2.2  Inhibitors targeting the cellular receptor
The genetic code of SARS‐CoV‐2 shares noticeable similarities with SARS‐CoV‐1, which caused the SARS epidemic in 2002. 26 ,  54  More importantly, both viruses have identical mechanisms of infection. SARS‐CoV‐1 uses the host's ACE2 as a portal to infect cells, which has high expression in the vascular endothelium 55  and the lung, particularly in type 2 alveolar epithelial cells. 56  SARS‐CoV‐2 shares 76% of its spike (S) protein with SARS‐CoV‐1. Despite a few amino acid differences in its RBD compared to the SARS‐CoV‐1 S protein, the SARS‐CoV‐2 S protein binds to ACE2 with even greater affinity 27  offering an explanation for its greater virulence and preference for the lung.
ACE, a highly glycosylated type I integral membrane protein, is an essential component of the renin‐angiotensin (Ang) system, which controls blood pressure homeostasis. Both ACE1 and ACE2 cleave Ang peptides. However, they differ markedly: ACE1 cuts and converts the inactive decapeptide Ang I into the octapeptide Ang II by removing the dipeptide His‐Leu. This Ang II induces vaso‐ and bronchoconstriction, increased vascular permeability, inflammation, and fibrosis, thus promoting acute respiratory distress syndrome (ARDS) and lung failure in patients infected with SARS‐CoV‐1 or SARS‐CoV‐2 57  (Figure 5). Therefore, ACE‐inhibitors (ACEis) and angiotensin II receptor blockers (ARBs) could block the disease‐propagating effect of Ang II. 58 ,  59 ,  60
Figure 5  The roles of ACE1 and 2 in the renin‐angiotensin system. A, Chemical structures of angiotensin‐related peptides and B, Schematic diagram of roles of ACE1 and 2 in renin‐angiotensin system. ACE, angiotensin‐converting enzyme [Color figure can be viewed at wileyonlinelibrary.com] ACE2, on the other hand, is a zinc‐containing metalloenzyme, and shares merely 42% of its amino acid sequence with ACE1. 61  It cleaves only one amino acid residue (Leu or Phe) from Ang I and Ang II, respectively, generating Ang (1–9) and Ang (1–7) (a vasodilator) (Figure 5). Thus, ACE2 has been considered a potential therapeutic target for cardiovascular diseases.
Virtual screening combined with a molecular docking approach targeting the ACE2 catalytic site with around 140 000 compounds led to the identification of inhibitor N‐(2‐aminoethyl)‐1 aziridine‐ethanamine (7; Figure 6) with an IC50 value of 57 µM and a K i of 459 µM. However, no information about the cytotoxicity of this compound is available so far. 62
Figure 6  Inhibitors for SARS‐CoV‐1 and 2 targeting ACE2. ACE, angiotensin‐converting enzyme; SARS‐CoV, severe acute respiratory syndrome coronavirus Chloroquine (8; Figure 6) is a relatively safe, cheap, and effective medication for the treatment of malaria and amebiasis. Savarino et al. 63  reported its antiviral effects. At a molecular level, it increases late endosomal and lysosomal pH, resulting in impaired liberation of virions from endosomes or lysosomes. The virus is therefore unable to release its genetic material into the cell and replicate. 64 ,  65  Furthermore, they hypothesize that chloroquine might block the production of proinflammatory cytokines (such as IL‐6), thereby blocking the pathway that subsequently leads to ARDS. 63
Chloroquine is reasonably active in vitro against SARS‐CoV‐1, MERS‐CoV, and SARS‐CoV‐2. It was found to inhibit SARS‐CoV‐2 with an EC50 value of 5.47 µM in vitro. 66  Antiviral activity against SARS‐CoV‐1 was reported with an IC50 of 8.8 μM in Vero cells, but it is unclear how this translates into activity in respiratory epithelial cells and in vivo. 67 ,  68  Mechanistic studies of chloroquine for SARS‐CoV‐1 infection revealed that it could also weaken the interaction between the RBD of SARS‐CoV‐1 and ACE2 by interfering with terminal glycosylation of ACE2, thereby reducing its affinity to SARS‐CoV‐1 S. 69
During the SARS‐CoV‐2 pandemic, chloroquine has been recommended by Chinese, South Korean, and Italian health authorities for the experimental treatment of COVID‐19, 70 ,  71  despite contraindications for patients with heart disease or diabetes. 72  However, health experts and agencies like the US FDA and European Medicines Agency warned against broad uncontrolled use after reports of misuse of low‐quality versions of chloroquine phosphate intended for fish.
Hydroxychloroquine (9; Figure 6) is being studied as an experimental treatment for COVID‐19. 73  However, the benefits of treatment with this drug are unclear. 74
Hydroxychloroquine was found to inhibit SARS‐CoV‐2 with an EC50 value of 0.74 µM in vitro. 66  Some studies imply synergistic effects of hydroxychloroquine and azithromycin. Azithromycin is active in vitro against Zika and Ebola virus 75 ,  76  and can be used to guard against life‐threatening bacterial superinfections when administered to patients suffering from viral infections. 77  A small study that compared hydroxychloroquine monotherapy and combination treatment with azithromycin found a significant advantage of the combination. While evaluating the efficacy of therapeutic intervention with hydroxychloroquine as monotherapy and its impact in combination with azithromycin, the number of patients testing negative in polymerase chain reaction (PCR) tests was substantially different in the two groups with 100% of patients cured (6 days post inclusion) in the combination arm of the study versus 57% in the monotherapy group. At the same time, 12% of patients in the control group receiving only standard care were cured. 78 ,  79
The WHO declared on 18 March that chloroquine and its derivative hydroxychloroquine will be among the four medicines studied in the solidarity clinical trial 80  for the treatment of COVID‐19. In April 2020, the US National Institutes of Health (NIH) also commenced a study with the drug for treating COVID‐19 patients. 81
The recent clinical trial involving 96 032 patients with COVID‐19 concluded that it was unable to confirm a benefit of hydroxychloroquine or chloroquine, when used alone or in combination with a macrolide such as azithromycin (or clarithromycin). 82  The study actually reported decreased survival rates for patients treated with each of these drug regimens. Additionally, patients had an increased risk of developing ventricular arrhythmia under treatment. However, still more evidence is needed to adequately assess the drugs' risks or benefits for the treatment or prevention of COVID‐19 (it is important to note that chloroquine and hydroxychloroquine are still considered safe treatment options in certain autoimmune diseases and malaria). Besides, the WHO announced the premature pause of its clinical trials using hydroxychloroquine as a safety precaution on 24 May 2020.
On a different note, it was found that ACE2 undergoes proteolytic shedding; releasing an enzymatic ectodomain during viral entry. 83  A disintegrin and metalloproteinase (ADAM), also known as TNF‐α converting enzyme (TACE), assisted the shedding regulation of ACE2. Inhibition of this enzyme led to reduced shedding of ACE2. GW280264X (10; Figure 6) was found to be a specific inhibitor of ADAM‐induced shedding of ACE2 at 1 nM. 84  Two TACE inhibitors, TAPI‐0 (11) and TAPI‐2 (12; Figure 6), reduced ACE2 shedding, with IC50 values of 100 and 200 nM, respectively. 83
MLN‐4760 (13; Figure 6) inhibited the catalytic activity of ACE2 with an IC50 of around 440 pM. 85  This is the most potent and selective small‐molecule inhibitor against soluble human ACE2 described to date, thus making it a very promising candidate for SARS‐CoV‐2 interference. It binds to the active site zinc and emulates the transition state peptide. However, no antiviral data for this compound is available at this time.
The interference of a virus‐host cell fusion, which is mediated by the viral S protein to its receptor ACE2 on host cells, may be a viable prevention strategy. Umifenovir (14; brand name Arbidol), a broad spectrum antiviral drug used against influenza, prevents viral entry by inhibiting virus‐host cell fusion. 86  It is currently being investigated in a clinical trial for the treatment of SARS‐CoV‐2. 87 ,  88
Do ACEis or ARBs amplify SARS‐CoV‐2 pathogenicity and aggravate the clinical course of COVID‐19? After ACE2 was recognized as the SARS‐CoV‐2 receptor, 14 ,  29  speculations emerged about potentially negative consequences of ACEi or ARB therapy in COVID‐19 patients. This theory caused confusion in the public and alarmed patients taking these medicines. One report said that the expression of ACE2 was increased in patients with heart disease compared to healthy individuals. It was also insisted that ACE2 expression could be increased by taking ACEis and ARBs, 89  although there is no supporting report of this happening in the lungs.
In another report, it was suggested that patients suffering from high blood pressure receiving “ACE2‐increasing drugs” have a higher risk for severe COVID‐19, since ACEis and ARBs could elevate levels of ACE2. 90
A joint declaration by the presidents of the HFSA/ACC/AHA on 17 March 2020, 91  followed by a similar statement of the European Medicines Agency, 92  clarified that there was no scientific basis for stopping ACEi or ARB therapy. 93 ,  94 ,  95  This was in accordance with the editors of the New England Journal of Medicine. 96
In case of SARS‐CoV, the experimental data showed that such medications may be beneficial rather than damaging, which led to a new therapeutic approach for lung diseases. 97