AN ETOPOSIDE-RESISTANT LUNG-CANCER SUBLINE OVEREXPRESSES THE MULTIDRUG RESISTANCE-ASSOCIATED PROTEIN

We have characterised an etoposide-resistant subline of the small-cell lung cancer cell line, UMCC-1, derived at our centre. Subline UMCC-1/ VP was developed by culturing the parent line in increasing concentrat ions of etoposide over 16 months. UMCC-1/VP is 20-fold resistant to et oposide by MTT assays, relative to the parent line, and is cross-resis tant to doxorubicin, vincristine and actinomycin D, but not to taxol, cisplatin, melphalan, thiotepa or idarubicin. Topoisomerase II immunob lotting demonstrates a 50% reduction of the protein in the resistant s ubline. The UMCC-1/VP subline demonstrates a marked decrease in the ac cumulation of [H-3]etoposide relative to the parent line, as well as a modest reduction in the accumulation of daunorubicin. Reverse transcr iption-polymerase chain reaction assays demonstrate no detectable mdr1 expression but marked expression of the multidrug resistance-associat ed protein (MRP) gene in the resistant subline. Northern blotting with an MRP cDNA probe confirms marked overexpression of the MRP gene only in the UMCC-1/VP subline. Western blotting with antisera against MRP peptide confirms a 195 kDa protein band in the UMCC-1/VP subline. Sout hern blotting experiments demonstrate a 10-fold amplification of the M RP gene in the resistant subline. Depletion of glutathione with buthio nine sulphoximine sensitised UMCC-1/VP cells to daunorubicin and etopo side. Our studies indicate that MRP gene expression may be induced by etoposide and may lead to reduced accumulation of the drug.