Peroxidasin Inhibition by Phloroglucinol and Other Peroxidase Inhibitors

Abstract

Human peroxidasin (PXDN) is a ubiquitous peroxidase enzyme expressed in most tissues in the body. PXDN represents an interesting therapeutic target for inhibition, as it plays a role in numerous pathologies, including cardiovascular disease, cancer and fibrosis. Like other peroxidases, PXDN generates hypohalous acids and free radical species, thereby facilitating oxidative modifications of numerous biomolecules. We have studied the inhibition of PXDN halogenation and peroxidase activity by phloroglucinol and 14 other peroxidase inhibitors. Although a number of compounds on their own potently inhibited PXDN halogenation activity, only five were effective in the presence of a peroxidase substrate with IC50 values in the low μM range. Using sequential stopped-flow spectrophotometry, we examined the mechanisms of inhibition for several compounds. Phloroglucinol was the most potent inhibitor with a nanomolar IC50 for purified PXDN and IC50 values of 0.95 μM and 1.6 μM for the inhibition of hypobromous acid (HOBr)-mediated collagen IV cross-linking in a decellularized extracellular matrix and a cell culture model. Other compounds were less effective in these models. Most interestingly, phloroglucinol was identified to irreversibly inhibit PXDN, either by mechanism-based inhibition or tight binding. Our work has highlighted phloroglucinol as a promising lead compound for the design of highly specific PXDN inhibitors and the assays used in this study provide a suitable approach for high-throughput screening of PXDN inhibitors.

The Versatility of Collagen in Pharmacology: Targeting Collagen, Targeting with Collagen

“…Phloroglucinol, a known inhibitor of PXDN [ 163 , 164 ], emerges as a promising compound already formulated for other purposes since a daily intraperitoneal injection of phloroglucinol has been shown to reduce interstitial fibrosis in a mouse model of unilateral ureteral obstruction [ 165 ]. In vitro studies have recently identified new inhibitors of PXDN [ 166 ]. One noteworthy feature of collagen IV is its structural diversity.…”