hTERT promoter targeting reverse drug resistance in breast cancer combined with RNAi

HUANG Jin-xiang; et al

doi:10.13406/j.cnki.cyxb.2010.10.040

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hTERT promoter targeting reverse drug resistance in breast cancer combined with RNAi

更新时间：2024-01-17

- hTERT promoter targeting reverse drug resistance in breast cancer combined with RNAi
- Journal of Chongqing Medical University Vol. 35, Issue 10, Pages: 1482-1485(2010)
- 作者机构：
  
  重庆医科大学附属第二医院普外科
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2010.10.040
  CLC： R737.9
- Published：2010，
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[1]黄金向,王继见,莫琳君.使用hTERT启动子联合RNAi靶向逆转乳腺癌的耐药性[J].重庆医科大学学报,2010,35(10):1482-1485.

HUANG Jin-xiang, et al. hTERT promoter targeting reverse drug resistance in breast cancer combined with RNAi. [J]. Journal of Chongqing Medical University 35(10):1482-1485(2010)
[1]黄金向,王继见,莫琳君.使用hTERT启动子联合RNAi靶向逆转乳腺癌的耐药性[J].重庆医科大学学报,2010,35(10):1482-1485. DOI： 10.13406/j.cnki.cyxb.2010.10.040.

HUANG Jin-xiang, et al. hTERT promoter targeting reverse drug resistance in breast cancer combined with RNAi. [J]. Journal of Chongqing Medical University 35(10):1482-1485(2010) DOI： 10.13406/j.cnki.cyxb.2010.10.040.

摘要

目的:克隆人端粒酶逆转录酶启动子（Human telomerase reverse transcriptase

hTERT）

探讨对乳腺癌细胞的特异性[1]

利用RNAi技术靶向沉默耐药乳腺癌细胞MCF-7/ADR乳腺癌耐药蛋白（Breast cancer resistance protein

BCRP）基因[2]的表达

观察BCRP的mRNA变化

BCRP蛋白的表达以及对阿霉素耐药性的变化[3]。方法:构建重组质粒（pGenesil-CMV-shRNA、pGenesil-hTERT-shRNA）

分别转染端粒酶阴性及阳性细胞

检测hTERT启动子的靶向性

应用RT-PCR及Western-blot技术检测不同组的细胞的RNA、蛋白差异

MTT法分析药敏性变化[4]。结果:端粒酶阳性的细胞可见很强的荧光

明显强于端粒酶阴性的细胞[5];细胞实验RT-PCR及Western blot检测结果显示:BCRP的mRNA和蛋白的表达受抑制与对照组其差异有显著性（P<0.05）;MTT比色法结果显示:对阿霉素的药物敏感性明显提高。结论:使用hTERT启动子联合RNAi成功靶向沉默BCRP基因

乳腺癌MCF-7/ADR对阿霉素的敏感性明显增加

为进一步开发肿瘤的特异性基因沉默治疗奠定实验基础。

Abstract

Objective:To clone the promoter of human telomerase reverse transcriptase (hTERT)and investigate its selective effect on breast cancer cell

and to observe changes of breast cancer resistance protein(BCRP) mRNA

its protein expression and changes of drug resistance to doxorubicin with RNAi technology used in cell (MCF-7/ADR). Methods:Plasmids (pGenesil-CMV-shRNA

pGenesil-hTERT-shRNA)were recombined and transfected into telomerase-negative and-positive cells

hTERT promoter targeting was detected.RT-PCR

and Western-blot were applied to detect RNA and protein of different groups of cells. MTT was applied to analyze drug sensitivity. Results:Highly visible fluorescent was seen in telomerase-positive cells

which was stronger than that of telom-erase-negative cells.The results of RT-PCR

and Western-blot showed that the mRNA and protein expression of BCRP were inhibited ; MTT results showed that the drug sensitivity to doxorubicin was significantly increased

which was of significant difference as compared with the control group ( P<0.05 ) . Conclusion : hTERT promoter can targeting silence BCRP gene combined with RNAi technology and drug resistance to doxorubicin is reversed

which lays a solid experimental basis for the further development of tumor specific therapy in gene silencing.

关键词

重组质粒启动子hTERTRNAi乳腺癌MCF-7/ADR端粒酶阳性细胞

Keywords

PlasmidhTERTpromoterRNAibreast cancerMCF-7/ADRTelomerace-positive cells

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