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Updated: Mar 25, 2026
Cytotoxic Efficacy of Photodynamic Therapy in Osteosarcoma Cells In Vitro
Published on: March 18, 2014
Sirolimus induces apoptosis and reverses multidrug resistance in human osteosarcoma cells in vitro via increasing
Yan Zhou1, Rui-hua Zhao2, Kuo-fu Tseng3
1Department of Oncology, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, China.
Aim:
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.
Methods:
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.
Results:
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.
Conclusion:
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.
Insights
Sirolimus, an mTOR inhibitor, enhances chemotherapy effectiveness in osteosarcoma by boosting miR-34b levels. This increases sensitivity to drugs like doxorubicin and indicates low miR-34b predicts poor prognosis.
Area of Science:
- Oncology
- Molecular Biology
- Pharmacology
Background:
- Multi-drug resistance is a major challenge in chemotherapy.
- The mTOR pathway is often upregulated in chemoresistant cancers.
Purpose of the Study:
- To investigate if sirolimus (rapamycin) can induce apoptosis in human osteosarcoma (OS) cells.
- To determine if sirolimus can increase the sensitivity of OS cells to anticancer drugs.
Main Methods:
- Human OS cells were treated with sirolimus alone or with doxorubicin, gemcitabine, or methotrexate.
- Cell proliferation, apoptosis, and miRNA expression were analyzed.
- Target genes of miR-34b were identified, and protein/mRNA levels were measured.
Main Results:
- Sirolimus suppressed proliferation and induced apoptosis in OS cells.
- Sirolimus significantly sensitized cells to doxorubicin, gemcitabine, and methotrexate.
- Sirolimus upregulated miR-34b, which targeted PAK1 and ABCB1, affecting cell cycle and drug resistance.
Conclusions:
- Sirolimus enhances OS cell sensitivity to chemotherapy by upregulating miR-34b, targeting PAK1 and ABCB1.
- Low miR-34 levels in OS patients correlate with a poor prognosis.
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