Isothiazolones interfere with normal matrix metalloproteinase activation and inhibit cartilage proteoglycan degradation.
A series of isothiazolones that inhibit pro-(matrix metallo-proteinase) (proMMP) activation but do not inhibit the active enzyme are effective as cartilage protectants in bovine nasal cartilage organ culture, preventing interleukin-1 (IL-1)-induced proteoglycan (aggrecan) degradation without affecting its synthesis. These compounds were found to bind to prostromelysin (proMMP-3) in a non-dialysable and stoichiometric manner. Preincubation with cartilage-protectant isothiazolones prevented the binding of [14C]iodoacetamide to Cys75 of the MMP-3 propeptide, suggesting that the activity of these compounds involves their binding to the Cys75 of the MMP zymogen. Studies following chymotrypsin activation of proMMP-3 by SDS/PAGE indicated that altered processing of the 57 kDa zymogen to the active form occurred in the presence of compound. The 53 kDa intermediate seen on normal activation was not formed; instead a different intermediate appeared with a molecular mass of approx. 46 kDa. N-terminal sequence analysis indicated that this intermediate was formed by cleavage at the putative 4-aminophenylmercuric acid cleavage site. Importantly the 45 kDa active MMP-3 species formed in the presence of compound was one amino acid residue shorter than the native MMP-3. These results suggest that the inhibition of cartilage proteoglycan degradation by isothiazolones might be due to their ability to bind to the Cys75 in the propeptide region of the MMP zymogen and interfere with its normal activation process.
Arner
E C
EC
DuPont Merck Pharmaceutical Company, Experimental Station, Wilmington, DE 19880-0400, USA.
Pratta
M A
MA
Freimark
B
B
Lischwe
M
M
Trzaskos
J M
JM
Magolda
R L
RL
Wright
S W
SW
D000595:Amino Acid Sequence; D000818:Animals; D002356:Cartilage / Q000378:metabolism*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D019278:Matrix Metalloproteinase 3 / Q000737:chemistry / Q000378:metabolism*; D061965:Matrix Metalloproteinase Inhibitors; D008969:Molecular Sequence Data; D009666:Nose; D009924:Organ Culture Techniques; D011480:Protease Inhibitors / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational; D011509:Proteoglycans / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013844:Thiazoles / Q000494:pharmacology*
D004792:Enzyme Precursors; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors; D011509:Proteoglycans; D013844:Thiazoles; D019278:Matrix Metalloproteinase 3