This research represents an investigation of the mechanisms of action of the cancer chemotherapeutic platinum coordination complexes, as well as an investigation of the mechanism of cellular sensitivity and resistance. Three murine leukemia L1210 cell lines will be used: a) L1210/0, which is sensitive to the action of both cis-diamminedichloroplatinum(II) (cis-DDP) and 1,2- diaminocyclohexaneplatinum (DACH-Pt) analogues, b) L1210/DDP, which is preferentially resistant to cis-DDP, c) L1210/DACH, which is preferentially resistant to DACH-Pt. The resistant cells exhibit some reduced accumulation of cis-DDP, but the majority of resistance is attributed to enhanced DNA repair. Two different assays of DNA repair have been used, one measuring direct removal of DNA bound adducts, the other measuring the ability of cells to express a transfected damaged gene. The latter assay demonstrated that one adduct was normally sufficient to inhibit transcription, suggesting this could contribute to the mechanism of toxicity rather than inhibition of DNA synthesis previously suggested.
The aims are, therefore, to further characterize the DNA repair process. using DNA repair inhibitors and probe for repair of specific genes. Attempts will be made to purify the protein(s) involved. An assay for inhibition of transcription on a defined gene is proposed, which will be compared to DNA repair of adducts within the same gene. Recombination, as a mechanism for replicative bypass of DNA adducts will also be investigated. In other experiments, attempts will be made to identify the chromosome(s) coding for resistance by using chromosome-mediated gene transfer. This is a preliminary to isolating the genes in- volved. Parallel studies will be performed to assess the mechanism of resistance to DACH-Pt in L1210/DACH and also to assess the mechanisms of hypersensitivity to cis-DDP in embryonal cells. These cells are representative of tumors that are very responsive to cis- DDP therapy.
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