A major cause of ineffective results in the chemotherapeutic treatment of human neoplasms is the development of clones of resistant cells, which are able to circumvent drug effects and repopulate a tumor. Cellular heterogeneity of tumor cells predicates that both genotypic and phenotypic variants will be present in a typical tumor. Cellular heterogeneity of tumor cells predicates that both genotypic and phenotypic variants will be present in a typical tumor. Such mutants are selected by drug treatment. By comparing an established Walker 256 rat carcinoma cell line resistant to bifunctional nitrogen mustards, but sensitive to nitrosoureas (WR) with its parent cell line (WS), and by developing other such resistant lines, this project is designed to achieve the following goals: (1) compare the nature of consistuent thiols in WR and WS (2) determine by quantitative and qualitative analysis of gluathione transferase enzymes if specific differences in the WR cells can account for their increased resistance to alkylating electrophiles (3) using nitroso-ureas with variable carbamoylating activity, determine if there are differential effects on the glutathione reductase isoenzyme pattern of WR and WS (a two-fold decrease in reductase activity has been found in WR and these cells have been found to be sensitive to carbamoylating agents) (4) using previously characterized nuclear lamin protein antibodies, determine if the extra nuclear matrix polypeptide band, found by SDS-gel electrophoresis in WS cells, is a novel protein or a post-translational modification of an existing matrix polypeptide (5) consider the nuclear matrix proteins as specific targets for alkylating agents and nitrosoureas by using labelled drugs and gel autoradiography (6) determine if there are differences in the effect of alkylating agents and nitrosoureas on poly(ADP-ribose) polymerase and DNA ligase, two enzymes with putative roles in repair of drug damage to DNA (7) develop cell lines with increased degrees of resistance to both nitrogen mustards and nitrosoureas, by exposing WR cells to escalting drug concentrations and selecting surviving cells in soft agar. Karyotypic screening of selected resistant lines may permit conclusions concerning general mechanisms of acquired resistance in selected cell lines and may assist in correlating changes in nuclear structure with the phenotypic expression of acquired resistance to nuclear reacting drugs.
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