title: High throughput screening of potentially selective MMP-13 exosite inhibitors utilizing a triple-helical FRET substrate.
abstract: The major components of the cartilage extracellular matrix are type II collagen and aggrecan. Matrix metalloproteinase 13 (MMP-13) has been implicated as the protease responsible for collagen degradation in cartilage during osteoarthritis (OA). In the present study, a triple-helical FRET substrate has been utilized for high throughput screening (HTS) of MMP-13 with the MLSCN compound library (n approximately 65,000). Thirty-four compounds from the HTS produced pharmacological dose-response curves. A secondary screen using RP-HPLC validated 25 compounds as MMP-13 inhibitors. Twelve of these compounds were selected for counter-screening with 6 representative MMP family members. Five compounds were found to be broad-spectrum MMP inhibitors, 3 inhibited MMP-13 and one other MMP, and 4 were selective for MMP-13. One of the selective inhibitors was more active against MMP-13 triple-helical peptidase activity compared with single-stranded peptidase activity. Since the THP FRET substrate has distinct conformational features that may interact with MMP secondary binding sites (exosites), novel non-active site-binding inhibitors may be identified via HTS protocols utilizing such assays.
authors: Department of Chemistry and Biochemistry, Florida Atlantic University, 777 Glades Road, Boca Raton, FL 33431-0991, USA.
authors: Janelle L
authors: JL
authors: Lauer-Fields
authors: Dmitriy
authors: D
authors: Minond
authors: Peter S
authors: PS
authors: Chase
authors: Pierre E
authors: PE
authors: Baillargeon
authors: S Adrian
authors: SA
authors: Saldanha
authors: Roma
authors: R
authors: Stawikowska
authors: Peter
authors: P
authors: Hodder
authors: Gregg B
authors: GB
authors: Fields
mesh_terms: D001665:Binding Sites; D004353:Drug Evaluation, Preclinical; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D053509:Matrix Metalloproteinase 13 / Q000378:metabolism; D061965:Matrix Metalloproteinase Inhibitors; D020782:Matrix Metalloproteinases / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011480:Protease Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D054852:Small Molecule Libraries; D013379:Substrate Specificity
chemical_list: D061965:Matrix Metalloproteinase Inhibitors; D010455:Peptides; D011480:Protease Inhibitors; D054852:Small Molecule Libraries; D053509:Matrix Metalloproteinase 13; D020782:Matrix Metalloproteinases