MOLECULAR STUDIES OF ESOPHAGEAL ADENOCARCINOMA

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The goals of this continuation application are to identify important, and potentially novel amplified cancer-related genes in esophageal adenocarcinomas. This project involves the collaboration of investigators with substantial expertise in cancer and molecular genetics and focuses upon esophageal adenocarcinoma, a deadly cancer which is increasing in incidence. The identification of novel genomic amplifications, and the resulting important genes overexpressed in these tumors, will provide significant insight into the molecular events associated with the development and progression of this disease. This work may provide new genetic tools for early detection, and potentially for identifying new avenues for therapeutic intervention. We have identified a number of potentially novel genomic amplification events in esophageal adenocarcinomas using the highly sensitive method of quantitative two-dimensional genomic scanning (2D gels). This technology involves the comparative analysis of several thousand genomic restriction fragments derived from functional components of the genome between the normal and tumor tissue from an individual patient. The amplified and overexpressed genes encompassed in the amplicons will be identified and characterized, using the procedures we have successfully demonstrated for a novel amplicon in esophageal adenocarcinomas. 2D gels are particularly well suited for the identification of amplified regions of DNA. Amplified fragments will be cloned using procedures that we have successfully implemented and their chromosomal locations mapped using fluorescence in situ hybridization and/or database searches. Utilizing an innovative quantitative PCR assay and over 200 adenocarcinomas collected to date, the "minimal region of common amplification" will be determined. This will effectively define the region to search for important amplified and overexpressed cancer-related genes. Potential candidates may include known genes, uncharacterized expressed sequence tags, or genes isolated from genomic clones that map within the minimal region. The cancer-relatedness of novel genes will be examined using functional assays involving transfection into immortalized cells and analysis of the effects upon cell growth, invasiveness, anchorage-independence and tumor formation in mice. These studies will determine both the identity and frequency of specific genes amplified in esophageal adenocarcinomas and also provide a greater insight into the mechanisms underlying the development and/or progression of this important cancer.