Coronavirus Susceptibility to the Antiviral Remdesivir

AlexInWonderland
February 5, 2020

Coronaviruses (CoVs) can cause significant disease in humans, as was evident in the severe acute respiratory syndrome (SARS) outbreak in 2002, the Middle Eastern respiratory syndrome (MERS) outbreak in 2012, and the ongoing outbreak in Central China of a yet to be genetically defined CoV (2). Currently, there are no approved therapeutics available, however, Agostini et al. details the effects of a nucleoside analogue that potently inhibits human and zoonotic CoVs in vitro. Remdesivir (GS-5734) acts as an inhibitor of viral replication and inhibits murine hepatitis virus (MHV), SARS-CoV, and MERS-CoV with similar 50% effective concentration values (EC50). Given the emergence potential of CoVs to cause deadly disease, novel antivirals will be critical in preventing the next epidemic. 

CoVs belong to the order Nidovirales and are enveloped, non-segmented, positive sense, single-stranded RNA viruses. CoVs account for the largest identified RNA genomes, with most being approximately 30kb. Two thirds of the genome encode for the replicase-nonstructural proteins, with the remaining third encoding for the structural units: spike (S), envelope (E), membrane (M), and nucleocapsid (N) (3). Nonstructural protein 14, ExoN, mediates exoribonuclease activity and provides a level of resistance to GS-5734.  At 0.25 µM of drug, MHV ExoN knockout strain (ExoN-) exhibits a 100-fold greater reduction in viral titer when compared to wild type (WT) MHV. The EC50 of ExoN- MHV was 0.019 μM, a 4.5-fold decrease compared to WT EC50 of 0.087 µM.  This increased sensitivity of ExoN- MHV is suggestive of a mechanism of action for GS-5734 that involves incorporation of drug into viral RNA that can be removed by the proofreading activity of ExoN.  Additionally, the EC50 for both SARS-CoV and MERS-CoV was approximately 0.074 μM for both viruses (1).

Following 23 passages of WT MHV in the presence of increasing concentrations of GS-441524, the parent nucleoside analogue to GS-5734, increased viral cytopathic effect (CPE) was observed, indicative of the virus’s ability to replicate.  GS-441524 and GS-5734 are metabolized in the same manner, but GS-441524 has a broader range of working concentrations and thus was used for this particular assay (6). Full genome sequencing revealed 6 non-synonymous mutations, two of which were found in the predicted fingers domain of the conserved right-hand structure of the RNA dependent RNA polymerase (F476L and V553L) (4,5).  Engineered MHV containing either mutation individually was more sensitive to GS-5734 than WT MHV, while recombinant MHV containing both mutations displayed a level of resistance similar to that of the P23 lineage.  In comparison to WT MHV, recombinant F476L MHV, V553L MHV, and F476L + V553L MHV displayed a 2.4, 5, and 5.6-fold resistance to GS-5734. However, recombinant F476L + V553L MHV could not compete with WT MHV via co-infection of murine DBT cells in the absence of GS-5734, suggesting that the increased resistance to drug comes at a fitness cost to the virus (1).

Broadly reactive antivirals against human CoV infections, such as the recently confirmed outbreak in East Asia, are imperative in the fight to control and prevent potential epidemics. Agostini et al. provides conclusive evidence that GS-5734 is highly active against CoVs and that resistant viruses endure a loss of competitive fitness in vitro. The pro-drug Remdesivir, GS-5734, has proved to be a promising contender in the family of small molecule, nucleoside analogue drugs. 

 

Resources:

  1. Agostini, M. L., Andres, E. L., Sims, A. C., Graham, R. L., Sheahan, T. P., Lu, X., Smith, E. C., Case, J. B., Feng, J. Y., Jordan, R., Ray, A. S., Cihlar, T., Siegel, D., Mackman, R. L., Clarke, M. O., Baric, R. S., & Denison, M. R. (2018). Coronavirus susceptibility to the antiviral remdesivir (GS-5734) is mediated by the viral polymerase and the proofreading exoribonuclease. MBio, 9(2), e00221-18, /mbio/9/2/mBio.00221-18.atom. https://doi.org/10.1128/mBio.00221-18
  2. Coronavirus | home | cdc. (2020, January 16). https://www.cdc.gov/coronavirus/index.html
  3. Fehr, A. R., & Perlman, S. (2015). Coronaviruses: An overview of their replication and pathogenesis. In H. J. Maier, E. Bickerton, & P. Britton (Eds.), Coronaviruses (Vol. 1282, pp. 1–23). Springer New York. https://doi.org/10.1007/978-1-4939-2438-7_1
  4. Sexton, N. R., Smith, E. C., Blanc, H., Vignuzzi, M., Peersen, O. B., & Denison, M. R. (2016). Homology-based identification of a mutation in the coronavirus rna-dependent rna polymerase that confers resistance to multiple mutagens. Journal of Virology, 90(16), 7415–7428. https://doi.org/10.1128/JVI.00080-16
  5. Xu, X. (2003). Molecular model of SARS coronavirus polymerase: Implications for biochemical functions and drug design. Nucleic Acids Research, 31(24), 7117–7130. https://doi.org/10.1093/nar/gkg916
  6. Warren TK, Jordan R, Lo MK, Ray AS, Mackman RL, Soloveva V, Siegel D, Perron M, Bannister R, Hui HC, Larson N, Strickley R, Wells J, Stuthman KS, Van Tongeren SA, Garza NL, Donnelly G, Shurtleff AC, Retterer CJ, Gharaibeh D, Zamani R, Kenny T, Eaton BP, Grimes E, Welch LS, Gomba L, Wilhelmsen CL, Nichols DK, Nuss JE, Nagle ER, Kugelman JR, Palacios G, Doerffler E, Neville S, Carra E, Clarke MO, Zhang L, Lew W, Ross B, Wang Q, Chun K, Wolfe L, Babusis D, Park Y, Stray KM, Trancheva I, Feng JY, Barauskas O, Xu Y, Wong P, Braun MR, Flint M, McMullan LK, Chen S-S, Fearns R, Swaminathan S, Mayers DL, Spiropoulou CF, Lee WA, Nichol ST, Cihlar T, Bavari S. 2016. Therapeutic efficacy of the small molecule GS-5734 against Ebola virus in rhesus monkeys. Nature 531:381–385. https://doi.org/10.1038/nature17180