Mutagenic antivirals: the evolutionary risk of low doses

Mutagenic antivirals: the evolutionary risk of low doses

Chase W. Nelson1, Sarah P. Otto2

  1. Institute for Comparative Genomics, American Museum of Natural History, New York, NY 10024, USA; [email protected]
  2. Department of Zoology, University of British Columbia, Vancouver BC Canada V6T 1Z4; [email protected]

Vaccines and antiviral drugs both have critical parts to play in the fight against COVID-19. Oral antivirals are of particular interest given their potential for equitable distribution, the occurrence of vaccine-breakthrough infections, and increasingly transmissible variants of concern.

Merck recently announced the mutagenic (mutation-inducing) antiviral molnupiravir, which improves COVID-19 outcomes when administered twice a day for five days (Merck & Co., Inc. 2021a; Merck & Co., Inc. 2021b). The drug is already authorized for emergency use in the U.K., and the U.S. Food and Drug Administration will meet to discuss emergency use authorization on Tuesday, November 30.

Antivirals can work via several mechanisms. Protease inhibitors, like masitinib (Drayman et al. 2021) and paxlovid (PF-07321332; ritonavir; Pfizer, Inc. 2021), prevent the production of mature viral proteins. Mutagens, like molnupiravir, instead increase the viral mutation rate beyond a tolerable limit. Specifically, molnupiravir is a nucleoside analog that incorporates into replicating RNA, preferentially inducing C→U mutations (Gordon et al. 2021). At the recommended doses, it is proposed to cause lethal mutagenesis (Loeb et al. 1999): newly replicated viral genomes accumulate so many errors as to become nonviable, an endpoint known as error catastrophe (Eigen and Schuster 1977) or mutational meltdown (Lynch and Gabriel 1990).

One drawback of self-administered oral medications is the risk of low drug concentrations due to missed doses, incomplete courses, or low initial drug penetrance at the site of action (e.g., mucosal membranes in the nasal passages or lungs). Critically, in the case of mutagenic antivirals like molnupiravir, low drug concentrations might increase the mutation rate without reaching the level required for error catastrophe, instead inducing only sublethal mutagenesis (Sadler et al. 2010). This could accelerate within-host evolution of the virus, potentiating new variants of concern that enhance transmissibility or immune escape (Pillai et al. 2008). Indeed, another antiviral, ribavirin, has mutagenic properties and induces adaptive mutations in other RNA viruses (Beaucourt and Vignuzzi 2014; Mejer et al. 2020). Moreover, previous SARS-CoV-2 variants of concern likely acquired adaptive combinations of mutations during single chronic infections (Kemp et al. 2021; Otto et al. 2021).

Given the potential for sublethal mutagenesis of SARS-CoV-2, steps should be taken to understand the evolutionary consequences of both drug concentration and improper administration for pathogen evolution. Among the issues to consider are:

  1. Peak viral shedding is likely to coincide with low initial drug concentration. Molnupiravir will often be used by patients with recent symptom onset, when viral shedding is near its peak. Because the drug is initially absent and builds over time, peak viral shedding is therefore likely to occur when there are still low drug concentrations at the sites of action (i.e., during the first days of administration). This could potentiate the release of mutant—but viable—virus.
  2. Molnupiravir has a short plasma half-life. Molnupiravir’s short plasma half-life (Painter et al. 2021) makes low concentrations easier to achieve, e.g., as a result of missed or inconsistently timed doses.
  3. Coronaviruses have a propensity for recombination. Recombination can help purge viral genomes of deleterious mutations, or generate adaptive combinations of beneficial or compensatory mutations. Thus, it is critical to include recombination and interactions between mutations (i.e., epistasis) in models and simulations.
  4. Sequence content limits mutation rate elevation. SARS-CoV-2 has a pre-existing bias for C→U mutations (Rice et al. 2020), a genomic G:C content of 38%, and a plus-strand C content of 18% (genotype Wuhan-Hu-1). These characteristics limit the extent to which molnupiravir can raise the mutation rate from its current value.
  5. Mutation and transmission limit evolution. Epidemiological spread of adaptive mutations is limited by both their generation via mutation within hosts, as well as the size of transmission bottlenecks between hosts, i.e., the number of viral genomes that establish a new infection.
  6. Individual virus genomes vary in their mutational burden. Even when the mean number of mutations per viral genome is high, the overall distribution of mutation counts can still include a class of minimally mutated viable genomes, due to either the mechanism of mutation or the existence of ‘compartments’ with low drug concentrations within the host.
  7. The mutational robustness of SARS-CoV-2 is not known. The highest mutation rate tolerated by a virus, just possibly 1-5 per genome, depends on the size of the functional genome and the expendability of ‘accessory’ genes.

Clinicians and patients must all be alerted to the critical importance of taking mutagenic antivirals as directed, and should strictly quarantine while doing so, lest transmission occur. Meanwhile, studies assessing the risk of onward transmission of mutated virus should be made a top priority for epidemiologists and evolutionary biologists.

It is easy to underestimate the potential of SARS-CoV-2 to mutate and adapt — a reality made clear by the repeated emergence of variants of concern. We therefore suggest that, until data are obtained on molnupiravir’s potential to generate viable mutated virus, measures are needed to ensure that SARS-CoV-2 is not inadvertently handed mutational resources for the accelerated generation of new variants.

Acknowledgments

The authors thank Richard J. A. Buggs for raising the main concern addressed herein to C.W.N.; Jesse D. Bloom for raising the issue of mutation- versus bottleneck-limited evolution; and Richard J. A. Buggs, Xinzhu (April) Wei, Zachary Ardern, Fabio Romerio, and Tony Goldberg for discussion and feedback on earlier versions of this manuscript. All views and any errors are our own.

Conflicts of Interest

None declared.

Submission History

  1. SUBMITTED to Nature as Correspondence on October 7, 2021; REJECTED on October 21, 2021
  2. SUBMITTED to Science as Letter on November 3, 2021; REJECTED on November 6, 2021
  3. SUBMITTED to The Lancet as Correspondence on November 9, 2021; WITHDRAWN FROM CONSIDERATION on November 29, 2021
  4. POSTED to Virological on November 29, 2021

References

Beaucourt S, Vignuzzi M. 2014. Ribavirin: a drug active against many viruses with multiple effects on virus replication and propagation. Molecular basis of ribavirin resistance. Current Opinion in Virology 8:10–15. https://doi.org/10.1016/j.coviro.2014.04.011

Drayman N, DeMarco JK, Jones KA, Azizi S-A, Froggatt HM, Tan K, Maltseva NI, Chen S, Nicolaescu V, Dvorkin S, et al. 2021. Masitinib is a broad coronavirus 3CL inhibitor that blocks replication of SARS-CoV-2. Science 373:931–936. https://doi.org/10.1126/science.abg5827

Eigen M, Schuster P. 1977. The hypercycle: a principle of natural self-organization. Part A: Emergence of the hypercycle. Naturwissenschaften 64:541–565. https://doi.org/10.1007/bf00450633

Gordon CJ, Tchesnokov EP, Schinazi RF, Götte M. 2021. Molnupiravir promotes SARS-CoV-2 mutagenesis via the RNA template. Journal of Biological Chemistry 297:100770. https://doi.org/10.1016/j.jbc.2021.100770

Kemp SA, Collier DA, Datir RP, Ferreira IATM, Gayed S, Jahun A, Hosmillo M, Rees-Spear C, Mlcochova P, Lumb IU, et al. 2021. SARS-CoV-2 evolution during treatment of chronic infection. Nature 592:277–282. https://doi.org/10.1038/s41586-021-03291-y

Loeb LA, Essigmann JM, Kazazi F, Zhang J, Rose KD, Mullins JI. 1999. Lethal mutagenesis of HIV with mutagenic nucleoside analogs. Proceedings of the National Academy of Sciences USA 96:1492–1497. https://doi.org/10.1073/pnas.96.4.1492

Lynch M, Gabriel W. 1990. Mutation load and the survival of small populations. Evolution 44:1725–1737. https://doi.org/10.1111/j.1558-5646.1990.tb05244.x

Mejer N, Fahnøe U, Galli A, Ramirez S, Weiland O, Benfield T, Bukh J. 2020. Mutations identified in the hepatitis C virus (HCV) polymerase of patients with chronic HCV treated with ribavirin cause resistance and affect viral replication fidelity. Antimicrobial Agents and Chemotherapy 64:e01417-20. https://doi.org/10.1128/aac.01417-20

Merck & Co., Inc. 2021a. Merck and Ridgeback announce submission of Emergency Use Authorization application to the U.S. FDA for molnupiravir, an investigational oral antiviral medicine, for the treatment of mild-to-moderate COVID-19 in at risk adults. https://www.merck.com/news/merck-and-ridgeback-announce-submission-of-emergency-use-authorization-application-to-the-u-s-fda-for-molnupiravir-an-investigational-oral-antiviral-medicine-for-the-treatment-of-mild-to-moderate-c/

Merck & Co., Inc. 2021b. Merck and Ridgeback Biotherapeutics provide update on results from move-out study of molnupiravir, an investigational oral antiviral medicine, in at risk adults with mild-to-moderate COVID-19. https://www.merck.com/news/merck-and-ridgeback-biotherapeutics-provide-update-on-results-from-move-out-study-of-molnupiravir-an-investigational-oral-antiviral-medicine-in-at-risk-adults-with-mild-to-moderate-covid-19/

Otto SP, Day T, Arino J, Colijn C, Dushoff J, Li M, Mechai S, Van Domselaar G, Wu J, Earn DJD, et al. 2021. The origins and potential future of SARS-CoV-2 variants of concern in the evolving COVID-19 pandemic. Current Biology 31:R918–R929. https://doi.org/10.1016/j.cub.2021.06.049

Painter WP, Holman W, Bush JA, Almazedi F, Malik H, Eraut NCJE, Morin MJ, Szewczyk LJ, Painter GR. 2021. Human safety, tolerability, and pharmacokinetics of molnupiravir, a novel broad-spectrum oral antiviral agent with activity against SARS-CoV-2. Antimicrobial Agents and Chemotherapy 65:e02428-20. https://doi.org/10.1128/aac.02428-20

Pfizer Inc. 2021. Pfizer’s novel COVID-19 oral antiviral treatment candidate reduced risk of hospitalization or death by 89% in interim analysis of phase 2/3 EPIC-HR study. https://www.pfizer.com/news/press-release/press-release-detail/pfizers-novel-covid-19-oral-antiviral-treatment-candidate

Pillai SK, Wong JK, Barbour JD. 2008. Turning up the volume on mutational pressure: Is more of a good thing always better? (A case study of HIV-1 Vif and APOBEC3). Retrovirology 5:26. https://doi.org/10.1186/1742-4690-5-26

Rice AM, Morales AC, Ho AT, Mordstein C, Mühlhausen S, Watson S, Cano L, Young B, Kudla G, Hurst LD. 2020. Evidence for strong mutation bias toward, and selection against, U content in SARS-CoV-2: implications for vaccine design. Molecular Biology and Evolution 38:67–83. https://doi.org/10.1093/molbev/msaa188

Sadler HA, Stenglein MD, Harris RS, Mansky LM. 2010. APOBEC3G contributes to HIV-1 variation through sublethal mutagenesis. Journal of Virology 84:7396–7404. https://doi.org/10.1128/jvi.00056-10