Dramatic changes in glycolipid metabolism associated with oncogenic transformation implicate a specific role for membrane glycolipids in regulation of cell growth and cellular interaction. 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 regulation of the biosynthetic mechanism which prevents expression of these antigens in normal cells and tissues and is activated to produce them in association with oncogenesis. This application proposes to study the regulation of synthesis of the X determinant antigen, important in many human adenocarcinomas, by detailed study of the glycosyltransferase enzymes associated with its biosynthesis and their differences in expression in normal tissues and tumors. A bl-4galactosyltransferase involved in the synthesis of type 2 glycolipid carbohydrate chain precursors and an al-3fucosyltransferase which utilizes type 2 chain structures as precursors for X determinant biosynthesis will be purified to homogeneity by affinity chromatographic procedures and their properties characterized. Antibodies specific for these enzymes will be prepared and used to study the tissue specificity of these enzymes in normal tissues and tumors. Glycolipids isolated from normal human colonic mucosa, colonic adenocarcinomas, and tumor cell lines will be characterized by thin layer chromatographic immunostain analysis for the presence of X determinant carrying structures and their precursors. These results will be correlated with the presence or absence of specific glycosyltransferase activities relating to synthesis of these antigens in order to determine the nature of the enzymatic block preventing expression in normal tissues. The nature of the regulatory properties of these transferase activities will be investigated with respect to other processes which may modulate their activity and be associated with alterations in the expression of X antigen. These studies will provide information relating to the biochemical changes occurring in association with oncogenesis which gives rise to an important and potentially immunologically useful human tumor associated antigen, the X determinant structure.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA041521-02
Application #
3182090
Study Section
Pathology B Study Section (PTHB)
Project Start
1986-01-07
Project End
1988-12-31
Budget Start
1987-01-01
Budget End
1987-12-31
Support Year
2
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Pacific Northwest Research Institute
Department
Type
DUNS #
City
Seattle
State
WA
Country
United States
Zip Code
98122
Amado, M; Almeida, R; Carneiro, F et al. (1998) A family of human beta3-galactosyltransferases. Characterization of four members of a UDP-galactose:beta-N-acetyl-glucosamine/beta-nacetyl-galactosamine beta-1,3-galactosyltransferase family. J Biol Chem 273:12770-8
Schwientek, T; Almeida, R; Levery, S B et al. (1998) Cloning of a novel member of the UDP-galactose:beta-N-acetylglucosamine beta1,4-galactosyltransferase family, beta4Gal-T4, involved in glycosphingolipid biosynthesis. J Biol Chem 273:29331-40
Holmes, E H; Xu, Z; Sherwood, A L et al. (1995) Structure-function analysis of human alpha 1-->3fucosyltransferases. A GDP-fucose-protected, N-ethylmaleimide-sensitive site in FucT-III and FucT-V corresponds to Ser178 in FucT-IV. J Biol Chem 270:8145-51
Hu, J; Stults, C L; Holmes, E H et al. (1994) Structural characterization of intermediates in the biosynthetic pathway of neolacto glycosphingolipids: differential expression in human leukaemia cells. Glycobiology 4:251-7
Meldgaard, P; Holmes, E H; Bennett, E P et al. (1994) Blood group ABO-related glycosylation of urothelial cell lines: immunocytological, enzymatic, and genetic characterization. Cancer Res 54:2440-7
Holmes, E H; Macher, B A (1993) Specificity of fucose transfer to GlcNAc residues of extended chain neolacto-series glycolipids catalyzed by human alpha 1-->3fucosyltransferases: effect of the lipidic environment on the myeloid enzyme form. Arch Biochem Biophys 301:190-9
Holmes, E H (1993) Human Lewis alpha 1-->3/4fucosyltransferase: specificity of fucose transfer to GlcNAc beta 1-->3Gal beta 1-->4Glc beta 1-->1Cer (LcOse3Cer). Glycobiology 3:77-81
Holmes, E H; Greene, T G (1993) De novo synthesis of type 1 lacto-series glycolipids in human colonic adenocarcinoma cells: efficient synthesis of the Le(a) antigen and absence of brefeldin A-induced inhibition of its synthesis in Colo 205 cells. Arch Biochem Biophys 305:328-40
Symington, F W; Holmes, E H; Symington, B E (1992) Human epidermal keratinocyte expression of sialyl-Lewis X. J Invest Dermatol 99:601-7
Sherwood, A L; Greene, T G; Holmes, E H (1992) Stable expression of a cDNA encoding a human beta 1 --> 3galactosyltransferase responsible for lacto-series type 1 core chain synthesis in non-expressing cells: variation in the nature of cell surface antigens expressed. J Cell Biochem 50:165-77

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