The emergence of drug resistant clones of tumor cells remains one of the major limitations of cancer chemotherapy. Relatively little is known of the genetic mechanisms which may underly the development of drug resistance. Model systems in which tissue culture cells have been selected for a drug resistance phenotype are very useful in the study of this phenomenon. In many instances, selection in vitro with antimetabolites has led to the emergence of drug resistant mutants in which resistance is the consequence of amplification of a specific drug resistance gene. A human colon cancer cell line, WIDR/s, and its mitoxantrone resistant sub-clone WIDR/r are described. Preliminary studies are presented which demonstrate that WIDR/r exhibits cytogenetic evidence of gene amplification, a chromosome 7 HSR. The presence of an amplified domain is verified on renaturation gel analysis, and Southern blotting with a P-glycoprotein probe demonstrates that the amplified domain does not include the P- glycoprotein. A series of experiments is proposed to characterize the amplified gene which may be responsible for resistance to mitoxantrone. We will obtain and characterize cDNA clones of the target gene and use DNA transfection to transfer the resistant phenotype to sensitive cells. These studies will lay the ground work for a mechanistic analysis of mitoxantrone resistance.
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