title: Identification of DNMT1 selective antagonists using a novel scintillation proximity assay.
abstract: A novel scintillation proximity high throughput assay (SPA) to identify inhibitors of DNA methyltransferases was developed and used to screen over 180,000 compounds. The majority of the validated hits shared a quinone core and several were found to generate the reactive oxygen species, H2O2. Inhibition of the production of H2O2 by the addition of catalase blocked the ability of this group of compounds to inhibit DNA methyltransferase (DNMT) activity. However, a related compound, SW155246, was identified that existed in an already reduced form of the quinone. This compound did not generate H2O2, and catalase did not block its ability to inhibit DNA methyltransferase. SW155246 showed a 30-fold preference for inhibition of human DNMT1 versus human or murine DNMT3A or -3B, inhibited global methylation in HeLa cells, and reactivated expression of the tumor suppressor gene RASSF1A in A549 cells. To our knowledge, this work represents the first description of selective chemical inhibitors of the DNMT1 enzyme.
authors.affiliation: Department of Biochemistry, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390, USA.
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Kilgore
authors.forename: Xinlin
authors.initials: X
authors.lastname: Du
authors.forename: Lisa
authors.initials: L
authors.lastname: Melito
authors.forename: Shuguang
authors.initials: S
authors.lastname: Wei
authors.forename: Changguang
authors.initials: C
authors.lastname: Wang
authors.forename: Hang Gyeong
authors.initials: HG
authors.lastname: Chin
authors.forename: Bruce
authors.initials: B
authors.lastname: Posner
authors.forename: Sriharsa
authors.initials: S
authors.lastname: Pradhan
authors.forename: Joseph M
authors.initials: JM
authors.lastname: Ready
authors.forename: Noelle S
authors.initials: NS
authors.lastname: Williams
mesh_terms: D000818:Animals; D001681:Biological Assay; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000090205:DNA Methyltransferase 3A; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D051379:Mice; D016877:Oxidants / Q000494:pharmacology; D061987:Sf9 Cells; D018411:Spodoptera; D025521:Tumor Suppressor Proteins / Q000096:biosynthesis / Q000235:genetics; D000097584:DNA Methyltransferase 3B
chemical_list: D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D004791:Enzyme Inhibitors; D006861:Hydrogen Peroxide; D016877:Oxidants; D025521:Tumor Suppressor Proteins; D000097584:DNA Methyltransferase 3B; C000717967:DNMT3A protein, human; C000715535:Dnmt3a protein, mouse; C411762:RASSF1 protein, human; C496882:RASSF1 protein, mouse; C000619380:DNMT1 protein, human; C000619381:Dnmt1 protein, mouse
keywords: DNA Methylation; DNA Methyltransferase; Gene Regulation; High Throughput Screening (HTS); Reactive Oxygen Species (ROS); Scintillation Proximity Assay