Dramatic changes in glycolipid metabolism associated with oncogenic transformation implicate a specific role for membrane glycolipids in regulation of cell growth and cellular recognition. These changes give rise to tumor specific membrane antigens which are useful diagnostically and as potential targets for immunotherapy. Although many examples of these tumor antigens have been documented, not much information is available relating to the mechanism of regulation of biosynthesis which prevents expression of these carbohydrate structures in normal cells and tissues and is activated to produce them in association with oncogenesis. A variety of lacto-series based carbohydrate antigens have been described to occur in human colonic adenocarcinomas. Recent evidence has shown that activation of a normally unexpressed beta1->3N-acetyl- glucosaminyltransferase required for lacto-series chain synthesis coupled with activation of type 2 chain synthesis via regulation of enzyme function in Golgi membranes occurs in colonic epithelial cells and results in accumulation of these antigens. This application proposes to continue studies of this system by detailed study of enzyme regulation directing core chain synthesis associated with formation of a variety of end-stage structures which occur in these tumors. The beta1- >3galactosyltransferase which defines the type 1 chain structure and the beta1->3N-acetylglucosaminyltransferase which is activated in association with oncogenesis will be extensively studied. Utilizing expression cloning of cDNA containing plasmids via antibody-cell surface carbohydrate selection procedures, cDNA encoding the enzymes will be isolated and sequenced. Based on this information and enzyme derived from classical purification or gene expression procedures, studies will be conducted to define the nature of the transferases via active site studies and monoclonal antibody production and analysis. Studies will continue concerning the basis for relative synthesis of type 1 or 2 chain structures in normal mucosa and adenocarcinoma tumors building on present findings of the participation of membrane organization in this process. These studies will provide important information relating to the genetic and epigenetic regulation of enzymes associated with lacto-series core chains during development and oncogenesis.
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