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Published on: August 12, 2015
Modulation of breast cancer resistance protein (BCRP/ABCG2) gene expression using RNA interference
P L Rachel Ee1, Xiaolong He, Douglas D Ross
1Department of Biopharmaceutical Sciences, University of Illinois at Chicago, Chicago, Illinois 60612, USA.
Abstract:
Overexpression of the breast cancer resistance protein (BCRP/ABCG2) confers multidrug resistance (MDR) to tumor cells and often limits the efficacy of chemotherapy. To circumvent BCRP-mediated MDR, a common approach is the use of potent and specific inhibitors of BCRP transport such as fumitremorgin C, novobiocin, and GF120918. Here, we evaluated a new approach using RNA interference for the specific knockdown of BCRP. We designed and synthesized small interfering RNA (siRNA) using T7 RNA polymerase and showed that siRNAs markedly down-regulated both exogenous and endogenous expression of BCRP. As a functional consequence, knockdown of BCRP by siRNAs increased the sensitivity of human choriocarcinoma BeWo cells to mitoxantrone and topotecan by 10.5- and 8.2-fold, respectively. Using flow cytometry, we found that introduction of siRNAs also enhanced the intracellular accumulation of topotecan. We have previously identified an estrogen response element in the BCRP promoter and have shown that 17beta-estradiol increased BCRP mRNA expression. Furthermore, in the present study, we found that expression of BCRP protein was inducible by 17beta-estradiol and that this effect was ameliorated by the introduction of siRNAs. These studies indicate that siRNAs could modulate MDR in vitro and may present a new approach to overcome BCRP-mediated drug resistance.
Insights
Small interfering RNA (siRNA) effectively reduces breast cancer resistance protein (BCRP/ABCG2) expression, overcoming multidrug resistance (MDR) in cancer cells. This approach enhances chemotherapy efficacy by increasing drug sensitivity and intracellular accumulation.
Area of Science:
- Molecular Biology
- Cancer Research
- Pharmacology
Background:
- Overexpression of breast cancer resistance protein (BCRP/ABCG2) leads to multidrug resistance (MDR) in tumor cells, limiting chemotherapy effectiveness.
- Current strategies to overcome BCRP-mediated MDR involve using specific BCRP transport inhibitors.
Purpose of the Study:
- To evaluate RNA interference (RNAi) using small interfering RNA (siRNA) as a novel approach to down-regulate BCRP expression and overcome BCRP-mediated MDR.
- To assess the impact of BCRP knockdown on the sensitivity of cancer cells to chemotherapeutic agents and their intracellular accumulation.
Main Methods:
- Design and synthesis of siRNAs targeting BCRP using T7 RNA polymerase.
- Evaluation of siRNA efficacy in down-regulating both exogenous and endogenous BCRP expression in human choriocarcinoma BeWo cells.
- Assessment of cellular sensitivity to mitoxantrone and topotecan using flow cytometry and measurement of intracellular drug accumulation.
Main Results:
- siRNAs significantly down-regulated both exogenous and endogenous BCRP expression.
- Knockdown of BCRP by siRNAs increased the sensitivity of BeWo cells to mitoxantrone (10.5-fold) and topotecan (8.2-fold).
- siRNA introduction enhanced the intracellular accumulation of topotecan and ameliorated 17beta-estradiol-induced BCRP protein expression.
Conclusions:
- siRNAs represent a potent tool for down-regulating BCRP expression and overcoming BCRP-mediated multidrug resistance in vitro.
- RNA interference offers a promising new strategy to enhance the efficacy of chemotherapy in BCRP-overexpressing tumors.
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