Inhibition of norovirus 3CL protease by bisulfite adducts of transition state inhibitors

Abstract

Noroviruses are the most common cause of acute viral gastroenteritis, accounting for >21 million cases annually in the U.S. alone. Norovirus infections constitute an important health problem for which there are no specific antiviral therapeutics or vaccines. In this study, a series of bisulfite adducts derived from representative transition state inhibitors (dipeptidyl aldehydes and α-ketoamides) was synthesized and shown to exhibit anti-norovirus activity in a cell-based replicon system. The ED50 of the most effective inhibitor was 60 nM. This study demonstrates for the first time the utilization of bisulfite adducts of transition state inhibitors in the inhibition of norovirus 3C-like protease in vitro and in a cell-based replicon system. The approach described herein can be extended to the synthesis of the bisulfite adducts of other classes of transition state inhibitors of serine and cysteine proteases, such as α-ketoheterocycles and α-ketoesters.

Potent inhibition of norovirus by dipeptidyl α-hydroxyphosphonate transition state mimics

“…Consistent with previous findings, 11 the replacement of the P2 leucine with cyclohexyl alanine enhanced the inhibitory activity of the compounds (compare 7a/7d and 7b/7e in Table 1). The effect of varying R 2 (assumed to be projecting toward the S’ subsites) on potency was also explored.…”

“…Processing of the polyprotein by 3CLpro is essential for virus replication. 8 We have recently described the structure-based design, synthesis, and evaluation of transition state inhibitors of norovirus 3CLpro, including peptidyl aldehydes, 9 peptidyl α-ketoamides and α-ketoheterocycles and their corresponding bisulfite salts, 10–11 and macrocyclic inhibitors. 12 These studies have demonstrated that representative members of these classes of compounds potently inhibit norovirus 3CL proteases from various genogroups and exhibit significant anti-norovirus activity in a cell-based replicon system.…”

“…13,17–19 Mapping of the active site of 3CLpro using chromogenic and fluorogenic substrates has shown that the protease has a strong preference for a -D/E-F/Y-X-L-Q-G- sequence, where X is H, E or Q, and cleavage is at the Q-G (P 1 -P 1 ’) 20 bond; 17,21–23 (c) we have previously demonstrated that the presence of a P 2 cyclohexyl alanine residue results in a significant enhancement in potency and cellular permeability. 11,24 Based on the aforementioned considerations, it was envisaged that an inhibitor represented by general structure (I) (Figure 1) may display high anti-norovirus activity in a cell-based replicon system mediated via the inhibition of NV 3CLpro.…”

Quantitative electrochemical detection of cathepsin B activity in breast cancer cell lysates using carbon nanofiber nanoelectrode arrays toward identification of cancer formation

“…The synthesis of cathepsin B specific tetrapeptide H 2 N-(CH 2 ) 4 -CO-Leu-Arg-Phe-Gly-NH-CH 2 -Fc and other peptides including tripeptide Arg-Phe-Gly-NH-CH 2 -Fc, dipeptide Phe-Gly-NH-CH 2 -Fc and modified single amino acid Gly-NH-CH 2 -Fc for studying cleaving sites is also described in the SI (Scheme S1). The cathepsin B inhibitor GC-373 was prepared 17 in house as described in the SI.…”

Treatment of norovirus infections: Moving antivirals from the bench to the bedside

“…Most recently, the same group of investigators has described a class of molecules, generally termed dipeptidyl or tripeptidyl transition state inhibitors, which display potent, broad-spectrum activity in enzymatic and cell culture-based assays against several 3C-like proteases of viruses from the families Picornaviridae, Coronaviridae , and Caliciviridae (including NV), which share conserved features in the protease catalytic site (Kim et al, 2012; Mandadapu et al, 2013; Prior et al, 2013; Takahashi et al, 2013). Mechanistically, these molecules have been shown to be covalently bound to the nucleophilic cysteine residue in the catalytic sites of the proteases of NV and poliovirus in co-crystallography studies (Kim et al, 2012).…”