The Environment as a Variable to Calibrate Mouse Models of Human Disease

描述(由申请人提供)：通过外部和内部环境暴露于具有遗传毒性的致癌物会增加人类患主要癌症的风险。这些化学物质与DNA形成共价加合物，被认为创造了一个复杂的遗传变化程序，作为恶变的起始事件，并促进了随后的肿瘤进展。加合物还可以破坏正常基因组修饰的形成或维持，改变受影响组织中的基因表达。因此，影响加合物形成或修复的因素很可能是人类对环境致癌易感性的重要决定因素。我们提出了一种方法来阐明DNA损伤的类型和导致正常细胞向恶性细胞转化的遗传变化的细胞过程。作为一种工具，我们的工作重点是黄曲霉毒素B1(AFB1)，这是一种公认的人类肝细胞癌的危险因素，在与乙肝病毒感染的协同作用中显著增加了风险。与人类肝细胞癌的发病模式一样，AFB1在雄性中比雌性更有效，而且两性动物在青少年时期比成年动物更敏感。我们提出的工作的假设是，不同的突变数量特征(突变频率)或质量特征(突变模式)是肿瘤发生的启动和促进阶段的决定因素。我们工作的目标是表征这些定量和定性特征，并确定它们在调节肝癌发生中的作用。为验证这一假设而设计的实验将采用一种新开发的双重测序协议，该协议能够应用下一代测序平台进行突变分析。在AFB1诱导的小鼠肝癌模型中，将在整个肿瘤形成过程的各个阶段确定选定基因组DNA的突变频率和光谱。这些分析将使我们能够确定在AFB1诱导的肿瘤发展过程中是否获得了“突变表型”，同时提供了它在不同基因之间的变异情况。突变子表型的证据在许多晚期人类肿瘤中都可以看到，并越来越多地被认为是早发药物或辐射耐药的可能来源。我们观察到的突变类型将提供协调肿瘤发生过程中基因变化的细胞过程的特征。我们的动物模型非常适合于研究突变子表型的发病时间，同样重要的是，也适用于其他研究，以调查可能延缓肿瘤发展和耐药性的干预措施。

DESCRIPTION (provided by applicant): Exposure to genotoxic carcinogens via both external and internal environments increases risks for major human cancers. Such chemicals form covalent adducts with DNA, which are thought to create a complex program of genetic change acting as the initiating event in malignant transformation and contributing to subsequent tumor progression. Adducts can also disrupt formation or maintenance of normal genome modifications, altering gene expression in affected tissues. Factors that influence formation or repair of adducts are thus likely to be important determinants of human susceptibility to environmental carcinogenesis. We propose an approach to elucidate types of DNA damage and cellular processes responsible for genetic changes that transform normal cells to malignant ones. As a tool, our work focuses on aflatoxin B1 (AFB1), an established risk factor for human hepatocellular carcinoma that strongly elevates risk in synergy with hepatitis B virus infection. As in the human HCC incidence pattern, AFB1 is more potent in males than females, and animals of both sexes are more sensitive as juveniles than as adults. The hypothesis underlying our proposed work is that different quantitative features of mutagenesis (mutation frequencies) or qualitative features (mutational patterns) are determinants of the initiation and promotion phases of tum origenesis. The goal of our work is to characterize these quantitative and qualitative features and define their roles in modulating liver carcinogenesis. Experiments designed to test this hypothesis will employ a newly developed Duplex Sequencing protocol that enables the application of Next-generation sequencing platforms for mutational analysis. Mutation frequencies and spectra will be determined in selected genomic DNAs at stages throughout the tumorigenic process in a mouse model of AFB1-induced liver cancer. These analyses will enable us to determine whether a "mutator phenotype" is acquired during tumor development induced by AFB1, while providing a picture of its variation among different genes. Evidence of mutator phenotypes is seen in many advanced human tumors, and is increasingly considered as a possible source of premature-onset drug or radiation resistance. The types of mutations that we observe will provide hallmarks of the cellular processes that orchestrate the genetic changes during tumorigenesis. Our animal model is well suited to study the time of onset of a mutator phenotype and, of equal importance, for additional studies to investigate interventions that could delay tumor development and drug resistance.