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Effective knock down of very high ABCG2 expression by a hammerhead ribozyme
Petra Kowalski1, Kate M Farley, Hermann Lage
1Wadsworth Center, Empire State Plaza, Albany, NY 12201, USA.
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.
Materials And 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.
Conclusion:
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.