Gastrointestinal cancer is a significant health problem, ranking fourth in incidence and second in death in the United States. Abnormal differentiation and increased proliferation are hallmarks of carcinogenesis. The molecular mechanisms that regulate cellular proliferation and differentiation in gastrointestinal development are far from being understood completely. Here we focus on the homeobox-transcription factors Cdx1 and Cdx2, which are primer candidates as central regulators of both intestinal development and GI carcinogenesis. While several hundred articles have appeared that suggest these genes as master regulators of early gut development and GI cancers, the definitive experiment, that is the analysis intestinal epithelia deficient for Cdx1, Cdx2 or both has not yet been performed. Therefore, we will develop and exploit new genetic tools to elucidate fundamental questions regarding the function of these two genes. In addition, in an innovative approach we will use the 1.8 million monoclonal crypts of the mouse intestine as a high- throughput """"""""laboratory"""""""" to identify novel proto-oncogenes, tumor suppressors, and cancer modifiers by transposon-mediated insertional mutagenesis.
Specific Aims :
In Aim 1, we will determine the function of Cdx genes in anterior-posterior patterning of gut endoderm. We will test our hypothesis that Cdx1 and Cdx2 are critical mediators of anterior-posterior patterning of the primitive gut by analyzing the development of early gut endoderm deficient for Cdx1, Cdx2 or Cdx2/Cdx1, which we will obtain by conditional gene ablation in mice.
In Aim 2, we will analyze the function of Cdx genes in intestinal differentiation and tumorigenesis. We hypothesize that Cdx1 and Cdx2 are critical regulators of intestinal differentiation, and that Cdx2 acts as a tumor suppressor in the intestinal epithelium. We will evaluate our hypotheses by deriving mice deficient for Cdx1, Cdx2 (or Cdx2/Cdx1) after cytodifferentiation has occurred. Mice will be analyzed for the elaboration of the mature intestinal cell types, for their rate of proliferation, and their propensity for tumorigenesis in multiple models.
In Aim 3, we will employ a novel genetic screen to uncover novel proto-oncogenes and tumor suppressors in colorectal cancer. We will utilize the innovative and powerful approach of retrotransposon-driven random insertional mutagenesis to mutagenize the 1.8 million crypts of the mouse intestine. The resulting polyps will be characterized histologically, and the tumor-causing mutation sequenced. Novel proto-oncogenes and tumor suppressor genes will be screen for mutations in human colorectal cancer. Together, this proposal will further our understanding of GI development and cancer, and allow for the development of novel diagnostic and possibly therapeutic tools in the future.
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