GENE EXPRESSION CHANGES IN METASTATIC COLON CANCER

Beth Israel Deaconess Medical Center, Boston MA

Investigators

Abstract

The most life-threatening aspects of colon cancer are invasion and metastasis. Genetic alterations likely play a crucial role in the pathogenesis of colon cancer metastasis. Therefore, one of the most important issues in cancer research is to understand the genetic alterations in metastasis. The goal of this application is to analyze gene expression changes in metastatic colon cancer. By using differential display, we have identified 19 genes expressed differentially between primary and metastatic colon cancer. In preliminary studies, we found that a novel gene called 055 is not expressed in metastatic colon cancer tissues or a metastatic colon cancer cell line (SW620). Overexpression of 055 in SW620 increased expression of several cell differentiation markers such as E-cadherin, CEA and alkaline phosphatase and induced morphological changes consistent with cell differentiation. The expression of 055 is upregulated by several known cell differentiation reagents. Our preliminary studies show that overexpression of 055 in SW620 colon cancer cells suppressed in vitro invasion into matrigel and in vivo liver metastasis in nude mice. Our workings hypothesis is that the 055 gene suppresses metastasis by inducing colon cancer cell differentiation and reversing the malignant phenotype. Therefore, we propose: (1) to investigate the expression and function of 055 protein in normal and neoplastic colonic epithelial cells. This will include studying: (a) subcellular localization of the 055 gene in colon cancer cell lines; the expression of 055 protein in normal colon tissues, adenomas, primary and metastatic colon cancers by immunohistochernistry. (b) the in vitro and in vivo adhesion and invasion properties of metastatic colon cancer cells transfected with 055 gene, or primary colon cancer cells transfected with 055 antisense DNA; (c) the mechanism of 055 in regulation of E-cadherin expression and cell differentiation. (2) to study the mechanism of aberrant regulation of 055 gene in metastaric colon cancer. This will be accomplished by: (a) nuclear run-off assay to determine the rate of transcription. Should run-off assay and promoter studies fail to detect difference in rate of transcription between primary and metastatic colon cancer cells, a mRNA decay assay to assess the mRNA stability will be performed. (B) cloning and analyzing the promoter region of the 055 gene. Chloramphenicol acetyltransferase (CAT) gene assay will be used to assess the activities of normal 055 promoter in both primary and metastatic colon cancer cells. (c) Analyzing the role of DNA methylation in regulation of 055 expression.