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MiR-193a regulates chemoresistance of human osteosarcoma cells via repression of IRS2.

ABSTRACT: Chemoresistance prevents curative potential of chemotherapy in most cases. MicroRNAs (miRNAs) are key players in regulating chemoresistance in osteosarcoma, which is the most common primary bone cancer. Bisulfite sequencing and quantitative real time PCR analyses showed that miR-193a expression is downregulated by DNA hypermethylation at its promoter region in a chemoresistant cell line, SJSA-1, compared to a chemosensitive cell line G-292. Introduction of a miR-193a mimic in SJSA-1 cells or an antagomir into G-292 cells confirmed the role of miR-193a in osteosarcoma chemoresistance. Bioinformatics together with biochemical assays showed that insulin receptor substrate 2 (IRS2) is a target of miR-193a. Our data concludes that miR-193a plays a role in the osteosarcoma chemoresistance and thus might serve as a useful biomarker for osteosarcoma prognosis.

SUBMITTER: Wang H

PROVIDER: S-EPMC6543196 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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MiR-193a regulates chemoresistance of human osteosarcoma cells via repression of IRS2.

Wang Haiyan H Zhao Fangfang F Cai Shanbao S Pu Youguang Y

Journal of bone oncology 20190511

Chemoresistance prevents curative potential of chemotherapy in most cases. MicroRNAs (miRNAs) are key players in regulating chemoresistance in osteosarcoma, which is the most common primary bone cancer. Bisulfite sequencing and quantitative real time PCR analyses showed that miR-193a expression is downregulated by DNA hypermethylation at its promoter region in a chemoresistant cell line, SJSA-1, compared to a chemosensitive cell line G-292. Introduction of a miR-193a mimic in SJSA-1 cells or an ...[more]

PMID: 31193934

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Project description:Translational control mediated by non-coding microRNAs (miRNAs) plays a key role in the mechanism of cellular resistance to anti-cancer drug treatment. Dihydrofolate reductase (DHFR) and thymidylate synthase (TYMS, TS) are two of the most important targets for antifolate- and fluoropyrimidine-based chemotherapies in the past 50 years. In this study, we investigated the roles of miR-215 in the chemoresistance to DHFR inhibitor methotrexate (MTX) and TS inhibitor Tomudex (TDX).The protein levels of both DHFR and TS were suppressed by miR-215 without the alteration of the target mRNA transcript levels. Interestingly, despite the down-regulation of DHFR and TS proteins, ectopic expression of miR-215 resulted in a decreased sensitivity to MTX and TDX. Paradoxically, gene-specific small-interfering RNAs (siRNAs) against DHFR or TS had the opposite effect, increasing sensitivity to MTX and TDX. Further studies revealed that over-expression of miR-215 inhibited cell proliferation and triggered cell cycle arrest at G2 phase, and that this effect was accompanied by a p53-dependent up-regulation of p21. The inhibitory effect on cell proliferation was more pronounced in cell lines containing wild-type p53, but was not seen in cells transfected with siRNAs against DHFR or TS. Moreover, denticleless protein homolog (DTL), a cell cycle-regulated nuclear and centrosome protein, was confirmed to be one of the critical targets of miR-215, and knock-down of DTL by siRNA resulted in enhanced G2-arrest, p53 and p21 induction, and reduced cell proliferation. Additionally, cells subjected to siRNA against DTL exhibited increased chemoresistance to MTX and TDX. Endogenous miR-215 was elevated about 3-fold in CD133+HI/CD44+HI colon cancer stem cells that exhibit slow proliferating rate and chemoresistance compared to control bulk CD133+/CD44+ colon cancer cells.Taken together, our results indicate that miR-215, through the suppression of DTL expression, induces a decreased cell proliferation by causing G2-arrest, thereby leading to an increase in chemoresistance to MTX and TDX. The findings of this study suggest that miR-215 may play a significant role in the mechanism of tumor chemoresistance and it may have a unique potential as a novel biomarker candidate.

Project description:Astrocyte elevated gene-1 (AEG-1) and endothelin-1 (ET-1)/endothelin A receptor (ETAR) signaling have been demonstrated to be important in osteosarcoma (OS) progression. In the present study, we explored the interaction between AEG-1 and ET-1/ETAR signaling in OS cells, and investigated the mechanism(s) through which the functional interaction may impact OS cell invasion and chemoresistance. Overexpression and knockdown of AEG-1 were performed in Saos-2 and MG-63 OS cells, respectively. Overexpression of AEG-1 in Saos-2 cells significantly increased ET-1 expression (at both the mRNA and protein levels), cell invasion, MMP-2 expression and cell survival against cisplatin. These effects were eradicated using a selective phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, or a selective ETAR inhibitor, BQ123. Knockdown of AEG-1 in MG-63 cells significantly decreased ET-1 expression (at both the mRNA and protein levels), cell invasion, MMP-2 expression and cell survival against cisplatin. Exogenous ET-1 restored cell invasion and MMP-2 expression levels in MG-63 cells, in which AEG-1 had been knocked down, in the presence of LY294002, but not in the presence of BQ123. However, exogenous ET-1 only partially rescued cell survival against cisplatin-induced apoptosis in the presence of LY294002, in cells in which AEG-1 had been knocked down. In conclusion, we have demonstrated that AEG-1 regulates ET-1 expression at the transcriptional level in a PI3K-dependent manner in OS cells. Downstream of PI3K, ET-1/ETAR signaling primarily mediates the promoting effect of AEG-1 on OS cell invasion, likely through the upregulation of MMP-2 expression, thus, ET-1/ETAR signaling partially, but significantly, mediates the AEG-1-induced chemoresistance in OS cells. To the best of our knowledge, this study has provided the first evidence of a functional association between AEG-1 and ET-1/ETAR signaling in OS cells, which adds novel insights into the molecular mechanism of OS metastasis and chemoresistance.

Dataset Information

MiR-193a regulates chemoresistance of human osteosarcoma cells via repression of IRS2.

Publications

MiR-193a regulates chemoresistance of human osteosarcoma cells via repression of IRS2.

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