Fragment-Sized and Bidentate (Immuno)Proteasome Inhibitors Derived from Cysteine and Threonine Targeting Warheads.

Levente Kollár,Izidor Sosič,Matic Proj,Dorottya Csányi,Tímea Imre,Péter Ábrányi-Balogh,György G Ferenczy,Stanislav Gobec,György M Keserű,Ágnes Gömöry,Damijan Knez,László Petri,Martina Gobec,Lara Smrdel

doi:10.3390/cells10123431

Levente Kollár, Izidor Sosič + Show 12 more

Open Access

https://doi.org/10.3390/cells10123431

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Abstract

Constitutive- and immunoproteasomes are part of the ubiquitin–proteasome system (UPS), which is responsible for the protein homeostasis. Selective inhibition of the immunoproteasome offers opportunities for the treatment of numerous diseases, including inflammation, autoimmune diseases, and hematologic malignancies. Although several inhibitors have been reported, selective nonpeptidic inhibitors are sparse. Here, we describe two series of compounds that target both proteasomes. First, benzoxazole-2-carbonitriles as fragment-sized covalent immunoproteasome inhibitors are reported. Systematic substituent scans around the fragment core of benzoxazole-2-carbonitrile led to compounds with single digit micromolar inhibition of the β5i subunit. Experimental and computational reactivity studies revealed that the substituents do not affect the covalent reactivity of the carbonitrile warhead, but mainly influence the non-covalent recognition. Considering the small size of the inhibitors, this finding emphasizes the importance of the non-covalent recognition step in the covalent mechanism of action. As a follow-up series, bidentate inhibitors are disclosed, in which electrophilic heterocyclic fragments, i.e., 2-vinylthiazole, benzoxazole-2-carbonitrile, and benzimidazole-2-carbonitrile were linked to threonine-targeting (R)-boroleucine moieties. These compounds were designed to bind both the Thr1 and β5i-subunit-specific residue Cys48. However, inhibitory activities against (immuno)proteasome subunits showed that bidentate compounds inhibit the β5, β5i, β1, and β1i subunits with submicromolar to low-micromolar IC50 values. Inhibitory assays against unrelated enzymes showed that compounds from both series are selective for proteasomes. The presented nonpeptidic and covalent derivatives are suitable hit compounds for the development of either β5i-selective immunoproteasome inhibitors or compounds targeting multiple subunits of both proteasomes.

Highlights

Introduction conditions of the Creative CommonsProtein degradation was considered a neglected field before the 1980s, but the discovery of ubiquitin pathways has put the topic in a different perspective [1]
One of the possible strategies for using structure-activity relationship data (SAR) information is based on the concept of group efficiency (GE) [37]
By synthesizing and testing a series of close analogues around the fragment core, the contributions of each substituent can be evaluated and positions prioritized according to their GE value [38]

Summary

Introduction

Protein degradation was considered a neglected field before the 1980s, but the discovery of ubiquitin pathways has put the topic in a different perspective [1]. Proteasome system (UPS) [2] and, in particular, the multicatalytic activity of the 26S protea-. The complex structure of the proteasome can be divided into smaller units. The 20S core particle (CP), which is responsible for the catalytic activity, consists of four stacked rings, each containing seven subunits. Three subunits of the inner β-rings are proteolytically active and are defined as the β1 subunit (caspase-like), the β2 subunit (trypsin-like), and the β5 subunit (chymotrypsin-like). The differential substrate preference of these subunits enables the degradation of diverse ubiquitinated proteins [11,12]

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