Development of an HTRF assay for the detection and characterization of inhibitors of catechol-O-methyltransferase

Martha Kimos, Maggi Burton, David Urbain, Didier Caudron, Murielle Martini, Michel Famelart, Michel Gillard, James Barrow, Martyn Wood

Research output: Contribution to journalArticlepeer-review

9 Scopus citations


Catechol-O-methyltransferase (COMT) plays an important role in the deactivation of catecholamine neurotransmitters and hormones. Inhibitors of COMT, such as tolcapone and entacapone, are used clinically in the treatment of Parkinson's disease. Discovery of novel inhibitors has been hampered by a lack of suitable assays for high-throughput screening (HTS). Although assays using esculetin have been developed, these are affected by fluorescence, a common property of catechol-type compounds. We have therefore evaluated a new homogenous time-resolved fluorescence (HTRF)-based assay from CisBio (Codolet, France), which measures the production of S-adenosyl-L-homocysteine (SAH). The assay has been run in both HTS and medium-throughput screening (MTS) modes. The assay was established using membranes expressing human membrane-bound COMT and was optimized for protein and time to give an acceptable signal window, good potency for tolcapone, and a high degree of translation between data in fluorescence ratio and data in terms of [SAH] produced. pIC50 values for the hits from the HTS mode were determined in the MTS mode. The assay also proved suitable for kinetic studies such as Km,app determination.

Original languageEnglish (US)
Pages (from-to)490-495
Number of pages6
JournalJournal of Biomolecular Screening
Issue number5
StatePublished - Jun 2016


  • COMT
  • HTRF
  • catechol-O-methyltransferase

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biotechnology
  • Biochemistry
  • Molecular Medicine
  • Pharmacology
  • Drug Discovery


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