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 特别容易发生突变，因为潜在的遗传 不稳定性和DNA修复缺陷。突变导致 化疗药物耐药性和肿瘤进展可能会出现 并限制有效治疗。然而，人们对 化疗混合物诱导肿瘤突变的能力 高于先前存在的背景频率。突变诱导可能是 在乳腺癌和其他肿瘤中尤其重要， 化疗最初，但最终成为耐药。 本项目的目标是确定1）突变是否 联合化疗的效果可以降低， 细胞毒性和2）诱变化疗是否诱导突变 导致肿瘤细胞自发产生耐药性 背景频率具体目标是：（1）确定是否 不同类型突变的诱导作为以下函数而变化： 化疗药物混合物和/或时间表 使用常规突变测定的细胞毒性，2）确定药物 混合和/或调度也影响不同的诱导， 乳腺癌细胞系中的突变类型，和3）相关药物 诱导突变，同时诱导稳定的蒽环类抗生素 乳腺癌的耐药性及其耐药机制 细胞系这些研究的长远目标是改善 使用常规化疗，通过减少有害的 突变的副作用 联合用药会诱发突变和耐药性 通常用于乳腺癌，根据不同的时间表给予， 混合物。将在tk（胸苷）时分析突变频率 激酶）和CAD（氨甲酰-P合酶，天冬氨酸转氨甲酰酶和 二氢乳清酸酶）酶基因座的TK 6人淋巴母细胞和修饰的 具有正常或突变p53的人乳腺癌细胞系。的频率 对阿霉素产生耐药性的乳腺癌细胞 用不同的突变诱导药物方案和混合物治疗 也将被衡量。耐药机制将在 通过表型分析的耐药细胞的代表性子集，单- 链构象多态性（SSCP）、斑点印迹分析和rtPCR。

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.