DELETERIOUS MUTATIONS FROM COMBINATION CHEMOTHERAPY

几乎所有用于治疗癌症的药物都会导致突变。癌细胞是 特别容易发生突变，因为潜在的遗传 不稳定和DNA修复缺陷。突变是导致 可出现化疗耐药和肿瘤进展 自发的，并限制有效的治疗。然而，人们对此知之甚少。 化疗混合物诱导肿瘤突变的能力 高于预先存在的背景频率的细胞。突变诱导可能是 对乳腺癌和其他对药物有反应的肿瘤尤其重要 最初是化疗，但最终产生了抗药性。 该项目的目标是确定1)突变 联合化疗的效果可以在不牺牲的情况下减少 细胞毒性和2)诱变化疗是否会导致突变 在自发产生的肿瘤细胞中引起耐药性 背景频率。具体目标是：1)确定是否 不同类别的突变的诱变是不同的 同等水平的化疗药物混合物和/或时间表 使用常规突变检测的细胞毒性，2)确定药物 混合和/或时间安排也会影响不同 乳腺癌细胞系的突变类型，以及3)相关药物 用稳定的蒽环类药物诱导诱变 乳腺癌中的耐药性及其机制 细胞系。这些研究的长远目标是改善 通过减少有害物质来使用常规化疗 突变的副作用。 联合用药会导致突变和抗药性 通常用于乳腺癌，根据不同的时间表和 混合物。突变频率将在tk(胸苷)进行分析 激酶)和CAD(氨基甲酰-P合成酶、天冬氨酸转氨酶和 TK6人淋巴母细胞二氢酶基因座及其修饰 P53正常或突变的人乳腺癌细胞株。这些频率 对阿霉素产生抗药性的乳腺癌细胞 用不同的诱变药物方案和混合物进行治疗 也将被测量。抗药性的机制将在 通过表型分析，抗性细胞的代表性亚群，单个- 链构象多态(SSCP)、斑点印迹分析和RT-PCR。

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.