Investigation of the Compatibility between Warheads and Peptidomimetic Sequences of Protease Inhibitors—A Comprehensive Reactivity and Selectivity Study

Abstract

Covalent peptidomimetic protease inhibitors have gained a lot of attention in drug development in recent years. They are designed to covalently bind the catalytically active amino acids through electrophilic groups called warheads. Covalent inhibition has an advantage in terms of pharmacodynamic properties but can also bear toxicity risks due to non-selective off-target protein binding. Therefore, the right combination of a reactive warhead with a well-suited peptidomimetic sequence is of great importance. Herein, the selectivities of well-known warheads combined with peptidomimetic sequences suited for five different proteases were investigated, highlighting the impact of both structure parts (warhead and peptidomimetic sequence) for affinity and selectivity. Molecular docking gave insights into the predicted binding modes of the inhibitors inside the binding pockets of the different enzymes. Moreover, the warheads were investigated by NMR and LC-MS reactivity assays against serine/threonine and cysteine nucleophile models, as well as by quantum mechanics simulations.

Dual Strategy to Design New Agents Targeting Schistosoma mansoni: Advancing Phenotypic and SmCB1 Inhibitors for Improved Efficacy

“…While two of them are vinylsulfonate-based cysteine protease inhibitors (SJ605, SJ606), originally developed for rhodesain, 18,19 the third is a 4-oxoenoate-based bortezomib congener with S,R-configuration. 24 All three compounds are highly active against NTS at 10 μM (100% efficacy, i.e., 100% dead NTS). Although SJ605 and SJ606 exhibit high SmCB1 inhibition with K i values in the low nanomolar range, a reduced efficacy against adult worms at 10 μM (55% dead worms for SJ605, 58% dead worms for SJ606) was achieved.…”

“…mansoni adults are shown in Figure . While two of them are vinylsulfonate-based cysteine protease inhibitors ( SJ605 , SJ606 ), originally developed for rhodesain, , the third is a 4-oxoenoate-based bortezomib congener with S,R -configuration . All three compounds are highly active against NTS at 10 μM (100% efficacy, i.e., 100% dead NTS).…”

“…In order to maximize their effect, we also focused on improving their physicochemical properties, such as lipophilicity, since the substances have to pass two cell membranes to reach their target site . This resulted in two approaches, the “phenotypic” approach focusing on KS309 derivatization and the “ Sm CB1” approach focusing on SJ605 / SJ606 derivatizations …”

“…25 This resulted in two approaches, the "phenotypic" approach focusing on KS309 derivatization and the "SmCB1" approach focusing on SJ605/ SJ606 derivatizations. 24 Chemistry. Compounds 1a−m were prepared by TBTU peptide coupling chemistry.…”

Recent Advances on Targeting Proteases for Antiviral Development

“…To better understand this, Müller et al led a study that aimed to investigate the influences of different known warhead groups and different peptide sequences on selectivity and reactivity on different proteases [ 74 ]. They synthesized different compounds containing specific peptide sequences coupled with different warhead groups, which were tested in vitro against five different proteins (uPA, CatS, β5-subunit of the proteasome, SARS-CoV-2 M pro and rhodesain), representing three groups of proteases: serine, cysteine and threonine.…”