THE GELATINASE INHIBITORY ACTIVITY OF TETRACYCLINES AND CHEMICALLY-MODIFIED TETRACYCLINE ANALOGS AS MEASURED BY A NOVEL MICROTITER ASSAY FOR INHIBITORS

A quantitative nonisotopic solution assay for gelatinases and inhibito rs was developed using biotinylated gelatin as enzyme substrate. In th is assay, residual biotinylated substrate is sandwiched between avidin -coated plates and streptavidin-peroxidase and is quantified by the pe roxidase reaction. This assay was useful for measuring gelatinase acti vities and defining the activities of gelatinase inhibitors. When 23 t etracycline analogues were compared, significant differences in gelati nase B inhibition were found between various compounds. 4-epioxytetrac yclin base, 4-epichlortetracycline, meclocyclinesulfosalicylate, and u nmodified metacycline and minocycline proved to be the most potent gel atinase B (EC 3.4.24.35) inhibitors. The gelatinase B inhibitory activ ity of tetracyclines was clearly dissociated from their antimicrobial activity. The effect of high-molecular-weight inhibitors, such as mono clonal antibodies, was also demonstrable in the microtiter plate assay . In view of the pathophysiological function of gelatinases, the defin ition of gelatinase inhibitors with known efficacy, safety, and side e ffects is crucial for the treatment of diseases such as rheumatoid art hritis and multiple sclerosis. Particular tetracyclines fulfil these c riteria and the described assay is useful for defining other gelatinas e inhibiting lead compounds.