The emergence of drug resistant clones of tumor cells remains one of the major limitations of cancer chemotherapy. Unfortunately, relatively little is known of the genetic mechanisms which may underlie the development of drug resistance. For this reason, we are proposing to examine a model system which may be very useful in studying this phenomena and which may lay the groundwork for mechanistic analysis of transformation associated drug resistance. The central focus of this proposal will be to understand the mechanism(s) generating a high frequency of genomic rearrangements in transformed cells and to determine whether this phenomena is directly related to the generation of drug resistant variants. The underlying framework for this proposal resides in the finding that transformed cells generate drug resistant mutants at a high frequency (1/10000-1/100000) while non-transformed cells (such as primary cultures of fibroblasts or PHA-stimulated lymphocytes) placed under similar drug selection schemes result in a frequency of drug resistant mutants so low as to be undetectable. This proposal will study a model system where expression of a single gene, a temperature sensitive mutation of the v-src gene (LA24) causes non-transformed cells to become transformed. Primary rat fibroblast (RAT-1) have been infected with avian sarcoma virus carrying the LA24 mutation. At the restrictive temperature, the v-src protein is not active and cells grow as a contact inhibited monolayer; at the permissive temperature, v-src is active and cells lose contact inhibition and form foci. Studies are proposed to characterize the association of genomic instability and drug resistance in transformed as compared to non- transformed cells, to find the frequency of various mechanisms of drug resistance in transformed vs. non-transformed cells and, finally, to determine whether specific DNA rearrangements can be implicated in altering the frequency of drug resistant variants.
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