miR-20b-5p mimics or miR-20b-5p inhibitors, and cell proliferation was measured by EdU and CCK-8. cell migration and invasion were detected by wound healing tests and Transwell. The potential target of miR-20b-5p was predicted and verified by luciferase assay. Relative expression levels of miR-20b-5p and TGFBR2 were detected by qRT-PCR. Protein expression level was detected by Western blot. In addition, A549 cell exosomes were injected into mice through the tail vein and the pathological changes of lung tissue were detected by HE staining. Expression levels of E-cadherin and Vimentin in lung tissues were detected by immunohistochemistry. Our results also showed that high levels of miR-20b-5p in exosomes generated from NSCLC cells conceivably enter the cytoplasm of their own cells and by downregulating TGFBR2, accelerate NSCLC invasion, migration and EMT while promoting NSCLC cell proliferation. Exosome analysis using clinical plasma specimens revealed that miR-20b-5p expression was considerably improved in exosomes from NSCLC patients compared with those from healthy controls. In vitro and in vivo, exosomes with high levels of miR-20b-5p were linked to the progression of NSCLC. Our data suggest that exosomes with high levels of miR-20b-5p can increase development and metastasis of NSCLC cells by downregulating TGFBR2, which would serve as a predictive and diagnostic marker for NSCLC.
Ma H
，Jiang B
，Ren Q
，Sun Y
，Wang M
，Xia S
，Wang D
，Zhang W
... -
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