Mismatch Repair Functions Affected During Tumorigenesis

简介(申请人提供)：结肠癌是影响男性和女性的第三种最常见的癌症，也是癌症死亡的第二大原因。最常见的疾病是遗传性非息肉病性结肠癌(HNPCC)，它是由DNA错配修复(MMR)基因突变引起的。此外，在10%-30%的散发性结直肠癌、子宫内膜癌和其他癌症中也检测到MMR缺陷。这项提案将解决为什么MMR途径缺陷会导致肿瘤发生的广泛问题。MMR系统纠正由于DNA聚合酶错误、异源序列之间的重组或内源和外源诱变剂之间的重组而导致的错配或损坏的DNA。因此，MMR功能的丧失会导致突变率升高(突变子表型)。然而，MMR系统也是激活细胞周期检查点和细胞凋亡以应对DNA损伤所必需的。我们推测，这种依赖于MMR的检查点/凋亡反应在肿瘤保护中发挥着重要作用，因为它的破坏将允许基因组不稳定性增加的细胞不受控制地生长。基于这一假设的预测是，肿瘤中含有MMR基因突变，破坏了损伤修复和检查点/凋亡反应功能。我们将通过确定MMR基因hMSH2和hMSH6的癌症相关错义突变是否会影响DNA修复和检查点/细胞凋亡反应来验证这一预测。 我们建议3旨在评估与癌症相关的hMSH2和hMSH6错义突变的功能后果。我们之前已经详细说明了与HNPCC相关的七个hMSH2错义突变对hMSH2-hMSH6异源二聚体生化功能的影响。在目标1中，我们将研究与癌症相关的hMSH6错义突变的生化效应。在目标2中，我们将设计一种体外实验来研究hMSH2-hMSH6异源二聚体如何与参与修复和检查点/凋亡信号的其他蛋白质相互作用，以及与癌症相关的错义突变是否影响这些相互作用。最后，在目标3中，我们将在人类细胞培养模型中检测hMSH2和hMSH6错义突变对DNA修复和检查点/凋亡功能的影响。通过这种生物化学和细胞生物学方法的彻底整合，我们将利用致癌错义突变作为工具，进一步了解MMR的分子机制以及这些机制在肿瘤发生过程中是如何被破坏的。结肠癌是男性和女性第三常见的恶性肿瘤，在癌症死亡人数中仅次于肺癌。错配修复途径基因的遗传突变会导致遗传性非息肉病性结肠癌，而在普通人群中，10%-30%的结直肠癌、子宫内膜癌和其他癌症也会检测到这一途径的缺陷。在分子水平上理解为什么这一途径的改变促进肿瘤发生将有助于开发更有针对性的治疗、预防和诊断策略。

DESCRIPTION (provided by applicant): Colon cancer is the third most common cancer affecting men and women and the second leading cause of cancer death. The most common disease predisposing patients to colorectal cancer is hereditary non-polyposis colon cancer (HNPCC) which stems from mutations in DNA mismatch repair (MMR) genes. In addition, MMR defects are also detected in 10-30% of sporadic colorectal, endometrial and other cancers. This proposal will address the broad question of why defects in the MMR pathway cause tumorigenesis. The MMR system corrects mispaired or damaged DNA resulting from DNA polymerase errors, recombination between heterologous sequences or endogenous and exogenous mutagens. Thus, loss of MMR function results in an elevated mutation rate (mutator phenotype). However, the MMR system is also required for the activation of cell cycle checkpoints and apoptosis in response to DNA damage. We hypothesize that this MMR-dependent checkpoint/apoptosis response plays a significant role in tumor protection as its disruption would allow cells with increased genomic instability to grow unchecked. A prediction based on this hypothesis is that tumors harbor mutations in MMR genes that disrupt damage repair and checkpoint/apoptosis response functions. We will test this prediction by determining whether cancer-associated missense mutations of the MMR genes hMSH2 and hMSH6 affect both DNA repair and checkpoint/apoptosis response. We propose 3 aims to evaluate the functional consequences of cancer- associated missense mutations in hMSH2 and hMSH6. We have previously detailed the effects of seven hMSH2 missense mutations associated with HNPCC on the biochemical functions of the hMSH2-hMSH6 heterodimer. In Aim 1, we will examine the biochemical effects of cancer-associated missense mutations in hMSH6. In Aim 2, we will devise an in vitro assay to examine how the hMSH2-hMSH6 heterodimer interacts with other proteins involved in repair and checkpoint/apoptosis signaling and whether cancer-associated missense mutations affect these interactions. Finally, in Aim 3 we will examine the effects of missense mutations in hMSH2 and hMSH6 on DNA repair and checkpoint/apoptosis functions in human cell culture models. Through this thorough integration of biochemical and cell biology approaches we will utilize cancer-causing missense mutations as tools to further understand the molecular mechanism of MMR and how those mechanisms are disrupted during tumorigenesis.Colon cancer is the third most common malignancy in men and women and ranks behind only lung cancer in cancer deaths. Inherited mutations in genes of the mismatch repair pathway cause the disease hereditary non-polyposis colon cancer, while defects in this pathway are also detected in 10-30% of colorectal, endometrial and other cancers in the general population. Understanding at the molecular level why alterations in this pathway promote tumorigenesis will aide the development of more targeted therapeutic, preventive and diagnostic strategies.