A novel resistant variant of murine P388 leukaemia, P388/SPR, was identified by de novo resistance to doxorubicin (DOX) in vivo. This mutant displayed a similar level of cross-resistance to etoposide (VP-16) and other topoisomerase II (topo II) inhibitors. Further analysis of the phenotype revealed a broad cross-resistance to vinca alkaloids, alkylating agents, antimetabolites, aphidicolin and UV light. Low-level expression of mdr1 and P-glycoprotein (P-gp), as well as a modest impairment of cellular drug accumulation and partial reversion of resistance to DOX and VP-16 by cyclosporine, confirmed a moderate role of P-gp in conferring drug resistance in P388/SPR cells. Consistent changes in neither topo II expression or activity nor glutathione metabolism could be detected. Induction of apoptosis was significantly reduced in P388/SPR cells, as indicated by minimal DNA fragmentation. Analysis of oncogenes regulating apoptotic cell death revealed a marked decrease of bcl-2 in combination with a moderate reduction of bax protein, but a striking overexpression of the long form of the bcl-X protein. Transfection of human bcl-X-L into P388 cells conferred drug resistance similar to that of P388/SPR cells. The data suggest that overexpression of bcl-X-L results in an unusual phenotype with broad cross-resistance to non-MDR-related cytotoxins in vitro, and provide an interesting example of spontaneous overexpression of another member of the bcl-2 gene family in cancer.