2.3  Proteolytic processing inhibitors
CoVs enter the host cells via both clathrin (endosomal) and nonclathrin pathways (nonendosomal); however, both pathways are dependent upon receptor binding. 98 ,  99
The clathrin‐mediated pathway involves the binding of CoV S protein to the host receptor followed by the internalization of vesicles that maturate to late endosomes. Acidification of the endosome promotes the H+‐dependent activation of cellular cathepsin L proteinase in late endosomes and lysosomes, which cleaves and activates the S protein, thus initiating viral fusion. Recent research shows that in addition to ACE2 SARS‐CoV‐2 can also use the host cell receptor CD147 to gain access into host cells. 100
Membrane fusion is also the crucial step for the CoV life cycle in the nonclathrin/endosomal route, in which host proteases such as cathepsin L, TMPRSS2, and TMPRSS11D (airway trypsin‐like protease) cut the S protein at the S1/S2 cleavage site to activate the S protein for membrane fusion. 101  Interference with this process by targeting these proteases could become an attractive strategy for combating CoV infections. A recent study confirms the role of TMPRSS2 for the viral life cycle in SARS‐CoV‐2‐infected VeroE6 cells. 5  Furin (a serine endoprotease) activates MERS‐CoV to initiate the nonclathrin mediated membrane fusion event. 102
The neurotransmitter receptor blockers chlorpromazine (15), promethazine (16), and fluphenazine (17; Figure 7), were reported to inhibit MERS‐CoV and SARS‐CoV‐1 most probably by impeding S protein‐induced fusion. 103  Chlorpromazine, a clathrin‐mediated viral entry inhibitor, was already described to inhibit human CoV‐229E, hepatitis C virus, infectious bronchitis virus, as well as mouse hepatitis virus‐2 (MHV2). 104 ,  105 ,  106 ,  107 ,  108
Figure 7  Neurotransmitter inhibitors targeting clathrin/nonclathrin pathways Matsuyama et al. identified the commercially available serine protease inhibitor camostat (18; Figure 8) to be a SARS‐CoV‐1 inhibitor, blocking TMPRSS2 activity at 10 µM. However, at a higher concentration (100 µM), inhibition of viral entry via SARS‐CoV‐1 S protein‐mediated cell fusion never exceeded 65% (inhibition efficiency), indicating that 35% of entry events take place via the endosomal cathepsin pathway. Interestingly, treatment with a combination of EST (a cathepsin inhibitor) and 18 resulted in remarkably blocked infection (>95%) activity of pseudotyped viruses. 109
Figure 8  Inhibitors targeting TMPRSS2. TMPRSS, transmembrane serine protease A similar approach has been investigated to prevent viral entry of SARS‐CoV‐2. Pöhlmann et al. reported the attainment of full inhibition efficiency with a combination of both 18 and E‐64d (a cathepsin inhibitor). Both studies indicate that SARS‐CoV‐1 and 2 enter cells in a similar manner showing the potential of 18 as a candidate for further development. 15
Recently, K11777 (19; Figure 8), a cysteine protease inhibitor, was shown in tissue cultures to inhibit SARS‐CoV‐1 and MERS‐CoV replication in the subnanomolar range. 110 ,  111  Future tissue culture and animal model studies should be conducted to clarify, whether its antiviral activity is mediated by targeting TMPRSS2.
Teicoplanin is a glycopeptide antibiotic used to prevent infections with Gram‐positive bacteria like methicillin‐resistant Staphylococcus aureus and Enterococcus faecalis. It was found that teicoplanin inhibits the entry of SARS‐CoV‐1, MERS‐CoV, and Ebola virus by specifically targeting cathepsin L. 112  This knowledge has also been used to block the entry of new SARS‐CoV‐2 pseudoviruses with an IC50 value of 1.66 µM. Therefore, teicoplanin could be considered a potential candidate for the treatment of COVID‐19. 113