DELETERIOUS MUTATIONS FROM COMBINATION CHEMOTHERAPY

几乎所有用于治疗癌症的药物都会引起突变。癌细胞是

Virtually all drugs used to treat cancer cause mutation. Cancer cells are especially prone to develop mutations because of underlying genetic instability and DNA repair deficiencies. Mutations responsible for chemotherapeutic drug-resistance and tumor progression can arise spontaneously and limit effective therapy. However, little is known about the capacity of chemotherapeutic mixtures to induce mutations in tumor cells above pre-existing background frequencies. Mutation induction may be particularly important in breast cancer and other tumors which respond to chemotherapy initially, but ultimately become resistant. The goals of this project are to establish whether 1) the mutational effects of combination chemotherapy can be reduced without sacrificing cytotoxicity and 2) whether mutagenic chemotherapy induces mutations causing drug resistance in tumor cells above spontaneously arising background frequencies. The specific aims are to 1) determine whether induction of different classes of mutation varies as a function of chemotherapeutic drug mixture and /or schedule at equivalent levels of cytotoxicity using conventional mutation assays, 2) determine whether drug mixture and/or scheduling also influences the induction of different classes of mutation in breast cancer cell lines, and 3) correlate drug induced mutation with both the induction of stable anthracycline resistance and the mechanisms underlying this resistance in breast cancer cell lines. The long term objectives of these studies are to improve the use of conventional chemotherapy through reduction of deleterious mutational side effects. Mutation and drug resistance will be induced by combinations of drugs typically used for breast cancer given according to varying schedules and mixtures. Mutation frequencies will be analyzed at the tk (thymidine kinase) and CAD (carbamyl-P synthase, aspartate transcarbamylase and dihydroorotase) enzyme loci in TK6 human lymphoblast cells and modified human breast cancer cell lines with normal or mutant p53. The frequencies of breast cancer cells which become resistant to doxorubicin alter treatment with different mutation inducing drug schedules and mixtures will also be measured. Mechanisms of resistance will be analyzed in representative subsets of resistant cells by phenotypic analysis, single- strand conformation polymorphism (SSCP), dot blot assays and rtPCR.