RESUMEN
BACKGROUND: Breast cancer is the leading cause of cancer mortality in women worldwide. Therefore, it is of great significance to identify the biological mechanism of tumorigenesis and explore the development of breast cancer to achieve a better prognosis for individuals suffering from breast cancer. MicroRNAs (miRNAs) have become a hot topic in cancer research, but the underlying mechanism of its involvement in cancer remains unclear. METHODS: The miRNA profile between breast cancer stem cells (BCSCs, CD44+CD24-/low) and control MCF-7 breast cancer cells was obtained in a previous study. Based on biological analysis, miR-20b-5p was hypothesized to be a key factor due to the malignant behavior of BCSCs. Then, agomir-20b-5p and antagomir-20b-5p were transfected into MCF-7 and T47D breast cancer cells to detect cell migration, wound healing and proliferation, and lentivirus vectors silencing or overexpressing miR-20b-5p were transfected into T47D-CSCs to detect proliferation and apoptosis. The effect of miR-20b-5p on xenograft growth was investigated in vivo by transfection of a lentivirus-overexpression vector into T47D cells. The target genes were predicted by the online programs picTar, miRanda and TargetScan and verified by dual luciferase assay, and changes in protein expression were detected by western blot. RESULTS: MiR-20b-5p had the highest degree in both the miRNA-gene network and miRNA-GO network to regulate BCSCs. Overexpression of miR-20b-5p significantly promoted the migration and wound healing ability of MCF-7 cells and T47D cells compared with the control (P < 0.05). In addition, miR-20b-5p facilitated the proliferation of MCF-7 cells and T47D-CSCs (P < 0.05) and inhibited the apoptosis of T47D-CSCs (P < 0.05). Moreover, miR-20b-5p promoted xenograft growth compared with the control group (P < 0.05). Accordingly, potential targets of both CCND1 and E2F1 were predicted by bioinformatics analysis. MiR-20b-5p directly targeted both CCND1 and E2F1 in a dual luciferase assay, while antagomir-20b-5p downregulated the protein levels of CCND1 and E2F1. CONCLUSIONS: Oncogenic miR-20b-5p was confirmed to promote the malignant behaviors of breast cancer cells and BCSCs. The underlying mechanism lies in that miR-20b-5p overall enhanced both CCND1 and E2F1 targets via bidirectional regulation probably involving direct downregulation and indirect upregulation.
Asunto(s)
Neoplasias de la Mama/genética , Ciclina D1/metabolismo , Factor de Transcripción E2F1/metabolismo , Células Madre Neoplásicas/metabolismo , Oncogenes/genética , Animales , Proliferación Celular , Femenino , Humanos , Ratones , Ratones Desnudos , TransfecciónRESUMEN
Acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitors, such as gefitinib and erlotinib, is a critical issue in the treatment of patients with epidermal growth factor receptor mutant-positive non-small-cell lung cancer. Recent evidence suggests that downregulation of gene of phosphatase and tensin homolog deleted on chromosome 10 plays an important role in acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitors in various types of cancers, including lung cancer. It was reported that the E3 ubiquitin ligase neural precursor cell expressed developmentally downregulated gene (NEDD4) (also known as NEDD4-1) negatively regulated phosphatase and tensin homolog deleted on chromosome 10 protein levels through poly-ubiquitination and proteolysis in carcinomas of the prostate, lung, and bladder. Whether this process plays a role in epidermal growth factor receptor-tyrosine kinase inhibitors resistance in non-small-cell lung cancer has not been studied extensively. In view of this, we investigated the involvement of NEDD4 and phosphatase and tensin homolog deleted on chromosome 10 in acquired erlotinib resistance with tyrosine kinase inhibitor-sensitive (HCC827) or tyrosine kinase inhibitor-resistant (Erlotinib-resistant HCC827/ER cells which harbored exon 19 deletion. Overexpression of NEDD4 in HCC827/ER cells was detected, and the reverse correlation between NEDD4 and phosphatase and tensin homolog deleted on chromosome 10 expression in these cells was also revealed. In HCC827/ER cells with knockdown of NEDD4, phosphatase and tensin homolog deleted on chromosome 10 and p-Akt expressions were decreased; the sensitivity of HCC827/ER cells to erlotinib was partially restored. Similar results were also observed in vivo. In H1650/ER cells harboring both exon 19 and phosphatase and tensin homolog deleted on chromosome 10 deletion, expression of p-Akt and sensitivity to erlotinib were not affected by simple knockdown of NEDD4 but affected after transfection of phosphatase and tensin homolog deleted on chromosome 10 into H1650/ER cells. Our results demonstrate that NEDD4 may promote the acquired resistance of non-small-cell lung cancer cells to erlotinib by decreasing phosphatase and tensin homolog deleted on chromosome 10 protein expression. Targeted decrease in NEDD4 expression may be a potential therapeutic strategy for tyrosine kinase inhibitor-resistant non-small-cell lung cancer.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Resistencia a Antineoplásicos/genética , Complejos de Clasificación Endosomal Requeridos para el Transporte/genética , Fosfohidrolasa PTEN/genética , Ubiquitina-Proteína Ligasas/genética , Apoptosis/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Clorhidrato de Erlotinib/administración & dosificación , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Mutación , Ubiquitina-Proteína Ligasas Nedd4 , Transducción de Señal , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PURPOSE: Acquired resistance is a bottleneck that restricts the efficacy of epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) for lung cancer. Ginsenoside Rg3 is an antiangiogenic agent which can down-regulate the expressions of vascular endothelial growth factor (VEGF) and EGFR. Combination of EGFR-TKI and ginsenoside Rg3 may be a promising strategy to delay acquired resistance. This retrospective study explored the efficacy and safety of this combined regimen in patients with EGFR mutation and advanced non-small cell lung cancer (NSCLC). RESULTS: By the deadline of March 31th 2016, the median follow-up period reached 22.9 months. The median PFS was significantly longer in group A than in group B (12.4 months vs 9.9 months, P = 0.017). In addition, ORR was significantly higher in group A than in group B (59.6% vs 41.7%, P = 0.049). The median OS in group A showed no extended tendency compared with that in group B (25.4 months vs 21.4 months, P = 0.258). No significant difference in side effects was found between the two groups. METHODS: A total of 124 patients with advanced NSCLC and EGFR active mutation were collected and analyzed. All of them were treated with first-line EGFR-TKI and divided into two groups. In group A (n=52), patients were administered EGFR-TKI plus ginsenoside Rg3 at standard doses. In group B (n=72), patients received EGFR-TKI alone. Progression-free survival (PFS), overall survival (OS), objective response rate (ORR) and side effects were analyzed. CONCLUSIONS: Ginsenoside Rg3 improves median PFS and ORR of first-line EGFR-TKI treatment in EGFR-mutant advanced NSCLC patients, thus providing a new regimen to delay acquired resistance of EGFR-TKI.