4  RdRP AND ITS INHIBITORS
The ability to produce new RNA copies from available template molecules is necessary for life on earth. RNA polymerases are therefore found in all living cells as well as many viruses. RdRP are essential enzymes to all RNA viruses, as they catalyze the synthesis of new RNA from a given RNA template. 206  Due to their importance for viral life cycles, and their high conservation among different RNA viruses, they have been attractive drug targets for antiviral therapy for a long time.
SARS‐CoV‐2 also uses an RdRP to replicate its genome within the host cell. Three nonstructural viral proteins (nsp) form its replication/transcription complex, with nsp12 forming the catalytic subunit. Bound to it are nsp7 and nsp8—accessory factors that facilitate template binding. 207 ,  208 ,  209  Their individual structures and that of the complex have been solved. 210 ,  211 ,  212 ,  213 ,  214  Interestingly, the only nsp that interacts directly with RNA seems to be nsp12, whereas nsp7 and nsp8 are needed to increase its efficiency. 210 ,  215
RdRP is the target of inhibitors like remdesivir (178), galidesivir (179), ribavirin (180), favipiravir (181), and EIDD‐2801 (182). These molecules have shown promise for the treatment of COVID‐19 patients. 87 ,  216 ,  217 ,  218  (For structures and biological data see Figure 38)
Figure 38  Nucleoside analogs with inhibition activity against SARS‐CoV‐2 RdRP (adenosine, guanosine, and sofosbuvir are included for comparison). RdRP, RNA‐dependent RNA polymerase; SARS‐CoV, severe acute respiratory syndrome coronavirus Remdesivir is a 1'‐cyano‐substitued adenine C‐nucleoside analog prodrug. The prodrug strategy used is similar to that of the FDA‐approved anti‐hepatitis C drug sofosbuvir (183; see Figure 38). Upon its diffusion into cells, the phosphoramidate undergoes an intracellular conversion process that results in the formation of the triphosphate active metabolite (RTP). The triphosphate is recognized as adenine by viral RdRP, which causes heavy disruptions in RNA synthesis.
The exact molecular mechanism of remdesivir's action against SARS‐CoV‐2 has recently been elucidated by Yin et al. 219  who reported cryo‐EM structures of SARS‐CoV‐2 RdRP with remdesivir monophosphate (RMP) covalently bound to the primer strand. As only a single RMP was incorporated in each observed primer strand the inhibition mechanism was shown to be nonobligate RNA chain termination. The addition of RTP led to a complete inhibition of RNA polymerization activity at a concentration of 1 mM, even in the presence of ATP in high concentrations of 100 mM. The authors further highlight the high conservation level of catalytic sites of RdRPs in different RNA viruses, which makes the discovery of future broad‐spectrum antiviral RdRP inhibitors seem likely.
Remdesivir was previously reported to inhibit SARS‐CoV‐1 and MERS‐CoV replication in multiple in vitro systems, with submicromolar IC50 values. 220  In primary human airway epithelial (HAE) cell cultures, the antiviral activity assessment of remdesivir against SARS‐CoV‐1 and MERS‐CoV showed a dose‐dependent reduction in replication with average IC50 values of 0.069 µM (SARS‐CoV‐1) and 0.074 µM (MERS‐CoV). In a mouse model of SARS‐CoV‐1 pathogenesis, remdesivir greatly decreased the virus concentration in the lung and mitigated clinical symptoms of infection and restored respiratory function.
Galidesivir is another C‐nucleoside analog that resembles adenosine. However, the base is not linked to a ribose, but to an aza‐sugar. Although it is recognized as adenosine by RdRP, its properties are different enough to cause a disruption in chain elongation. Galidesivir has been used in the treatment of Ebola and Marburg virus infections, and in vitro studies against SARS‐ and MERS‐CoVs have suggested efficacy against CoVs. 221  Therefore, it is a likely future anti SARS‐CoV‐2 agent, and currently being studied in clinical trials. 222 ,  223
Ribavirin is a nucleoside analog, which shows structural similarity to guanosine. But guanosine's 6‐membered ring is only hinted at by the amide group. As such, it is incorporated by viral RdRPs, but interrupts RNA polymerization. 224  It is an approved drug in most countries and used against a variety of viral infections. Although its efficacy against SARS‐CoV‐2 has not been determined in large clinical trials, ribavirin has shown some promise in the treatment of COVID‐19 patients. 225
Favipiravir (Avigan®) is an approved antiviral drug for the treatment of influenza in Japan and China. It is a pyrazinamide derivative that has shown some activity against a variety of RNA viruses. 226  Favipiravir inhibits viral RdRP via its similarity to guanine. After biotransformation into its active metabolite, favipiravir‐ribofuranosyl‐5'‐triphosphate, it is incorporated into newly synthesized RNA by RdRP, leading to premature chain termination 227  similar to remdesivir's mode of action. Favipiravir is currently being studied around the world as a treatment option against COVID‐19.
Very recently, Sheahan et al. 228  reported the discovery of EIDD‐1931 and its orally bioavailable prodrug EIDD‐2801. These nucleoside analogs have shown remarkable potency against SARS‐CoV‐2 and other related CoVs in vitro and in vivo, with IC50 values in the low nanomolar range, outperforming remdesivir 3–10‐fold. The reason for this increased potency could be additional interactions with viral RdRP involving the N4‐hydroxyl group of the cytidine ring. 219  The efficacy of EIDD‐2801 in COVID‐19 patients is being evaluated in a clinical trial. 229
Remdesivir and other potential RdRP inhibitors 230  are currently being studied in clinical trials around the world, but even though preliminary results appear promising, it is too early to assess their clinical value against COVID‐19.