Alternative modes of enzyme inhibition are covalently bound irreversible inhibitors. While chemically reactive affinity labels such as chloromethylketones (CMKs) have demonstrated potent protease inhibition and in certain instances in vivo activity, their inherent chemical reactivity precludes development as clinical agents due to safety concerns.24 However, acyloxymethylketones first reported by Krantz as “quiescent affinity labels” for cathepsin B have demonstrated potent levels of enzyme inhibition with relatively low chemical reactivity.25,26 Mechanistic studies of this class of inhibitors with the cysteine protease caspase-1 support the intermediacy of a thiohemiketal arising from the initial 1,2-carbonyl attack. Orientation and stabilization of the acyloxy leaving group within the S1′ domain (Figure 1) of the protease can facilitate SN2 displacement via sulfur migration, resulting in irreversible or bimodal (reversible inhibition followed by slow inactivation) inhibition.27