RETROVIRAL INSERTIONAL MUTAGENESIS AS A STRATEGY FOR THE IDENTIFICATION OF GENES ASSOCIATED WITH CIS-DIAMMINEDICHLOROPLATINUM(II) RESISTANCE

Expression of resistance to cis-diamminedichloroplatinum(II) (CDDP), o ne of the most effective chemotherapeutic drugs used to treat a variet y of malignancies, remains a serious obstacle for improving cancer tre atment. To study possible genetic mechanisms underlying the developmen t of CDDP resistance, we have adopted the approach of retroviral inser tional mutagenesis. An early-stage CDDP-sensitive human melanoma cell line, WM35, was infected with a defective amphotropic murine retroviru s (murine stem cell virus), and the pooled cells were subsequently sel ected for CDDP-resistant variants. Nine CDDP-resistant clones independ ently derived from murine stem cell virus infected WM35 cells were ana lyzed and it was found that five of these clones acquired an identical retroviral integration site, designated as CDDP resistance locus 1 (C RL-1), as revealed by isolation of retroviral flanking sequences. Furt hermore, using the flanking sequence as probe, we have detected a 3.5- 4.0-kilobase message, the expression of which is strongly increased in clones carrying a rearranged CRL-1 locus. These results strongly sugg est that overexpression of CRL-1 confers resistance to CDDP in these c lones. In addition, the present study indicates that retroviral insert ional mutagenesis represents a potential strategy to identify genes re sponsible for CDDP resistance and possibly other chemotherapeutic drug s as well.