DECREASED LEVELS OF TOPOISOMERASE II-ALPHA IN HUMAN RENAL-CELL CARCINOMA LINES RESISTANT TO ETOPOSIDE

Renal cell carcinoma (RCC) displays strong resistance against many che motherapeutic drugs, Overexpression of P-glycoprotein (Pgp) appears to be part of this resistance. The involvement of another resistance mec hanism, involving the decreased activity of DNA topoisomerase II (topo II), remains uncertain. By culturing the human RCC lines RC2 and RC21 in the presence of increasing concentrations of etoposide, we derived the valiant sublines RC2E, RC21A and RC21E, that had acquired approxim ately 30-, 60- and 90-fold resistance to this drug respectively. RC2E, RC21A and RC21E were approximately 50-, 5- and 400-fold cross-resista nt to doxorubicin respectively. RC2E and RC21E also showed cross-resis tance (approximately 200- and 3500-fold respectively) to vinblastine. Quantitative differences in MDR1 and Pgp expression (elevated in RC2E and RC21E) and topoII alpha (reduced in RC21E and RC21A) were demonstr ated using Western blotting and the reverse transcriptase/polymerase c hain reaction. Decreased amounts of topoII alpha were reflected in a r educed activity of RC21A and RC21E as measured by unknotting phage P4 DNA. Qualitative changes of the topoII alpha gene, such as point mutat ions in the motif B/DNBS and DNA-binding regions, or differences in me thylation status of the promoter gene of RC21E, were not found. These cell lines represent a model of a solid tumor in which overexpression of Pgp, a combination of increased Pgp and decreased topoII alpha, and a decrease of topoII alpha are represented.