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All Studies   Meta Analysis    Recent:   

Hydroxychloroquine-mediated inhibition of SARS-CoV-2 entry is attenuated by TMPRSS2

Ou et al., PLOS Pathogens, doi:10.1371/journal.ppat.1009212 (date from preprint)
Jul 2020  
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HCQ for COVID-19
1st treatment shown to reduce risk in March 2020
 
*, now known with p < 0.00000000001 from 421 studies, recognized in 42 countries.
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3,800+ studies for 60+ treatments. c19hcq.org
In Vitro analysis showing that HCQ efficiently blocks viral entry mediated by cathepsin L, but not by TMPRSS2, and that a combination of HCQ and a TMPRSS2 inhibitor prevents SARS-CoV-2 infection more potently than either drug alone.
Ou et al., 22 Jul 2020, peer-reviewed, 6 authors.
In Vitro studies are an important part of preclinical research, however results may be very different in vivo.
This PaperHCQAll
Hydroxychloroquine-mediated inhibition of SARS-CoV-2 entry is attenuated by TMPRSS2
Tianling Ou, Huihui Mou, Lizhou Zhang, Amrita Ojha, Hyeryun Choe, Michael Farzan
PLOS Pathogens, doi:10.1371/journal.ppat.1009212
Hydroxychloroquine, used to treat malaria and some autoimmune disorders, potently inhibits viral infection of SARS coronavirus (SARS-CoV-1) and SARS-CoV-2 in cell-culture studies. However, human clinical trials of hydroxychloroquine failed to establish its usefulness as treatment for COVID-19. This compound is known to interfere with endosomal acidification necessary to the proteolytic activity of cathepsins. Following receptor binding and endocytosis, cathepsin L can cleave the SARS-CoV-1 and SARS-CoV-2 spike (S) proteins, thereby activating membrane fusion for cell entry. The plasma membrane-associated protease TMPRSS2 can similarly cleave these S proteins and activate viral entry at the cell surface. Here we show that the SARS-CoV-2 entry process is more dependent than that of SARS-CoV-1 on TMPRSS2 expression. This difference can be reversed when the furincleavage site of the SARS-CoV-2 S protein is ablated or when it is introduced into the SARS-CoV-1 S protein. We also show that hydroxychloroquine efficiently blocks viral entry mediated by cathepsin L, but not by TMPRSS2, and that a combination of hydroxychloroquine and a clinically-tested TMPRSS2 inhibitor prevents SARS-CoV-2 infection more potently than either drug alone. These studies identify functional differences between SARS-CoV-1 and -2 entry processes, and provide a mechanistic explanation for the limited in vivo utility of hydroxychloroquine as a treatment for COVID-19.
media containing 2% FBS. Same infection procedures were applied on other cell lines without the plate coating and transfection steps. Cell surface expression and S protein analysis To measure surface TMPRSS2 expression of 293T-ACE2 transiently transfected with TMPRSS2 and the stable cell line 293T/ACE2/TMPRSS2, cells were detached by 1mM EDTA in PBS and then stained by 2 ug/ml of anti-Flag M2 antibody (Sigma-Aldrich, F1804) and 2 μg/ml of goat anti-mouse IgG (H+L) conjugated with Alexa 647 (Jackson ImmunoResearch Laboratories, #115-606-146). Flow cytometry analysis was done using Accuri C6 (BD Biosciences). To measure the endogenous TMPRSS2 expression of Vero, H1299, H1975 and Calu-3 cells, cells were permeabilized with PBS including 0.5% Triton X-100 (Sigma-Aldrich) at room temperature for 10 min, and detected by 2 μg/ml monoclonal rabbit Anti-TMPRSS2 antibody [EPR3861] (Abcam, ab92323) and goat anti-rabbit IgG conjugated with HRP (Sigma-Aldrich, A0545). To determine the cleavage of S proteins, 293T cells were transfected with 2 μL of lipofectamine 2000 (Life Technologies) in complex with 1 μg plasmid expressing the indicated S protein variant. Cells were harvested for western blot analysis 48 hours post transfection. Cells were permeabilized with PBS including 0.5% Triton X-100 (Sigma-Aldrich) at room temperature for 10 min, and detected by 1 μg/ml anti-Flag M2 antibody (Sigma-Aldrich, F1804) and goat anti-mouse IgG (Fab only) conjugated with HRP (Sigma-Aldrich, A9917). ..
References
Belouzard, Chu, Whittaker, Activation of the SARS coronavirus spike protein via sequential proteolytic cleavage at two distinct sites, Proceedings of the National Academy of Sciences, doi:10.1073/pnas.0809524106
Bertram, Heurich, Lavender, Gierer, Danisch et al., Influenza and SARS-coronavirus activating proteases TMPRSS2 and HAT are expressed at multiple sites in human respiratory and gastrointestinal tracts, PloS one, doi:10.1371/journal.pone.0035876
Bo ¨ttcher, Matrosovich, Beyerle, Klenk, Garten et al., Proteolytic activation of influenza viruses by serine proteases TMPRSS2 and HAT from human airway epithelium, Journal of virology, doi:10.1128/JVI.01118-06
Boulware, Pullen, Bangdiwala, Pastick, Lofgren et al., A randomized trial of hydroxychloroquine as postexposure prophylaxis for Covid-19, New England Journal of Medicine
Chen, Hu, Zhang, Jiang, Han et al., Efficacy of hydroxychloroquine in patients with COVID-19: results of a randomized clinical trial, MedRxiv, doi:10.1001/jama.2020.22240
Chen, Liu, Liu, Liu, Xu et al., A pilot study of hydroxychloroquine in treatment of patients with moderate COVID-19, Zhejiang da xue xue bao Yi xue ban = Journal of Zhejiang University Medical sciences, doi:10.3785/j.issn.1008-9292.2020.03.03
Coutard, Valle, De Lamballerie, Canard, Seidah et al., The spike glycoprotein of the new coronavirus 2019-nCoV contains a furin-like cleavage site absent in CoV of the same clade, Antiviral research, doi:10.1016/j.antiviral.2020.104742
Gautret, Lagier, Parola, Meddeb, Mailhe et al., Hydroxychloroquine and azithromycin as a treatment of COVID-19: results of an open-label non-randomized clinical trial, International journal of antimicrobial agents, doi:10.1016/j.ijantimicag.2020.105949
Geleris, Sun, Platt, Zucker, Baldwin et al., Observational study of hydroxychloroquine in hospitalized patients with Covid-19, New England Journal of Medicine, doi:10.1056/NEJMoa2012410
Glowacka, Bertram, Mu ¨ller, Soilleux, Pfefferle, Evidence that TMPRSS2 activates the severe acute respiratory syndrome coronavirus spike protein for membrane fusion and reduces viral control by the humoral immune response, Journal of virology, doi:10.1128/JVI.02232-10
Hasan, Paray, Hussain, Qadir, Attar et al., A review on the cleavage priming of the spike protein on coronavirus by angiotensin-converting enzyme-2 and furin, Journal of Biomolecular Structure and Dynamics, doi:10.1080/07391102.2020.1754293
Hoffmann, Kleine-Weber, Schroeder, Kru ¨ger, Herrler et al., SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 and is blocked by a clinically proven protease inhibitor, Cell, doi:10.1016/j.cell.2020.02.052
Hu, Frieman, Insights from nanomedicine into chloroquine efficacy against COVID-19, Nature Nanotechnology, doi:10.1038/s41565-020-0674-9
Keyaerts, Vijgen, Maes, Neyts, Van Ranst, In vitro inhibition of severe acute respiratory syndrome coronavirus by chloroquine, Biochemical, doi:10.1016/j.bbrc.2004.08.085
Li, Greenough, Moore, Vasilieva, Somasundaran et al., Efficient replication of severe acute respiratory syndrome coronavirus in mouse cells is limited by murine angiotensin-converting enzyme 2, Journal of virology, doi:10.1128/JVI.78.20.11429-11433.2004
Li, Moore, Vasilieva, Sui, Wong et al., Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus, Nature, doi:10.1038/nature02145
Liu, Cao, Xu, Wang, Zhang et al., Hydroxychloroquine, a less toxic derivative of chloroquine, is effective in inhibiting SARS-CoV-2 infection in vitro, Cell discovery, doi:10.1038/s41421-019-0132-8
Matsuyama, Nagata, Shirato, Kawase, Takeda et al., Efficient activation of the severe acute respiratory syndrome coronavirus spike protein by the transmembrane protease TMPRSS2, Journal of virology, doi:10.1128/JVI.01542-10
Millet, Whittaker, Host cell entry of Middle East respiratory syndrome coronavirus after two-step, furin-mediated activation of the spike protein, Proceedings of the National Academy of Sciences, doi:10.1073/pnas.1407087111
Ou, Liu, Lei, Li, Mi et al., Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV, Nature communications, doi:10.1038/s41467-019-13993-7
Qi, Qian, Zhang, Zhang, Single cell RNA sequencing of 13 human tissues identify cell types and receptors of human coronaviruses, Biochemical and biophysical research communications, doi:10.1016/j.bbrc.2020.03.044
Sanders, Monogue, Jodlowski, Cutrell, Pharmacologic treatments for coronavirus disease 2019 (COVID-19): a review, Jama, doi:10.1001/jama.2020.6019
Savarino, Boelaert, Cassone, Majori, Cauda, Effects of chloroquine on viral infections: an old drug against today's diseases, The Lancet infectious diseases, doi:10.1016/s1473-3099%2803%2900806-5
Savarino, Trani, Donatelli, Cauda, Cassone, New insights into the antiviral effects of chloroquine, The Lancet infectious diseases, doi:10.1016/S1473-3099%2806%2970361-9
Shang, Ye, Shi, Wan, Luo et al., Structural basis of receptor recognition by SARS-CoV-2, Nature, doi:10.1038/s41586-020-2179-y
Shirato, Kawase, Matsuyama, Middle East respiratory syndrome coronavirus infection mediated by the transmembrane serine protease TMPRSS2, Journal of virology, doi:10.1128/JVI.01890-13
Shirato, Kawase, Matsuyama, Wild-type human coronaviruses prefer cell-surface TMPRSS2 to endosomal cathepsins for cell entry, Virology, doi:10.1016/j.virol.2017.11.012
Simmons, Gosalia, Rennekamp, Reeves, Diamond et al., Inhibitors of cathepsin L prevent severe acute respiratory syndrome coronavirus entry, Proceedings of the National Academy of Sciences, doi:10.1073/pnas.0505577102
Spinelli, Pellino, COVID-19 pandemic: perspectives on an unfolding crisis, The British Journal of Surgery
Sungnak, Huang, Be ´cavin, Berg, Queen et al., SARS-CoV-2 entry factors are highly expressed in nasal epithelial cells together with innate immune genes, Nature medicine, doi:10.1038/s41591-020-0868-6
Vincent, Bergeron, Benjannet, Erickson, Rollin et al., Chloroquine is a potent inhibitor of SARS coronavirus infection and spread, Virology journal, doi:10.1186/1743-422X-2-69
Walls, Park, Tortorici, Wall, Mcguire et al., Structure, function, and antigenicity of the SARS-CoV-2 spike glycoprotein, Cell
Wang, Cao, Zhang, Yang, Liu et al., Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro, Cell research, doi:10.1038/s41422-020-0282-0
Wrobel, Benton, Xu, Roustan, Martin et al., SARS-CoV-2 and bat RaTG13 spike glycoprotein structures inform on virus evolution and furin-cleavage effects, Nature Structural & Molecular Biology
Yao, Ye, Zhang, Cui, Huang et al., In vitro antiviral activity and projection of optimized dosing design of hydroxychloroquine for the treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Clinical Infectious Diseases
Zhang, Jackson, Mou, Ojha, Rangarajan et al., The D614G mutation in the SARS-CoV-2 spike protein reduces S1 shedding and increases infectivity, bioRxiv, doi:10.1101/2020.06.12.148726
Zhang, Penninger, Li, Zhong, Slutsky, Angiotensin-converting enzyme 2 (ACE2) as a SARS-CoV-2 receptor: molecular mechanisms and potential therapeutic target. Intensive care medicine, doi:10.1007/s00134-020-05985-9
Zhao, Zhao, Wang, Zhou, Ma et al., Single-cell RNA expression profiling of ACE2, the putative receptor of Wuhan 2019-nCov
Zhou, Vedantham, Lu, Agudelo, Carrion R Jr et al., Protease inhibitors targeting coronavirus and filovirus entry, Antiviral research, doi:10.1016/j.antiviral.2015.01.011
Ziegler, Allon, Nyquist, Mbano, Miao et al., SARS-CoV-2 receptor ACE2 is an interferon-stimulated gene in human airway epithelial cells and is detected in specific cell subsets across tissues, Cell, doi:10.1016/j.cell.2020.04.035
Zou, Chen, Zou, Han, Hao et al., Single-cell RNA-seq data analysis on the receptor ACE2 expression reveals the potential risk of different human organs vulnerable to 2019-nCoV infection, Frontiers of medicine, doi:10.1007/s11684-020-0754-0
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