Dataset Information

A-770041 reverses paclitaxel and doxorubicin resistance in osteosarcoma cells.

ABSTRACT:

Background

Reversing multidrug resistance (MDR) has been an important goal for clinical and investigational oncologists. In the last few decades, significant effort has been made to search for inhibitors to reverse MDR by targeting ATP-binding cassette (ABC) transporters (Pgp, MRP) directly, but these efforts have achieved little clinical success. Protein kinases play important roles in many aspects of tumor cell growth and survival. Combinations of kinase inhibitors and chemotherapeutics have been observed to overcome cancer drug resistance in certain circumstances.

Methods

We screened a kinase specific inhibitor compound library in human osteosarcoma MDR cell lines to identify inhibitors that were capable of reversing chemoresistance to doxorubicin and paclitaxel.

Results

We identified 18 small molecules that significantly increase chemotherapy drug-induced cell death in human osteosarcoma MDR cell lines U-2OSMR and KHOSR2. We identified A-770041 as one of the most effective MDR reversing agents when combined with doxorubicin or paclitaxel. A-770041 is a potent Src family kinase (Lck and Src) inhibitor. Western blot analysis revealed A-770041 inhibits both Src and Lck activation and expression. Inhibition of Src expression in U-2OSMR and KHOSR2 cell lines using lentiviral shRNA also resulted in increased doxorubicin and paclitaxel drug sensitivity. A-770041 increases the intracellular drug accumulation as demonstrated by calcein AM assay.

Conclusions

These results indicate that small molecule inhibitor A-770041 may function to reverse ABCB1/Pgp-mediated chemotherapy drug resistance. Combination of Src family kinase inhibitor with regular chemotherapy drug could be clinically effective in MDR osteosarcoma.

SUBMITTER: Duan Z

PROVIDER: S-EPMC4177239 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Publications

A-770041 reverses paclitaxel and doxorubicin resistance in osteosarcoma cells.

Duan Zhenfeng Z Zhang Jianming J Ye Shunan S Shen Jacson J Choy Edwin E Cote Gregory G Harmon David D Mankin Henry H Hua Yingqi Y Zhang Yu Y Gray Nathanael S NS Hornicek Francis J FJ

BMC cancer 20140919

<h4>Background</h4>Reversing multidrug resistance (MDR) has been an important goal for clinical and investigational oncologists. In the last few decades, significant effort has been made to search for inhibitors to reverse MDR by targeting ATP-binding cassette (ABC) transporters (Pgp, MRP) directly, but these efforts have achieved little clinical success. Protein kinases play important roles in many aspects of tumor cell growth and survival. Combinations of kinase inhibitors and chemotherapeutic ...[more]

PMID: 25236161

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Project description:Multi-drug resistance poses a critical bottleneck in chemotherapy. Given the up-regulation of mTOR pathway in many chemoresistant cancers, we examined whether sirolimus (rapamycin), a first generation mTOR inhibitor, might induce human osteosarcoma (OS) cell apoptosis and increase the sensitivity of OS cells to anticancer drugs in vitro.Human OS cell line MG63/ADM was treated with sirolimus alone or in combination with doxorubicin (ADM), gemcitabine (GEM) or methotrexate (MTX). Cell proliferation and apoptosis were detected using CCK-8 assay and flow cytometry, respectively. MiRNAs in the cells were analyzed with miRNA microarray. The targets of miR-34b were determined based on TargetScan analysis and luciferase reporter assays. The expression of relevant mRNA and proteins was measured using qRT-PCR and Western blotting. MiR-34, PAK1 and ABCB1 levels in 40 tissue samples of OS patients were analyzed using qRT-PCR and in situ hybridization assays.Sirolimus (1-100 nmol/L) dose-dependently suppressed the cell proliferation (IC50=23.97 nmol/L) and induced apoptosis. Sirolimus (10 nmol/L) significantly sensitized the cells to anticancer drugs, leading to decreased IC50 values of ADM, GEM and MTX (from 25.48, 621.41 and 21.72 μmol/L to 4.93, 73.92 and 6.77 μmol/L, respectively). Treatment of with sirolimus increased miR-34b levels by a factor of 7.5 in the cells. Upregulation of miR-34b also induced apoptosis and increased the sensitivity of the cells to the anticancer drugs, whereas transfection with miR-34b-AMO, an inhibitor of miR-34b, reversed the anti-proliferation effect of sirolimus. Two key regulators of cell cycle, apoptosis and multiple drug resistance, PAK1 and ABCB1, were demonstrated to be the direct targets of miR-34b. In 40 tissue samples of OS patients, significantly higher miR-34 ISH score and lower PAK5 and ABCB1 scores were detected in the chemo-sensitive group.Sirolimus increases the sensitivity of human OS cells to anticancer drugs in vitro by up-regulating miR-34b interacting with PAK1 and ABCB1. A low miR-34 level is an indicator of poor prognosis in OS patients.

Project description:BackgroundHeat Shock Transcription Factor 1 (HSF1) is activated under stress conditions. In turn, it induces expression of Heat Shock Proteins (HSPs), which are well-known regulators of protein homeostasis. Elevated levels of HSF1 and HSPs were observed in many types of tumors. The aim of the present study was to determine whether HSF1 could have an effect on the survival of cancer cells treated with chemotherapeutic cytotoxic agents.MethodsWe constructed mouse (B16F10) and human (1205Lu, WM793B) melanoma cells overexpressing full or mutant form of human HSF1: a constitutively active one with a deletion in regulatory domain or a dominant negative one with a deletion in the activation domain. The impact of different forms of HSF1 on the expression of HSP and ABC genes was studied by RT-PCR and Western blotting. Cell cultures were treated with increasing amounts of doxorubicin, paclitaxel, cisplatin, vinblastine or bortezomib. Cell viability was determined by MTT, and IC50 was calculated. Cellular accumulation of fluorescent dyes and side population cells were studied using flow cytometry.ResultsCells overexpressing HSF1 and characterized by increased HSPs accumulation were more resistant to doxorubicin or paclitaxel, but not to cisplatin, vinblastine or bortezomib. This resistance correlated with the enhanced efflux of fluorescent dyes and the increased number of side population cells. The expression of constitutively active mutant HSF1, also resulting in HSPs overproduction, did not reduce the sensitivity of melanoma cells to drugs, unlike in the case of dominant negative form expression. Cells overexpressing a full or dominant negative form of HSF1, but not a constitutively active one, had higher transcription levels of ABC genes when compared to control cells.ConclusionsHSF1 overexpression facilitates the survival of melanoma cells treated with doxorubicin or paclitaxel. However, HSF1-mediated chemoresistance is not dependent on HSPs accumulation but on an increased potential for drug efflux by ABC transporters. Direct transcriptional activity of HSF1 is not necessary for increased expression of ABC genes, which is probably mediated by HSF1 regulatory domain.

Dataset Information

A-770041 reverses paclitaxel and doxorubicin resistance in osteosarcoma cells.

Background

Methods

Results

Conclusions

Publications

A-770041 reverses paclitaxel and doxorubicin resistance in osteosarcoma cells.

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