Camostat mesylate inhibits SARS-CoV-2 activation by TMPRSS2-related proteases and its metabolite GBPA exerts antiviral activity

Hoffmann, Markus; Hofmann-Winkler, Heike; Smith, John K.; Krüger, Nadine; Arora, Prerna; Sørensen, Lambert K.; Søgaard, Ole Schmeltz; Hasselstrøm, Jørgen B.; Winkler, Michael; Hempel, Tim; Raich, Lluís; Olsson, Simon; Danov, Olga; Jonigk, Danny; Yamazoe, Takashi; Yamatsuta, Katsura; Mizuno, Hirotaka; Ludwig, Stephan; Noé, Frank; Kjølby, Mads; Braun, Armin; Sheltzer, Jason M.; Pöhlmann, Stefan

doi:10.1016/j.ebiom.2021.103255

Cited by 284 publications

(258 citation statements)

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“…TMPRSS2 activated the S proteins of all four CoVs, in keeping with published data [31,[59][60][61][62][63][64][65]. Intriguingly, TMPRSS13 enhanced entry driven by the S proteins of the highly virulent SARS-CoV, SARS-CoV-2 and MERS-CoV (in line with other reports [33,34,58]), but not the common cold virus HCoV-229E. MERS-S and 229E-S were both activated by HAT, as reported earlier [63,66], with roughly the same efficiency as TMPRSS2.…”

Section: Among All 18 Ttsps Tmprss2 and Tmprss13 Are The Best Activators Of Sars-2-ssupporting

confidence: 90%

“…HAT proved effective on 229E-S and MERS-S, however it was less active on SARS-S and SARS-2-S, as reported earlier [33,34,58,63]. We did not see the recently reported activation of SARS-2-S by DESC1 [33,34] and TMPRSS11F [33], which belong to the same subfamily as HAT. This suggests that the level of TTSP activation depends on assay conditions, in particular the level of protease that is generated by plasmid transfection.…”

Section: Plos Pathogenscontrasting

confidence: 46%

“…The Type II Transmembrane Serine Protease (TTSP) family, to which TMPRSS2 belongs, contains in total 18 proteases, many of which are expressed in human airways [13]. Two recent analyses with a subset of TTSPs identified TMPRSS13 as a second prominent activator of the SARS-CoV-2 S protein (subsequently referred to as SARS-2-S) [33,34].…”

Section: Introductionmentioning

confidence: 99%

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The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

et al. 2021

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The high transmissibility of SARS-CoV-2 is related to abundant replication in the upper airways, which is not observed for the other highly pathogenic coronaviruses SARS-CoV and MERS-CoV. We here reveal features of the coronavirus spike (S) protein, which optimize the virus towards the human respiratory tract. First, the S proteins exhibit an intrinsic temperature preference, corresponding with the temperature of the upper or lower airways. Pseudoviruses bearing the SARS-CoV-2 spike (SARS-2-S) were more infectious when produced at 33°C instead of 37°C, a property shared with the S protein of HCoV-229E, a common cold coronavirus. In contrast, the S proteins of SARS-CoV and MERS-CoV favored 37°C, in accordance with virus preference for the lower airways. Next, SARS-2-S-driven entry was efficiently activated by not only TMPRSS2, but also the TMPRSS13 protease, thus broadening the cell tropism of SARS-CoV-2. Both proteases proved relevant in the context of authentic virus replication. TMPRSS13 appeared an effective spike activator for the virulent coronaviruses but not the low pathogenic HCoV-229E virus. Activation of SARS-2-S by these surface proteases requires processing of the S1/S2 cleavage loop, in which both the furin recognition motif and extended loop length proved critical. Conversely, entry of loop deletion mutants is significantly increased in cathepsin-rich cells. Finally, we demonstrate that the D614G mutation increases SARS-CoV-2 stability, particularly at 37°C, and, enhances its use of the cathepsin L pathway. This indicates a link between S protein stability and usage of this alternative route for virus entry. Since these spike properties may promote virus spread, they potentially explain why the spike-G614 variant has replaced the early D614 variant to become globally predominant. Collectively, our findings reveal adaptive mechanisms whereby the coronavirus spike protein is adjusted to match the temperature and protease conditions of the airways, to enhance virus transmission and pathology.

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Section: Among All 18 Ttsps Tmprss2 and Tmprss13 Are The Best Activators Of Sars-2-ssupporting

confidence: 90%

Section: Plos Pathogenscontrasting

confidence: 46%

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The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

et al. 2021

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“…33 Rapid hydrolysis to (4-(4-guanidinobenzoyl-oxy)phenylacetic acid), also known as the protease inhibitor FOY 251 ( Figure 3A), occurs both in vitro in serum as well as in vivo. [34][35][36] We observe similar potencies of FOY 251 and camostat in our biochemical assay, with apparent IC50 = 4.3 ± 0.9 nM and 2.7 ± 0.4 nM, respectively ( Figure S4). The half-life of FOY 251 is longer than camostat, though it is metabolized to 4-guanidinobenzoic acid, which shows minimal TMPRSS2 inhibition.…”

Section: Analysis Of Covalent Tmprss2 Inhibitorsmentioning

confidence: 80%

“…The half-life of FOY 251 is longer than camostat, though it is metabolized to 4-guanidinobenzoic acid, which shows minimal TMPRSS2 inhibition. 34 Thus, it would be advantageous to identify novel inhibitors that have alternative molecular scaffolds.…”

Section: Analysis Of Covalent Tmprss2 Inhibitorsmentioning

confidence: 99%

TMPRSS2 inhibitor discovery facilitated through anin silicoand biochemical screening platform

Jm²,

Wu³

et al. 2021

Preprint

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The COVID-19 pandemic has highlighted the need for new antiviral targets, as many of the currently approved drugs have proven ineffective against mitigating SARS-CoV-2 infections. The host transmembrane serine protease TMPRSS2 is a highly promising antiviral target, as it plays a direct role in priming the spike protein before viral entry occurs. Further, unlike other targets such as ACE2, TMPRSS2 has no known biological role. Here we utilize virtual screening to curate large libraries into a focused collection of potential inhibitors. Optimization of a recombinant expression and purification protocol for the TMPRSS2 peptidase domain facilitates subsequent biochemical screening and characterization of selected compounds from the curated collection in a kinetic assay. In doing so, we demonstrate that serine protease inhibitors camostat, nafamostat, and gabexate inhibit through a covalent mechanism. We further identify new non-covalent compounds as TMPRSS2 protease inhibitors, demonstrating the utility of a combined virtual and experimental screening campaign in rapid drug discovery efforts.

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MPI8 is Potent against SARS‐CoV‐2 by Inhibiting Dually and Selectively the SARS‐CoV‐2 Main Protease and the Host Cathepsin L**

Xinyu

Alugubelli

et al. 2021

ChemMedChem

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A number of inhibitors have been developed for the SARS‐CoV‐2 main protease (M Pro ) as potential COVID‐19 medications but little is known about their selectivity. Using enzymatic assays, we characterized inhibition of TMPRSS2, furin, and cathepsins B/K/L by more than a dozen of previously developed M Pro inhibitors including MPI1‐9, GC376, 11a, 10–1, 10–2, and 10–3. MPI1‐9, GC376 and 11a all contain an aldehyde for the formation of a reversible covalent hemiacetal adduct with the M Pro active site cysteine and 10–1, 10–2 and 10–3 contain a labile ester to exchange with the M Pro active site cysteine for the formation of a thioester. Our data revealed that all these inhibitors are inert toward TMPRSS2 and furin. Diaryl esters also showed low inhibition of cathepsins. However, all aldehyde inhibitors displayed high potency in inhibiting three cathepsins. Their determined IC 50 values vary from 4.1 to 380 nM for cathepsin B, 0.079 to 2.3 nM for cathepsin L, and 0.35 to 180 nM for cathepsin K. All aldehyde inhibitors showed similar inhibition levels toward cathepsin L. A cellular analysis indicated high potency of MPI5 and MPI8 in inhibiting lysosomal activity, which is probably attributed to their inhibition of cathepsins. Among all aldehyde inhibitors, MPI8 shows the best selectivity toward cathepsin L. With respect to cathepsins B and K, the selective indices are 192 and 150, respectively. MPI8 is the most potent compound among all aldehyde inhibitors in cellular M Pro inhibition potency and anti‐SARS‐CoV‐2 activity in Vero E6 cells. Cathepsin L has been demonstrated to play a critical role in the SARS‐CoV‐2 cell entry. By selectively inhibiting both SARS‐CoV‐2 M Pro and the host cathepsin L, MPI8 potentiates dual inhibition effects to synergize its overall antiviral potency and efficacy. Due to its high selectivity toward cathepsin L that reduces potential toxicity toward host cells and high cellular and antiviral potency, we urge serious consideration of MPI8 for preclinical and clinical investigations for treating COVID‐19.

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Camostat mesylate inhibits SARS-CoV-2 activation by TMPRSS2-related proteases and its metabolite GBPA exerts antiviral activity

Cited by 284 publications

References 58 publications

The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

TMPRSS2 inhibitor discovery facilitated through anin silicoand biochemical screening platform

MPI8 is Potent against SARS‐CoV‐2 by Inhibiting Dually and Selectively the SARS‐CoV‐2 Main Protease and the Host Cathepsin L**

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