Reversion of multidrug resistance in the P-glycoprotein positive breast cancer cell line (MCF-7/ADR) by introduction of hammerhead ribozyme.
A hammerhead ribozyme which site-specifically cleaved the GUC position in codon 880 of the mdr1 mRNA was designed. The target site was chosen between the two ATP binding sites, which may be important for the function of the P-Gp as an ATP-dependent pump. A DNA sequence encoding the ribozyme gene was then incorporated into a eukaryotic expression vector (pH beta Apr-1 neo) and transfected into the breast cancer cell line MCF-7/Adr, which is resistant to adriamycin and expresses the MDR phenotype. The ribozyme was stably expressed in the cell line by the RNA dot blotting assay. The result of Northern blot assay showed that the expressed ribozyme could decrease the level of mdr1 mRNA expression by 83.5%; and the expressed ribozyme could inhibit the formation of P-glycoprotein detected by immuno-cytochemistry assay and could reduce the cell's resistance to adriamycin; this means that the resistant cells were 1,000-fold more resistant than the parental cell line (MCF-7), whereas those cell clones that showed ribozyme expression were only 6-fold more resistant than the parental cell line. These results show that a potentially useful tool is at hand which may inactivate MDR1 mRNA and revert the multidrug resistance phenotype.
Yuan
Y
Y
Department of Oncology, Zhujiang Hospital, First Military Medical University, Guangzhou 510282.
Zhang
J
J
Scanlon
K J
KJ
Lu
Z
Z
Qi
G
G
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis*; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D019450:Genes, MDR; D006801:Humans; D016337:RNA, Catalytic / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D016337:RNA, Catalytic; D012333:RNA, Messenger; D004317:Doxorubicin