A High-Throughput Glycosyltransferase Inhibition Assay for Identifying Molecules Targeting Fucosylation in Cancer Cell-Surface Modification

ACS Chem Biol. 2019 Apr 19;14(4):715-724. doi: 10.1021/acschembio.8b01123. Epub 2019 Mar 14.


In cancers, increased fucosylation (attachment of fucose sugar residues) on cell-surface glycans, resulting from the abnormal upregulation of the expression of specific fucosyltransferase enzymes (FUTs), is one of the most important types of glycan modifications associated with malignancy. Fucosylated glycans on cell surfaces are involved in a multitude of cellular interactions and signal regulation in normal biological processes, as well as in disease. For example, sialyl LewisX is a fucosylated cell-surface glycan that is abnormally abundant in some cancers where it has been implicated in facilitating metastasis, allowing circulating tumor cells to bind to the epithelial tissue within blood vessels and invade into secondary sites by taking advantage of glycan-mediated interactions. To identify inhibitors of FUT enzymes as potential cancer therapeutics, we have developed a novel high-throughput assay that makes use of a fluorogenically labeled oligosaccharide as a probe of fucosylation. This probe, which consists of a 4-methylumbelliferyl glycoside, is recognized and hydrolyzed by specific glycoside hydrolase enzymes to release fluorescent 4-methylumbelliferone, yet when the probe is fucosylated prior to treatment with the glycoside hydrolases, hydrolysis does not occur and no fluorescent signal is produced. We have demonstrated that this assay can be used to measure the inhibition of FUT enzymes by small molecules, because blocking fucosylation will allow glycosidase-catalyzed hydrolysis of the labeled oligosaccharide to produce a fluorescent signal. Employing this assay, we have screened a focused library of small molecules for inhibitors of a human FUT enzyme involved in the synthesis of sialyl LewisX and demonstrated that our approach can be used to identify potent FUT inhibitors from compound libraries in microtiter plate format.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Drug Screening Assays, Antitumor / methods
  • Enzyme Inhibitors / analysis*
  • Enzyme Inhibitors / chemistry
  • Enzyme Inhibitors / pharmacology
  • Enzyme Inhibitors / therapeutic use
  • Fucose / chemistry*
  • Fucosyltransferases / antagonists & inhibitors*
  • Fucosyltransferases / metabolism*
  • Glycosylation
  • High-Throughput Screening Assays*
  • Humans
  • Neoplasms / drug therapy
  • Neoplasms / enzymology
  • Small Molecule Libraries / chemistry
  • Small Molecule Libraries / pharmacology
  • Triazoles / chemistry
  • Triazoles / metabolism


  • Enzyme Inhibitors
  • Small Molecule Libraries
  • Triazoles
  • 1,2,4-triazole
  • Fucose
  • Fucosyltransferases