Drug Repurposing for the SARS‐CoV‐2 Papain‐Like Protease

Abstract

As the pathogen of COVID‐19, SARS‐CoV‐2 encodes two essential cysteine proteases that process the pathogen's two large polypeptide products pp1a and pp1ab in the human cell host to form 15 functionally important, mature nonstructural proteins. One of the two enzymes is papain‐like protease or PL Pro . It possesses deubiquitination and deISGylation activities that suppress host innate immune responses toward SARS‐CoV‐2 infection. To repurpose drugs for PL Pro , we experimentally screened libraries of 33 deubiquitinase and 37 cysteine protease inhibitors on their inhibition of PL Pro . Our results showed that 15 deubiquitinase and 1 cysteine protease inhibitors exhibit strong inhibition of PL Pro at 200 μM. More comprehensive characterizations revealed seven inhibitors GRL0617, SJB2‐043, TCID, DUB‐IN‐1, DUB‐IN‐3, PR‐619, and S130 with an IC 50 value below 40 μM and four inhibitors GRL0617, SJB2‐043, TCID, and PR‐619 with an IC 50 value below 10 μM. Among four inhibitors with an IC 50 value below 10 μM, SJB2‐043 is the most unique in that it does not fully inhibit PL Pro but has a noteworthy IC 50 value of 0.56 μM. SJB2‐043 likely binds to an allosteric site of PL Pro to convene its inhibition effect, which needs to be further investigated. As a pilot study, the current work indicates that COVID‐19 drug repurposing by targeting PL Pro holds promise, but in‐depth analysis of repurposed drugs is necessary to avoid omitting critical allosteric inhibitors.

Mass Spectrometric Assays Reveal Discrepancies in Inhibition Profiles for the SARS‐CoV‐2 Papain‐Like Protease

“…By contrast, PF‐07321332 (nirmatrelvir), a selective SARS‐CoV‐2 M pro inhibitor which is in clinical development and which inhibits covalently by reaction with the active site cysteine, [2f] does not inhibit PL pro (Table 3, entry 2). Similarly, MK‐0822 (odanacatib), a reported selective covalent inhibitor of the human cysteine protease cathepsin K, [33] does not inhibit PL pro (Table 3, entry 3), in accord with a recent report [34] . SDZ‐224015, which is an investigational inhibitor of the human cysteine protease caspase‐1 [35] and which is also reported to efficiently inhibit SARS‐CoV‐2 M pro by irreversible alkylation of the active site cysteine, [8b] does not inhibit PL pro (Table 3, entry 4), in agreement with its reported inability to inhibit PL pro [8b] …”

Potential Inhibitors Targeting Papain-Like Protease of SARS-CoV-2: Two Birds With One Stone

“…Compared with drug repurposing, screenings for hit compounds, either in vitro or in silico, incorporate a much larger scale of molecules including microbial metabolites, natural products and marine-derived bioactive compounds (Cragg and Newman, 2013;Kumar et al, 2021;Quimque et al, 2021). Since the outbreak of COVID-19, the widespread use of combinatorial screening has revealed a large mount of hit compounds that specifically target SARS-CoV-2 PLpro (Amin et al, 2021;Gogoi et al, 2021;Goyzueta-Mamani et al, 2021;Hajbabaie et al, 2021;Jade et al, 2021;Jamalan et al, 2021;Kumar et al, 2021;Li et al, 2021;Quimque et al, 2021;Rahul and Sarkar, 2021;Rudrapal et al, 2021;Stasiulewicz et al, 2021). Hits generated through the initial screens are further validated with an aim to choose the best ones to serve as leads for drug developments.…”

“…Smith et al developed a cell-based luciferase complementation assay to evaluate the inhibition of known drugs against SARS-CoV-2 viral PLpro ( Smith et al, 2020 ). Based on the fluorescence-based enzymatic inhibition assay, several groups have identified potential inhibitors of SARS-CoV-2 PLpro with the IC 50 value under 10 μM ( Cho et al, 2021 ; Xu et al, 2021 ; Zhao et al, 2021 ). Although not all the identified compounds exhibited good performance in a cell-based assay, they could serve as a starting point for further chemical modification.…”

Invalidation of dieckol and 1,2,3,4,6-pentagalloylglucose (PGG) as SARS-CoV-2 main protease inhibitors and the discovery of PGG as a papain-like protease inhibitor

“…For example, boceprevir [ 10 , 11 ], calpain inhibitors [ 10 ], GC-376 [ 10 , 12 ], and masitinib [ 13 ] were among the first hits reported as M pro inhibitors. GRL0617 [ 14 , 15 ], YM155 [ 16 ], 6-thioguanine [ 17 ], SJB2-043 [ 18 ], and others were identified as PL pro inhibitors. Natural products have always been a rich source of modern medicine [ 19 ], and multiple natural products have been reported as M pro and PL pro inhibitors [ 20 ].…”