Mitigating the Metabolic Liability of Carbonyl Reduction: Novel Calpain Inhibitors with P1′ Extension

Abstract

Dysregulation of calpains 1 and 2 has been implicated in a variety of pathological disorders including ischemia/reperfusion injuries, kidney diseases, cataract formation, and neurodegenerative diseases such as Alzheimer's disease (AD). 2-(3-Phenyl-1)-pyrazol-1-yl)nicotinamides represent a series of novel and potent calpain inhibitors with high selectivity and efficacy. However, carbonyl reduction leading to the formation of the inactive hydroxyamide was identified as major metabolic liability in monkey and human, a pathway not reflected by routine absorption, distribution, metabolism, and excretion (ADME) assays. Using cytosolic clearance as a tailored ADME assay coupled with hepatocyte metabolism enabled the identification of analogues with enhanced stability against carbonyl reduction. These efforts led to the identification of P1' modified calpain inhibitors with significantly improved pharmacokinetic profile including P1'-methoxyamide as potential candidate compound for non-central nervous system indications.

The Alpha Keto Amide Moiety as a Privileged Motif in Medicinal Chemistry: Current Insights and Emerging Opportunities

“…18 The α-ketoamide moiety was then further modified on the nitrogen with a set of different alkyl-, O-alkyl-, aryl-, and heteroaryl residues to identify calpain inhibitors with enhanced stability against carbonyl reduction, which should translate into an improved pharmacokinetic profile in humans. 201 N-Alkyl extension presented a strong increase in cytosolic stability but also a significant reduction in calpain inhibition, as in compounds 126−130 (Table 12). Aromatic moieties were better tolerated, in terms of calpain inhibition, as exemplified by compounds 129 and 130 (Table 12).…”

“…The α-ketoamide moiety was then further modified on the nitrogen with a set of different alkyl-, O -alkyl-, aryl-, and heteroaryl residues to identify calpain inhibitors with enhanced stability against carbonyl reduction, which should translate into an improved pharmacokinetic profile in humans. 201 …”

“…Even if they showed a diminished calpain inhibition in vitro, IC 50 values of 750 ( 128 ) and 2150 nM ( 131 ) were comparable to primary amide 124 in terms of cellular efficacy. 201 …”

“…On the other hand, these analogues were not suitable for further advancement due to insufficient stability in liver microsomes, probably due to the enhanced lipophilicity. P1′ N -alkoxy products ( 131 – 133 , Table ) were generally more potent than the corresponding N -alkyl analogues . From this series containing more than 80 derivatives, the cycloalkyl amide 128 and N -methoxy amide 131 emerged as those molecules with the best balance between calpain inhibition, microsomal and cytosolic stability, and selectivity versus cysteine protease cathepsins.…”

“…Several P1′ alkyl, o -alkyl, aryl, and heteroaryl amides were then synthesized (as examples, 140 – 143 in Table ) to increment the cytosolic stability, even though the ability to inhibit the primary target for most of the analogues decreased. Some aromatic P1′ modifications ( 141 – 142 ) had a positive impact on cathepsin selectivity while also retaining calpain inhibition, but the advancement was abandoned because of the low stability in liver microsomes …”