Cell lines derived from the human ovarian tumor, which is sensitive to taxol, and TR-21, which is highly resistant, were clonally isolated. The latter clones were found to express P-glycoprotein (P-GP) by fluorescent staining with anti-P-GP monoclonal antibody and Western blotting. Transfection of the multidrug resistence type 1 (MDR-1) gene into TAOV cells resulted in moderate resistance to taxol and expression of P- glycoprotein (P-GP), but the resistance was not dominant as in TR-21 subclones. The influx of 3H-taxol into TR-21 cells was 37-fold lower than that of TAOV cells, and 16-fold lower than that of the MDR-1 gene transfected cell line. High performance liquid chromatagraphy analysis of taxol metabolites in the taxol exposed cells as well as in culture media indicated that taxol was converted to one or two metabolites in TAOV cells, but the amounts of metabolites in the cells and media were only a fraction of the total taxol. Metabolites of taxol in the resistant cells were undetectable because of the low level of 3H-taxol influx and the high level of efflux. Treatment of the resistant cells with verapamil, a calcium channel blocker, resulted in a significantly high level of cell death and active taxol metabolism, indicating that the transport (influx and efflux) and metabolic conversion are under kinetic coordination. The resistant cells were found to proliferate markedly faster than the TAOV or MDR-1 gene-inserted cells. Some clones derived from TR-21 cells were insensitive to the treatment with 8-azaguanine (8- AZ), whereas 8-AZ killed all of the TAOV cells. About 50% of the 8-AZ- resistant clones were resistant to a combined treatment of azaserine/hypoxanthine, suggesting that taxol resistance and nucleotide synthesis through de novo and/or salvage pathways are closely interrelated.
