Epoxomicin, a potent and selective proteasome inhibitor, exhibits<i>in vivo</i>antiinflammatory activity

TLDR

The proteasome regulates cellular processes such as cell cycle progression and NF‑κB activation. This study demonstrates that the natural product epoxomicin specifically targets the proteasome. Biotinylated‑epoxomicin covalently binds to the LMP7, X, MECL1, and Z catalytic subunits, potently inhibiting chymotrypsin‑like activity while only slowly inhibiting trypsin‑like and peptidyl‑glutamyl activities, and does not affect non‑proteasomal proteases. Epoxomicin effectively blocks NF‑κB activation in vitro and reduces inflammation in a murine ear edema assay, outperforming lactacystin and NLVS, and thus represents a novel, selective proteasome inhibitor for in vivo and in vitro applications.

Abstract

The proteasome regulates cellular processes as diverse as cell cycle progression and NF-κB activation. In this study, we show that the potent antitumor natural product epoxomicin specifically targets the proteasome. Utilizing biotinylated-epoxomicin as a molecular probe, we demonstrate that epoxomicin covalently binds to the LMP7, X, MECL1, and Z catalytic subunits of the proteasome. Enzymatic analyses with purified bovine erythrocyte proteasome reveal that epoxomicin potently inhibits primarily the chymotrypsin-like activity. The trypsin-like and peptidyl-glutamyl peptide hydrolyzing catalytic activities also are inhibited at 100- and 1,000-fold slower rates, respectively. In contrast to peptide aldehyde proteasome inhibitors, epoxomicin does not inhibit nonproteasomal proteases such trypsin, chymotrypsin, papain, calpain, and cathepsin B at concentrations of up to 50 μM. In addition, epoxomicin is a more potent inhibitor of the chymotrypsin-like activity than lactacystin and the peptide vinyl sulfone NLVS. Epoxomicin also effectively inhibits NF-κB activation in vitro and potently blocks in vivo inflammation in the murine ear edema assay. These results thus define epoxomicin as a novel proteasome inhibitor that likely will prove useful in exploring the role of the proteasome in various in vivo and in vitro systems.