Dataset Information

Overexpression of miR-15b Promotes Resistance to Sunitinib in Renal Cell Carcinoma.

ABSTRACT: Aim: Sunitinib remains the frontline treatment for metastatic clear-cell renal cell carcinoma (ccRCC). Drug resistance is inevitable and related mechanism warrant insightful elaboration. Methods: In silico data mining of GEO and TCGA datasets was performed to identify potential target micro-RNA. In vitro and in vivo studies were performed to validate findings. Results: Reproduction of GEO datasets revealed miR-15b significantly upregulated in sunitinib- resistant ccRCC. Five out of seven ccRCC cell lines demonstrated significantly overexpressed miR-15b after sunitinib treatment. Vector-mediated overexpression of miR-15b significantly induced resistance to sunitinib in ccRCC cells. Overexpression of miR-15b significantly induced less population in G1 phase of cell cycle and less apoptosis in cells treated sunitinib. Expression of genes negatively correlated with miR-15b in TCGA ccRCC (KIRC) dataset were cross-referenced with predicted targets of miR-15b and CCNC was selected as potential target for resistance mediation. Overexpression of miR-15b suppressed CCNC expression and protein (Cyclin C) levels. Cyclin C-associated proteins CDK19 and CDK8 were also suppressed following miR-15b overexpression. Silencing of CCNC mimicked overexpression of miR-25 inducing cell cycle progression passing G1 phase and less apoptosis in ccRCC cells treated by sunitinib. Overexpression of miR-15b also counteracted suppression of migration and colony formation by sunitinib in ccRCC cell lines. In vivo mouse xenograft models showed recovered tumor growth with miR-15b expression in mice treated with sunitinib. Conclusion: We here show miR-15b as a possible culprit for sunitinib resistance in ccRCC. Targeting miR-15b could potentially overcome drug resistance and related mechanism warrants further investigation.

SUBMITTER: Lu L

PROVIDER: S-EPMC6603409 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Overexpression of miR-15b Promotes Resistance to Sunitinib in Renal Cell Carcinoma.

Lu Lu L Li Yanmin Y Wen Hui H Feng Chenchen C

Journal of Cancer 20190609 15

Aim: Sunitinib remains the frontline treatment for metastatic clear-cell renal cell carcinoma (ccRCC). Drug resistance is inevitable and related mechanism warrant insightful elaboration. Methods: In silico data mining of GEO and TCGA datasets was performed to identify potential target micro-RNA. In vitro and in vivo studies were performed to validate findings. Results: Reproduction of GEO datasets revealed miR-15b significantly upregulated in sunitinib- ...[more]

PMID: 31293642

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Project description:Rationale: Although sunitinib has been shown to improve the survival rate of advanced renal cell carcinoma (RCC) patients, poor drug response is a major challenge that reduces patient benefit. It is important to elucidate the underlying mechanism so that the therapeutic response to sunitinib can be restored. Methods: We used an Illumina HumanMethylation 850K microarray to find methylation-differentiated CpG sites between sunitinib-nonresponsive and -responsive RCC tissues and Sequenom MassARRAY methylation analysis to verify the methylation chip results. We verified glutaminyl peptide cyclotransferase (QPCT) expression in sunitinib-nonresponsive and -responsive RCC tissues via qRT-PCR, western blot and immunohistochemical assays. Then, cell counting kit 8 (CCK-8), plate colony formation and flow cytometric assays were used to verify the function of QPCT in RCC sunitinib resistance after QPCT intervention or overexpression. Chromatin immunoprecipitation (ChIP) was performed to clarify the upstream regulatory mechanism of QPCT. A human proteome microarray assay was used to identify downstream proteins that interact with QPCT, and co-immunoprecipitation (co-IP) and confocal laser microscopy were used to verify the protein chip results. Results: We found that the degree of methylation in the QPCT promoter region was significantly different between sunitinib-nonresponsive and -responsive RCC tissues. In the sunitinib-nonresponsive tissues, the degree of methylation in the QPCT promoter region was significantly reduced, and the expression of QPCT was upregulated, which correlated with a clinically poor response to sunitinib. A knockdown of QPCT conferred sunitinib sensitivity traits to RCC cells, whereas an overexpression of QPCT restored sunitinib resistance in RCC cells. Mechanistically, reducing the methylation degree of the QPCT promoter region by 5-aza-2'-deoxycytidine (decitabine) in RCC cells could increase the expression of QPCT and NF-?B (p65) bound to the QPCT promoter region, positively regulating its expression, while the hypermethylation in the QPCT promoter region could inhibit the binding of NF-?B (p65). QPCT could bind to HRAS and attenuate the ubiquitination of HRAS, thus increasing its stability and leading to the activation of the ERK pathway in RCC cells. Conclusion: QPCT may be a novel predictor of the response to sunitinib therapy in RCC patients and a potential therapeutic target.

Dataset Information

Overexpression of miR-15b Promotes Resistance to Sunitinib in Renal Cell Carcinoma.

Publications

Overexpression of miR-15b Promotes Resistance to Sunitinib in Renal Cell Carcinoma.

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