Effective knock down of very high ABCG2 expression by a hammerhead ribozyme.
BACKGROUND
Ribozymes are an effective tool to reduce the mRNA levels of specific target genes. Overexpression of the drug transport protein, ABCG2, has been associated with multidrug resistance in cancer cells.


METHODS
An expression plasmid encoding a hammerhead ribozyme against the ABCG2 gene was stably transfected into multidrug-resistant MCF7/MX cells that express very high levels of the ABCG2 protein. The effect of the ribozyme was determined by quantitative real-time RT-PCR, Western blot and cytotoxicity assays.


RESULTS
The ribozyme reduced ABCG2 mRNA levels to less than 10% of control values, which resulted in the concomitant reduction of ABCG2 protein levels and sensitization of the cells to mitoxantrone and methotrexate.


CONCLUSIONS
The ribozyme used was highly effective in reducing the expression of its target gene, ABCG2, and was able to modulate the associated multidrug-resistant phenotype.
Wadsworth Center, Empire State Plaza, Albany, NY 12201, USA.
Petra
P
Kowalski
Kate M
KM
Farley
Hermann
H
Lage
Erasmus
E
Schneider
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D000970:Antineoplastic Agents / Q000494:pharmacology; D001943:Breast Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D008942:Mitoxantrone / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D016337:RNA, Catalytic / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019772:Topotecan / Q000494:pharmacology; D014162:Transfection
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000970:Antineoplastic Agents; D009363:Neoplasm Proteins; D016337:RNA, Catalytic; C427204:hammerhead ribozyme; D019772:Topotecan; D008942:Mitoxantrone; D008727:Methotrexate