Reversal of MDR1 gene-dependent multidrug resistance in HL60/HT9 cells using short hairpin RNA expression vectors.
Multidrug resistance (MDR) is a serious obstacle to cancer chemotherapy. Overexpression of P-glycoprotein (P-gp), the MDR1 gene product, confers MDR to tumor cells. This study explored the possibility of reducing drug resistance by targeting the mdr1 gene using short hairpin RNA (shRNA). Two different shRNAs were designed and constructed in a pSilencer 2.1-U6 neo plasmid. The shRNA recombinant plasmids were transfected into HT9 leukemia cells. Real-time polymerase chain reaction and Western blotting were used to characterize the inhibited expression of MDR1 mRNA and P-gp, and the drug sensitivity of the transfected cells was assessed using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. The results indicated that the inhibition of P-gp expression by small interfering RNA selectively restored sensitivity to the drugs transported by P-gp. Evaluation of chemosensitivity showed 52.58% reversal by p2.1-shRNA1 and 73.07% reversal by p2.1-shRNA2 in drug resistance for harringtonine, and 84.87% reversal by p2.1-shRNA1 and 94.23% reversal by p2.1-shRNA2 in drug resistance for curcumin in the transfected cells. The results demonstrated the efficacy and selectivity of shRNA in reversing MDR in drug-resistant HT9 cells in vitro.
Shao
Shu-Li
SL
College of Life Science and Engineering, Qiqihar University, Qiqihaer, Hei Longjiang Province, China.
Zhang
Wei-Wei
WW
Li
Xu-Yan
XY
Zhang
Zhen-Zhu
ZZ
Yun
Dong-Ze
DZ
Fu
Bo
B
Zuo
Ming-Xue
MX
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000235:genetics; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D045744:Cell Line, Tumor; D003474:Curcumin / Q000494:pharmacology; D018432:Drug Resistance, Multiple / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D005822:Genetic Vectors / Q000627:therapeutic use*; D006801:Humans; D007938:Leukemia / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000627:therapeutic use*; D020133:Reverse Transcriptase Polymerase Chain Reaction
C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D003474:Curcumin