Blockade of SARS-CoV-2 infection by recombinant soluble ACE2

Abstract

Viruses use specific host-cell plasma membrane molecules as receptors to enter the cell. SARS-CoV-2, the coronavirus responsible for the current COVID-19 pandemic, is believed to bind mainly or exclusively to angiotensin-converting enzyme 2 (ACE2), a cell-surface protein otherwise known for its enzymatic, carboxypeptidase-type activity and its physiological role in the renin-angiotensin system.1Wrapp D. Wang N. Corbett K.S. et al.Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation.Science. 2020; 367: 1260-1263Crossref PubMed Scopus (95) Google Scholar,2Wan Y. Shang J. Graham R. et al.Receptor recognition by the novel coronavirus from Wuhan: an analysis based on decade-long structural studies of SARS coronavirus.J Virol. 2020; 94 (e00127-20)Crossref Scopus (3201) Google Scholar ACE2 activity per se does not seem to be involved in infectivity, but its role in lung, heart, and kidney function has putative consequences for disease severity. ACE2 is a transmembrane protein with a large extracellular N-terminal domain bearing the active site. It was discovered through its structural homology with the dipeptidyl-peptidase angiotensin I-converting enzyme (kininase II, ACE). ACE2 removes the carboxyterminal amino acid of angiotensin II (and also, but with reduced efficiency, of angiotensin I), inactivating angiotensin II and generating angiotensin 1-7.3Danilczyk U. Penninger J.M. Angiotensin-converting enzyme II in the heart and the kidney.Circ Res. 2006; 98: 463-471Crossref PubMed Scopus (229) Google Scholar Angiotensin 1-7, which activates a specific receptor, MAS, is a vasodilator and has antioxidant and anti-inflammatory properties.4Varagic J. Ahmad S. Nagata S. et al.ACE2: angiotensin II/angiotensin-(1–7) balance in cardiac and renal injury.Curr Hypertens Rep. 2014; 16: 420Crossref PubMed Scopus (95) Google Scholar ACE2 is thus considered to have a counter-regulatory role, shifting to some extent the renin-angiotensin system from a vasoconstrictor to a vasodilator, endothelium-activator system, with favorable consequences for end-organ protection in experimental settings such as ischemia or diabetes. Both SARS-CoV-2 and the coronavirus responsible for a more limited pandemic of respiratory syndromes 17 years ago, SARS-CoV, use ACE2 for host cell entry and infection.1Wrapp D. Wang N. Corbett K.S. et al.Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation.Science. 2020; 367: 1260-1263Crossref PubMed Scopus (95) Google Scholar,2Wan Y. Shang J. Graham R. et al.Receptor recognition by the novel coronavirus from Wuhan: an analysis based on decade-long structural studies of SARS coronavirus.J Virol. 2020; 94 (e00127-20)Crossref Scopus (3201) Google Scholar Nonetheless, there does not seem to be strong evolutionary rationale for these viruses targeting ACE2 and the renin-angiotensin system, as another coronavirus involved in respiratory disease, MERS-CoV, does not bind ACE2 but dipeptidyl peptidase IV, another membrane-bound peptidase with different enzymatic specificity. Both peptidases are expressed in alveolar epithelial cells of the lung, although not in great abundance. In contrast, they are abundant in the kidney, especially in the proximal tubule. They are also present in podocytes. ACE2 is also expressed in the heart and is involved in myocardial function.3Danilczyk U. Penninger J.M. Angiotensin-converting enzyme II in the heart and the kidney.Circ Res. 2006; 98: 463-471Crossref PubMed Scopus (229) Google Scholar These organs might therefore be particularly prone to the effects of coronavirus infection. To date, no specific treatment is available for COVID-19. The race is on among academic teams and pharmaceutical companies to identify effective agents and develop a vaccine. Besides strategies targeting the virus with antibodies to viral proteins or chemical compounds aimed at inhibiting viral replication, another approach is to focus on the cellular target of the virus, ACE2. Binding of SARS-Cov-2 to ACE2 involves the viral Spike protein, which protrudes from the viral surface and is activated by human proteases. The molecular interaction between ACE2 and Spike has been modeled,1Wrapp D. Wang N. Corbett K.S. et al.Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation.Science. 2020; 367: 1260-1263Crossref PubMed Scopus (95) Google Scholar,2Wan Y. Shang J. Graham R. et al.Receptor recognition by the novel coronavirus from Wuhan: an analysis based on decade-long structural studies of SARS coronavirus.J Virol. 2020; 94 (e00127-20)Crossref Scopus (3201) Google Scholar and synthetic compounds or antibodies interfering in that interaction could be designed. Protease inhibitors preventing Spike activation could also be tested. Yet another therapeutic approach is the use of soluble ACE2 as a virus trap and inactivator. Soluble ACE2 generated by proteolytic cleavage of the membrane anchor is normally present in plasma, although at low concentration. Increasing its availability at tissue sites would shift the competition with membrane-bound ACE2 toward the soluble protein that cannot promote viral entry into the cell. It is also expected that this approach would preserve tissue ACE2.5Monteil V, Kwon H, Prado P, et al. Inhibition of SARS-CoV-2 infections in engineered human tissues using clinical-grade soluble human ACE2 [e-pub ahead of print]. Cell. https://doi.org/10.1016/j.cell.2020.04.004. Accessed April 2, 2020.Google Scholar,6Batlle D. Wysocki J. Satchell K. Soluble angiotensin-converting enzyme 2: a potential approach for coronavirus infection therapy?.Clin Sci (Lond). 2020; 134: 543-545Crossref PubMed Scopus (347) Google Scholar This sound theoretical rationale has now received experimental support from in vitro studies by Monteil et al.5Monteil V, Kwon H, Prado P, et al. Inhibition of SARS-CoV-2 infections in engineered human tissues using clinical-grade soluble human ACE2 [e-pub ahead of print]. Cell. https://doi.org/10.1016/j.cell.2020.04.004. Accessed April 2, 2020.Google Scholar in SARS-CoV-2–infected monkey kidney-derived epithelial cells and engineered human blood vessel and kidney organoids. The investigators isolated the SARS-CoV-2 from a nasopharyngeal sample of a patient with confirmed COVID-19. After successful culture on Vero-E6 cells (a line of African green monkey kidney cells), the isolated virus was sequenced by next-generation sequencing and showed the prototypic coronal shape of viral particles by electron microscopy. The investigators then infected the Vero-E6 cells with SARS-CoV-2 and examined the effect of human recombinant soluble ACE2 (hrsACE2) added to the culture milieu. They found that exposure of infected cells to hrsACE2 during the first hour, followed by washing and incubation without hrsACE2, significantly inhibited SARS-CoV-2 replication 15 hours post infection. This inhibition was dependent on the initial quantity of the virus in the inoculum and the concentration of hrsACE2, establishing dose-dependency. In contrast to hsrACE2, mouse recombinant soluble ACE2 did not inhibit the infection. Because the virus has a size of 80 to 100 nm, indicating that SARS-CoV-2 must first infect blood vessels prior to local tissue infections, the investigators went on to infect human capillary organoids with the SARS-CoV-2 isolate. After demonstrating active viral replication in this experimental model, they found that the addition of hrsACE2 markedly reduced SARS-CoV-2 infection. Finally, because ACE2 is strongly expressed in tubular epithelium, the investigators examined the effect of hrsACE2 in kidney organoids generated from human embryonic stem cells. They first demonstrated ACE2 expression and active SARS-CoV-2 replication in organoids composed of proximal tubular cell and podocyte clusters. They then added hrsACE2 to the culture medium and found that it reduced SARS-CoV-2 infection in a dose-dependent manner. What is still lacking is the demonstration that hrsACE2 can also protect pulmonary cells against SARS-CoV-2 infection. Figure 1 presents a schematic view of the renin-angiotensin system, emphasizing ACE2 with its 2 molecular forms, membranous and soluble, and their putative role in SARS-CoV-2 cellular infection and virus trapping, respectively. These observations are exciting in view of their potential clinical relevance. It remains to be seen whether hrsACE2 administration in vivo can overcome massive SARS-CoV-2 infection, considering also the stoichiometry of the association, with several Spike proteins per viral particle. There is some hope that this will be tested clinically, because hrsACE2 has been approved for human use and is currently being developed as treatment for acute respiratory distress syndrome (ARDS). A pharmacokinetic study in healthy volunteers and a phase II clinical trial in patients with ARDS have been completed.7Haschke M. Schuster M. Poglitsch M. et al.Pharmacokinetics and pharmacodynamics of recombinant human angiotensin- converting enzyme 2 in healthy human subjects.Clin Pharmacokinet. 2013; 52: 783-792Crossref PubMed Scopus (295) Google Scholar,8Khan A. Benthin C. Zeno B. et al.A pilot clinical trial of recombinant human angiotensin-converting enzyme 2 in acute respiratory distress syndrome.Crit Care. 2017; 21: 234Crossref PubMed Scopus (481) Google Scholar Studies have suggested that ACE2 protects the lung in experimental ARDS. This includes syndromes caused by chemical injury of the lung, bacterial infection, or infection by influenza virus or respiratory syncytial virus (see bibliography in Khan et al.8Khan A. Benthin C. Zeno B. et al.A pilot clinical trial of recombinant human angiotensin-converting enzyme 2 in acute respiratory distress syndrome.Crit Care. 2017; 21: 234Crossref PubMed Scopus (481) Google Scholar). Genetic ACE2 deficiency worsens the condition in mice, while administration of recombinant ACE2 in mice or pigs improves experimental ARDS. Conversely, inactivation of ACE or angiotensin II AT1 receptor protects the lung. Genetic variation in ACE level has also been reported to be a prognostic factor for ARDS in humans. The balance between angiotensin II and angiotensin 1-7 may play a role in alveolar function in disease, with angiotensin II exerting deleterious and angiotensin 1-7 beneficial actions, but this remains to be further studied. In any case, SARS-CoV infection and Spike activation result in ACE2 depletion, which might contribute to the severity of the respiratory disease in COVID-19. ACE2 has also been shown to exert beneficial action in noninfectious experimental kidney disease, especially diabetic nephropathy, probably through both inactivation of angiotensin II and formation of angiotensin 1-7. Genetic deficiency of ACE2 in mice aggravates kidney pathology, and use of recombinant ACE2 has been advocated for treatment of angiotensin II-mediated kidney diseases.6Batlle D. Wysocki J. Satchell K. Soluble angiotensin-converting enzyme 2: a potential approach for coronavirus infection therapy?.Clin Sci (Lond). 2020; 134: 543-545Crossref PubMed Scopus (347) Google Scholar SARS-CoV-2 can infect the kidney,9Su H, Yang M, Wan C, et al. Renal histopathological analysis of 26 postmortem findings of patients with COVID-19 in China [e-pub ahead of print]. Kidney Int. 10.1016/j.kint.2020.04.003. Accessed April 9, 2020.Google Scholar and the study by Monteil et al.5Monteil V, Kwon H, Prado P, et al. Inhibition of SARS-CoV-2 infections in engineered human tissues using clinical-grade soluble human ACE2 [e-pub ahead of print]. Cell. https://doi.org/10.1016/j.cell.2020.04.004. Accessed April 2, 2020.Google Scholar shows that hrsACE2 dramatically reduces SARS-CoV-2 infection of human kidney organoids. In SARS-CoV-2 infection, ACE2 depletion might also aggravate preexisting kidney disease. The rationale for hrsACE2 therapy in COVID-19 is based primarily on its expected inhibitory effect on pulmonary infection, but it may have additional beneficial effects in the lung and other organs such as the kidney. It is noteworthy that sACE2 administration was well tolerated in clinical phase I and phase II trials in ARDS.7Haschke M. Schuster M. Poglitsch M. et al.Pharmacokinetics and pharmacodynamics of recombinant human angiotensin- converting enzyme 2 in healthy human subjects.Clin Pharmacokinet. 2013; 52: 783-792Crossref PubMed Scopus (295) Google Scholar,8Khan A. Benthin C. Zeno B. et al.A pilot clinical trial of recombinant human angiotensin-converting enzyme 2 in acute respiratory distress syndrome.Crit Care. 2017; 21: 234Crossref PubMed Scopus (481) Google Scholar Because ACE2 therapy is expected to result in angiotensin II depletion, attention should be paid to the impact on blood pressure and kidney function. Although additional studies are needed, the potentially protective action of hrsACE2 against viral infection gives new hope in the fight against COVID-19. TBD reports having received advisor, consultant, speaker, and/or travel fees from Akebia, Amgen, Astellas, Chugai, F. Hoffman-La Roche, FMC, GlaxoSmithKline, Kyowa Hakko Kirin, and Vifor. The other author declared no competing interests. Renal histopathological analysis of 26 postmortem findings of patients with COVID-19 in ChinaKidney InternationalVol. 98Issue 1PreviewAlthough the respiratory and immune systems are the major targets of Coronavirus Disease 2019 (COVID-19), acute kidney injury and proteinuria have also been observed. Currently, detailed pathologic examination of kidney damage in critically ill patients with COVID-19 has been lacking. To help define this we analyzed kidney abnormalities in 26 autopsies of patients with COVID-19 by light microscopy, ultrastructural observation and immunostaining. Patients were on average 69 years (19 male and 7 female) with respiratory failure associated with multiple organ dysfunction syndrome as the cause of death. Full-Text PDF Open Access

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