TUMOR CELL DEATH INDUCED BY INHIBITION OF REPLICATION

描述：许多用于癌症化疗的细胞毒性药物抑制DNA 复制并诱导肿瘤细胞中的程序性细胞死亡（凋亡）。 对DNA复制本身的抑制程度尚不清楚 负责抗癌药物诱导细胞死亡。 细胞 可以通过用已知的抑制剂处理来诱导循环细胞的死亡 DNA复制，如aphidicolin，一种真菌毒素， 通过DNA聚合酶α和δ延长复制，或 含羞草素，一种植物氨基酸，可阻止复制起始， 哺乳动物细胞这个应用程序的主要目标是识别基因 以及参与诱导细胞死亡的遗传途径 DNA复制的抑制剂。 这些基因将通过 基因抑制元件（GSE）的表达选择，短cDNA 抑制基因表达的片段，通过编码显性作用于 肽或抑制性反义RNA。 GSE的标准化文库， 将逆转录病毒载体转导入HeLa细胞， 保护细胞免受蚜虫霉素或含羞草素诱导的细胞死亡将是 与世隔绝这些GSE将根据其功效、效果 对细胞周期进程的影响，保护细胞免受不同DNA 复制抑制剂和细胞类型特异性。 全长cDNA 将分离对应于所选GSE的人类基因，并且 它们在细胞周期中的结构和表达模式将被 表征了 孤立的GSE也将被测试其影响 关于细胞对复制抑制化疗药物的反应， 以确定细胞毒性的复制机制在 其抗肿瘤作用。

DESCRIPTION: Many cytotoxic drugs used in cancer chemotherapy inhibit DNA replication and induce programmed cell death (apoptosis) in tumor cells. It is unknown to what extent the inhibition of DNA replication per se is responsible for the induction of cell death by anticancer drugs. Cell death in cycling cells can be induced by treatment with known inhibitors of DNA replication, such as aphidicolin, a mycotoxin which inhibits elongation of replication by DNA polymerases alpha and delta, or mimosine, a plant amino acid which prevents initiation of replication in mammalian cells. The main goal of this application is to identify genes and genetic pathways that are involved in the induction of cell death by inhibitors of DNA replication. These genes will be identified through expression selection of genetic suppressor elements (GSE), short cDNA fragments that inhibit gene expression by encoding dominantly acting peptides or inhibitory antisense RNA. A normalized library of GSEs in a retroviral vector will be transduced into HeLa cells, and GSEs which protect cells from aphidicolin- or mimosine-induced cell death will be isolated. These GSEs will be characterized for their efficacy, effects on cell cycle progression, ability to protect cells against different DNA replication inhibitors and cell-type specificity. Full-length cDNAs for human genes corresponding to the selected GSEs will be isolated, and their structure and expression patterns in the cell cycle will be characterized. The isolated GSEs will also be tested for their effects on cellular response to replication-inhibiting chemotherapeutic drugs, to determine the role of replication-based mechanisms of cytotoxicity in their antitumor effect.