Remdesivir has been approved as a stand-alone therapy for the treatment and prevention of COVID-19. One of the issues that arises with any single drug treatment of an RNA virus is the level of resistance. Resistance is documented for remdesivir in vitro, however, to date, no examples of resistance to remdesivir have been documented. Here we describe research by a team at Yale University, which documents SARS-CoV-2 resistance to remdesivir in a patient with an abnormally prolonged infection.
Gandhi et al. treated a 70-year-old, immunocompromised patient who developed a prolonged infection with SARS-CoV-2. Figure 1A shows the patient’s time course and symptoms. During the initial period, she suffered from persistent fever, anosmia with intermittent cough and rhinorrhea. The patient’s viral load was monitored throughout her infection, a peak was detected on day 148. At this point, it was decided to treat the patient intravenously with a ten-day course of remdesivir. Treatment resulted in cessation of the patient’s fever, normal CRP, and improvement of opacities on her chest computed tomography (CT) scan. However, by day 160, there was a steady increase in the amount of virus produced, as judged by PCR. The patient was treated with antibodies on day 163 for one week, specifically casirivimab-indevimab, until symptoms and tests returned to normal. After antibody treatment, his recovery was carefully monitored for the rest of his stay.
Isolation of virus-resistant remdesivir
Although the antiviral drug alleviated symptoms, there was a recurrence of viral shedding, suggesting that resistance had occurred. To confirm this suspicion, the virus was isolated from nasopharyngeal tissue, saliva, stool and blood samples of infected patients to clone the viral genome. When sequencing the virus, Gandhi et al. noted the appearance of a mutation at E802D seven days after the initial dose of remdesivir. The E802A mutation had previously been associated with resistance to remdesivir in vitro. In addition to E802D, mutations A504V in nsp14 (exonuclease) and I115L in nsp14 (endoRNAse) were also found in the isolated virus.
Remdesivir resistance decreases viral form
To determine whether E802D indeed conferred resistance, the mutation was cloned into an isogenic strain of SARS-CoV-2 and resistance to remdesivir was measured. Figure 2 shows that both E802D/A mutations are resistant to remdesivir. E802D causes an overall 4.2-fold increase in drug resistance while E802A has a 2.7-fold resistance. Like Gandhi et al. noted, the fitness disadvantage is only noticed in the absence of remdesivir. In the presence of remdesivir, the E802D variant shows a marked growth advantage. The implication is that this particular mutation is unlikely to spread through the population.
Figure 3 illustrates the three-dimensional diagram of the location of the E802D mutation within the RNA-dependent RNA polymerase nsp12 as determined by cryo-electron microscopy. The E802D mutation occurs in the active site. The substitution at E802D is a substitution from aspartic acid to glutamic acid in the active site of polymerase nsp12. Note that the two amino acids that lead to a mutation, alanine and aspartic acid, are shorter than glutamic acid. The mechanism by which this leads to resistance to remdesivir is not immediately obvious. The authors believe that it can either allow elongation at the remdesivir insertion site or exclude remdesivir from binding. Of the two, it seems more likely that a more open configuration would allow elongation of the growing chain beyond the remdesivir insertion.
As mentioned, other mutations associated with RDV therapy were found: A504V in nsp14 (exonuclease) and I115L in nsp14 (endoRNAse) as well as four other lower grade mutations. Both have demonstrated an increase in allele frequencies from remdesivir treatment, but still need to be tested further. This study notes the inactive nature of the virus under treatment with remdesivir despite the increase in viral shedding in immunocompromised patients. As we have known for some time with viruses, single drug treatment is not optimal because very often mutations occur, remdesivir being no exception. As is often the case, the resistance mutation incurs a fitness cost, the implication of which is that the mutation is unlikely to spread through the population. For now, that means monitoring the effects of Remdesivir as an antiviral drug for SARS-CoV-2 and considering multi-antibody therapies for immunocompromised patients.
A take-home message from this study is that using single drugs to treat RNA viruses often leads to resistance. The future utility of remdesivir as a first-line treatment for SARS-CoV-2 should include combination with another active antiviral drug, as well as attempts to ensure that the drug can be administered intramuscularly or subcutaneously. skin.