Reduction of expression of the multidrug resistance protein (MRP) in human tumor cells by antisense phosphorothioate oligonucleotides

Multidrug resistance protein (MRP) is a member of the ATP-binding cassette superfamily of transport proteins which has been demonstrated to cause multidrug resistance when transfected into previously sensitive cells. Sixteen elcosomeric oligonucleotides complementary to different regions along the entire length of the MRP mRNA reduced MRP mRNA and protein levels in drug resistant small cell lung cancer cells that highly overexpress this protein. In MRP-transfected HeLa cells that express intermediate levels of MRP, one oligonucleotide, ISIS 7597, targeted to the coding region of the MRP mRNA, decreased the levels of MRP mRNA to <10% of control levels in a concentration dependent manner. This effect was rapid but transient with a return to control levels of MRP mRNA 72 hr after treatment. A double treatment with ISIS 7597 produced a sustained inhibition, resulting in a greater than 90% reduction in MRP mRNA for 72 hr and a comparable decrease in protein levels. Increased sensitivity to doxorubicin was observed under these conditions. Northern blotting analyses using two DNA probes corresponding to sequences 5' and 3' of the ISIS 7597 target sequence, respectively, revealed the presence of low levels of two smaller sized RNA fragments as expected from an RNase H-mediated mechanism of action of the antisense oligonucleotide. These studies indicate that a specific reduction in MRP expression can be achieved with antisense oligonucleotides, a finding that has potential implications for the treatment of drug resistant tumors.