ANTIMETABOLITES AND NUCLEAR MATRIX-BOUND DNA

最近的证据支持这样的概念，即在真核细胞中， 复制附着在核基质上的固定位点上， 模板DNA在复制时通过这些位点。 由于许多 用于癌症化疗的抗代谢物诱导链断裂， 片段化，以及DNA的其他改变，这些似乎是合理的， 代理人可能有直接和重要的影响， 核基质上的模板DNA和DNA复制酶。 然而，在这方面， 在大多数情况下，这些潜在的机制， 抑制DNA复制仍然没有被探索。 拟议的研究是 旨在揭示5-氟-2 '-脱氧尿苷（FdUrd）、5，8- 二脱氮异叶酸（IAHQ）或替尼泊苷（VM-26）特异性抑制 人CCRF-CEM白血病细胞中核基质结合的DNA复制，和 无论这种抑制是否由DNA结合受损引起 聚合酶α和DNA引发酶与细胞核相关的新生DNA 矩阵 将进行研究，以确定是否有直接的 核基质结合DNA的抑制程度之间的关系 复制，DNA单链断裂的形成，以及 菌落形成软琼脂糖。 这些抗癌药物对 核基质结合的DNA复制也将在一定条件下进行检查 其中所有脱氧核糖核苷的细胞内浓度 在未处理的细胞中，使三磷酸盐相等并饱和用于DNA合成。 和L-溶血素透化后药物处理的CCRF-CEM细胞。 计划进行更多的研究，以确定FdUrd、IAHQ或VM-26 通过干扰DNA合成来抑制新生DNA的延伸， 通过干扰DNA固定新生DNA延伸的位点 在核基质上的固定位点合成。 将采取各种办法， 为了验证这些抗癌药物最终阻断 核基质结合的DNA复制通过干扰的协会 DNA复制酶与核基质。 在这些阿尔法和DNA中 核基质部分中的引发酶，以及DNA聚合酶的结合 在对照组和药物组中，α- DNA引发酶与新复制的DNA形成复合物， 处理的细胞。 对这一假设的进一步检验将涉及定量 这些抗代谢物对RNA引发的比活性的任何影响 核基质上的新生DNA 这些结果可能具有新颖性， 对更清楚地了解机制的重要影响， 这些抗代谢物抑制DNA复制。

Recent evidence supports the concept that in eucaryotic cells DNA replication are attached to fixed sites on the nuclear matrix, and that template DNA passes through these sites as it is replicated. Since many antimetabolites used in cancer chemotherapy induce strand-breaks, fragmentations, and other alterations in DNA, it seems reasonable that these agents might have direct and important effects on the interactions of template DNA and DNA replication enzymes on the nuclear matrix. However, for the most part these potential mechanisms by which antimetabolites might inhibit DNA replication have remained unexplored. The proposed studies are designed to reveal if either 5-fluoro-2'-deoxyuridine (FdUrd), 5,8- dideazaisofolic acid (IAHQ),or teniposide (VM-26) specifically inhibit nuclear matrix-bound DNA replication in human CCRF-CEM leukemia cells, and whether or not this inhibitation results from impaired binding of DNA polymerase alpha and DNA primase to nascent DNA associated with the nuclear matrix. Studies will be carried out to determine if there is a direct relationship among the degree of inhibition of nuclear matrix-bound DNA replication, the formation of DNA single-strand breaks, and the inhibition of colony formation soft agarose. The effects of these anticancer agents on nuclear matrix-bound DNA replication will also be examined under conditions in which the intracellular concentrations of all the deoxyribonucleoside triphosphates are made equal and saturating for DNA synthesis in untreated and drug-treated CCRF-CEM cells following L-lysolecithin permeabilization. Additional studies are planned to determine if either FdUrd, IAHQ, or VM-26 inhibit the elongation of nascent DNA by interfering with DNA synthesis at fixed sites on the elongation of nascent DNA by interfering with DNA synthesis at fixed sites on the nuclear matrix. Various approaches will be taken to test the hypothesis that these anticancer agents ultimately block nuclear matrix-bound DNA replication by interfering with the association of DNA replication enzymes with the nuclear matrix. In these alpha and DNA primase in the nuclear matrix fraction, and on the binding of DNA polymerase alpha - DNA primase complexes to newly replicated DNA in control and drug- treated cells. Further testing of this hypothesis will involve quantitating any effects of these antimetabolites on the specific activity of RNA-primed nascent DNA on the nuclear matrix. These results could have novel and important implications for more clearly understanding the mechanisms by which antimetabolites inhibit DNA replication.