MOLECULAR PATHOGENESIS OF CANCER OF MUTATOR PHENOTYPE

在此优秀奖所涵盖的时间内，我们已按计划在竞争性续展申请中继续进行研究项目。我们的具体目标(SA？)有三个，每个都有三个子项目。第一个子项目一般是描述性的，第二个子项目一般是机械性的，第三个子项目代表了先前子项目中的实验在癌症预后方面的应用。具体地说，按照优先顺序，我们将通过几种方法测试累积单倍体充分性模型，包括分离单倍体充分性候选基因突变的单细胞克隆，使用与展示Bax和MSH6突变功能相同的策略。这将得到另一种方法的补充，该方法使用不同基因杂合的基因敲除小鼠的杂交。我们还将通过使用MS-AFLP DNA指纹对全基因组DNA甲基化改变进行无偏见的分析，来检验甲基化表型作为突变子表型表现的先前致病事件的假设。我们还将继续寻找与一些HNPCC和散发性MMP癌有关的新的突变子基因，以及MSH5的筛选和功能分析。这项研究的翻译方面(预测应用)将以累积的方式继续进行，以进行验证。针对特定目的1，我们提出了微卫星不稳定性肿瘤(MSI)代表结肠癌分子通路的假说，并探讨了微卫星突变表型(MMPs)致癌途径的机制。在特定的目的2中，我们提出了一种假设，即通过参与基因组完整性的多个基因的突变失活，包括错配修复(MMR)基因家族的不同成员，来检验这一假说，并确定其功能后果。在特定的目标3中，我们提出了一种假设，即逃脱细胞凋亡是基质金属蛋白酶途径在肿瘤发生中的关键事件，并分析其潜在的机制。

During the period covered by this merit award we have continued the research project as planned in the competitive renewal application. Our specific aims (SA?) were three, each with three subprojects. The first subprojects were generally descriptive, the second generally mechanistic and the third represented the applications for cancer prognosis of the experiments from the previous subprojects. Specifically, by order of priorities, we will test the accumulative haploid sufficiency model for gene inactivation in tumors of the MMP by several approaches, including isolation of single cell clones with mutations in some of the candidate genes for haploid sufficiency, using the same strategy used for the demonstration of the functionality of Bax and MSH6 mutations. This will be complemented with another approach using crosses of knockout mice heterozygous in different genes. We will also test the hypothesis of a methylator phenotype as the preceding causative event for the manifestation of the mutator phenotype, by the unbiased analysis if genome-wide DNA methylation alteration using MS-AFLP DNA fingerprinting. We will also continue the search for novel mutator genes responsible for some of the HNPCC and sporadic cancers of the MMP, and the screening and functional analysis of MSH5. The translational aspects of the research (prognostic applications) will be continued in an accumulative manner with validation purposes. AIMS In specific aim 1 we proposed to test the hypothesis that tumors with microsatellite instability (MSI) represent a distinct molecular pathway for colon cancer, and to investigate the mechanisms underlying this microsatellite mutator phenotype (MMP) tumorigenic pathway. In specific aim 2 we proposed to test the hypothesis that cancer of the MMP unfolds gradually by the mutational inactivation of multiple genes involved in genome integrity, including different members of the mismatch repair (MMR) gene family, and to determine the functional consequences. In specific aim 3 we proposed to test the hypothesis that the escape from apoptosis represented a critical event in tumorigenesis of the MMP pathway and to analyze the underlying mechanisms.