Inducible colon-specific transgenic mouse for cancer research

描述（由申请人提供）：在过去的20年里，我们对结肠癌分子病因学的理解已经急剧扩展。尽管如此，结肠癌仍然是美国的一个重大健康问题。该领域进展的障碍之一是缺乏概括人类结肠癌病因学的良好表征的动物模型。虽然我们已经从各种化学诱导或遗传编程的啮齿动物模型产生的癌症中学到了很多，但在这些模型中发展的癌症在潜伏期，肠道位置或分子特征方面通常与人类结肠癌显着不同。机制和预防为重点的结肠癌研究需要发展的动物模型，允许精确定时，结肠特异性修改肠道生物学。我们所提出的研究的目标是使用驱动碳酸酐酶1（CA 1）基因的大的精氨酸特异性表达的启动子来创建具有Cre重组酶（Cre）的结肠上皮细胞特异性表达的转基因小鼠，所述Cre重组酶（Cre）融合至仅对雌激素类似物他莫昔芬（CAC-ERT 2）具有高亲和力的经修饰的雌激素受体（ER）配体结合结构域。融合基因的ERT 2部分赋予转基因以泰索米芬诱导性，并允许小鼠基因组中floxed等位基因的时间控制性缺失。一个转基因载体已经产生，它将被用来生产在普渡转基因小鼠核心设施的小鼠。将通过PCR评估转基因表达水平和组织分布。将CAC-ERT 2小鼠与ROSA 26 R指示小鼠杂交后，评估他莫昔芬对转基因功能的诱导。将在与具有一个或两个floxed APC等位基因的小鼠杂交的CAC-ERT 2小鼠中评估他莫昔芬诱导肿瘤形成的能力。将在存在葡聚糖硫酸钠（DSS）诱导的结肠炎症的情况下进行其他研究。该项目完成后，CAC-ERT 2小鼠将成为唯一的转基因模型，既限制Cre表达大肠上皮细胞，并允许控制Cre功能时删除floxed等位基因。这将允许结肠癌研究人员容易地组合联合收割机基因突变（即与结肠癌病因相关的多个floxed等位基因）并在生命的任何阶段启动结肠癌。这将允许更仔细地评估致癌机制，并改进成年小鼠中化学预防或治疗剂的测试。虽然我们的兴趣是将该模型用于结肠癌，但该模型也将对对下肠道炎症性疾病（即IBD、克罗恩病）感兴趣的研究人员有用。1 公共卫生相关性：结肠癌在美国和科学家中是一种常见的癌症。由于缺乏能再现人类散发性结肠癌特征的动物模型，开发有效治疗和预防策略的研究受到阻碍。我们提出的研究将创造一种新的转基因小鼠模型，克服现有结肠癌研究动物模型的局限性。

DESCRIPTION (provided by applicant): Over the past 20 years our understanding of the molecular etiology of colon cancer has expanded dramatically. Despite this, colon cancer remains a significant health problem in the US. One of the barriers to progress in the field is the lack of well-characterized animal models that recapitulate the etiology of human colon cancer. While we have learned a great deal from cancers resulting from various chemically induced- or genetically programmed-rodent models, the cancer that develops in these models is often significantly different from human colon cancer in terms of latency, intestinal location, or molecular signature. Mechanistic and prevention focused colon cancer research requires the development of animal models that permit precisely timed, colon-specific modification of intestinal biology. Our goal for the proposed research is to use the promoter that drives large intestine-specific expression of the carbonic anhydrase 1 (CA1) gene to create a transgenic mouse with colon-epithelial cell-specific expression of Cre recombinase (Cre)-fused to a modified estrogen receptor (ER) ligand binding domain with high affinity only for the estrogen analog tamoxifen (CAC- ERT2). The ERT2 portion of the fusion gene confers taxomifen-inducibility to the transgene and permits temporally controlled deletion of floxed alleles in the mouse genome. A transgene vector has been produced and it will be used to produce mice at the Purdue Transgenic Mouse Core Facility. Transgene expression level and tissue distribution will be assessed by PCR. Inducibility of transgene function by tamoxifen will be assessed after crossing the CAC-ERT2 mouse to the ROSA26R indicator mouse. The ability of tamoxifen to induce tumor formation will be assessed in CAC-ERT2 mice crossed to mice with one or two floxed APC alleles. Additional studies will be done in the presence of colonic inflammation induced by dextran sulfate sodium (DSS). Upon completion of the project, the CAC-ERT2 mouse will be the only transgenic model that both limits Cre expression to the epithelial cells of the large intestine and permits control over when Cre functions to delete floxed alleles. This will permit colon cancer researchers to easily combine genetic mutations (i.e. with multiple floxed alleles relevant to colon cancer etiology) and initiate colon cancer at any stage of life. This will permit more careful assessment of mechanisms of carcinogenesis and improved testing of chemopreventative or therapeutic agents in adult mice. While our interests are in using this model for colon cancer, this model will also be useful for researchers interested in inflammatory conditions of the lower bowel (i.e. IBD, Crohn's disease). 1 PUBLIC HEALTH RELEVANCE: Colon cancer is a common form of cancer in the US and scientists. Research to develop effective treatment and prevention strategies is hampered by the lack of animal models that reproduce the features of human sporadic colon cancer. Our proposed research will create a new genetically modified mouse model that overcomes the limitations of existing animal models for colon cancer research.