title: Glycosylation-dependent inhibition of cutaneous lymphocyte-associated antigen expression: implications in modulating lymphocyte migration to skin.
abstract: Constitutive E-selectin expression on dermal microvascular endothelial cells plays a critical role in mediating rolling adhesive interactions of human skin-homing T cells and in pathologic accumulation of lymphocytes in skin. The major E-selectin ligand on human skin-homing T cells is cutaneous lymphocyte-associated antigen (CLA), a specialized glycoform of P-selectin glycoprotein ligand-1 (PSGL-1) defined by monoclonal antibody HECA-452. Since HECA-452 reactivity, and not PSGL-1 polypeptide itself, confers the specificity of human T cells to enter dermal tissue, inhibition of HECA-452 expression is a potential strategy for modulating lymphocyte migration to skin. In this study, we examined the efficacy of several well-characterized metabolic inhibitors of glycosylation and of a novel fluorinated analog of N-acetylglucosamine (2-acetamido-1,3,6-tri-O-acetyl-4-deoxy-4-fluoro-D-glucopyranose [4-F-GlcNAc]) to alter HECA-452 expression on human CLA(+) T cells and prevent cell tethering and rolling on selectins under shear stress. At concentrations that did not affect PSGL-1 expression, we found that swainsonine (inhibitor of complex-type N-glycan synthesis) had no effect on HECA-452 expression or selectin ligand activity, whereas benzyl-O-N-acetylgalactosamide (BAG; inhibitor of O-glycan biosynthesis) ablated HECA-452 expression on PSGL-1 and significantly lowered selectin ligand activity. We found that 4-F-GlcNAc (putative inhibitor of poly-N-acetyllactosamine biosynthesis) was more potent than BAG at lowering HECA-452 expression and selectin binding. In addition, we show that 4-F-GlcNAc was directly incorporated into native CLA expressed on T cells, indicating direct inhibition on poly-N-acetyllactosamine elongation and selectin-binding determinants on PSGL-1 O-glycans. These observations establish a potential treatment approach for targeting pathologic lymphocyte trafficking to skin and indicate that 4-F-GlcNAc may be a promising agent for treatment of dermal tropism associated with malignancies and inflammatory disorders.
authors.affiliation: Department of Dermatology and the Harvard Skin Disease Research Center at Brigham and Women's Hospital, Boston, MA, USA.
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Dimitroff
authors.forename: Ralph J
authors.initials: RJ
authors.lastname: Bernacki
authors.forename: Robert
authors.initials: R
authors.lastname: Sackstein
mesh_terms: D000117:Acetylglucosamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D036904:Leukocyte Rolling / Q000187:drug effects*; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects*; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000187:drug effects / Q000502:physiology; D011134:Polysaccharides / Q000096:biosynthesis; D012871:Skin Diseases / Q000188:drug therapy / Q000276:immunology; D013329:Structure-Activity Relationship; D017026:Swainsonine / Q000494:pharmacology; D014415:Tunicamycin / Q000494:pharmacology
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm; C063419:CTAGE1 protein, human; D008562:Membrane Glycoproteins; C084882:P-selectin ligand protein; D011134:Polysaccharides; D014415:Tunicamycin; C089178:2-acetamido-1,3,6-tri-O-acetyl-4-deoxy-4-fluoroglucopyranose; C037199:poly-N-acetyllactosamine; D017026:Swainsonine; D000117:Acetylglucosamine