17 Aug Episode 282 Update on Ensitrelvir

Wed, 08/17/2022 - 17:23

ENSITRELVIR (S-217622) a Noncovalent Oral SARS-CoV‑2 3CL Protease Inhibitor Candidate to Treat COVID-19


Dear colleagues,

In Episode 266, I already introduced this novel inhibitor of the main M or 3CL protease of SARS-CoV-2.  There are more data now both pre-clinically and in phase 2.  All in all, Ensitrelvir has a favorable profile:

  • Orally available sub-micromolar activity in cell culture with long half-life time and no activity against human proteases (Ep 282-1)
  • Proven anti-viral activity in mouse, even after 24 h delayed start (Ep 282-2).  In hamster models also anti-disease and anti-transmission activity (Ep 282-3)
  • Equally active against variants, including BA.2 and more active on weight basis than Molnupiravir and Nirmatrelvir (Ep 282-4). 
  • After short course within 5 days of PCR positivity in human phase 2, significant earlier negativation of PCR.  Decrease of viral load (including omicron) is at least as good or superior to the polymerase inhibitors Molnupiravir and Remdesivir as well as the “established” protease inhibitor , Paxloxid (= Nirmatrelvir + Ritonavir) (Ep 282-5 and -6)
  • Clinical benefit not yet very clear; acceptable side effects (Ep 282-5 and -6)
  • Phase 3 ongoing (Ep 282-7)
  • It is a single compound: no need for inhibitor of liver metabolism, such as Ritonavir (which is part of Paxlovid), thus avoiding many potential drug-drug interactions


Although formal studies on the resistance profile of Ensitrelvir have not yet been published, it seems that low level resistance is present in some circulating variants (Ep 282-8).  Moreover, after in vitro induction of resistance to Nirmatrelvir, some of the mutants show cross-resistance to Ensitrelvir (Ep 282-9).  

In the context of resistance induction, the long half-life could be a disadvantage, especially if given repeatedly to immune suppressed subjects with chronic or recurrent infection.     





Ep 282-1: Yuto Unoh Development of S217622 J Med Chem May 2022 vol 65 p. 6499





  1. Optimization of antiviral activity, oral bio-availability and pharmacokinetics of original hit




Compound 3 (S-217622 or Ensitrelvir) has 13 nanomolar activity against the 3CL SARS-CoV-2 protease and very high oral bioavailability


  1. Broad spectrum sub-micromolar activity of the optimized S-217623 compounds against human beta-CoV




  1. No activity against human proteases




  1. In vivo antiviral activity and pharmacokinetics profile





S-217622 is the first nonpeptidic, noncovalent, oral 3CLpro inhibitor clinical candidate for treating COVID-19.


It has a favorable preclinical profile as a once-daily oral therapeutic agent for COVID-19 with

  • promising antiviral activities to known variants of concern,
  • a long elimination half-life in vivo, especially in monkeys and dogs,
  • excellent oral bioavailability,
  • strong efficacy in an in vivo mouse model infected with SARS-CoV-2.



Ep 282-2:  Haruaki Nobori Efficacy of Ensitrelvir against SARS-CoV-2 in a 24 Hrs delayed-treatment

mouse model  JAC August 2022








Ensitrelvir has the potential to suppress viral replication and clinical disease caused by SARS-CoV-2 infection, when started 1 day after infection



Ep 282-3: Sasaki bioRxiv 15 Feb 2022: extensive studies in hamsters showing clear-cut antiviral effect of S-217622, but also mitigation of body weight loss and lung pathology of infected animals, treated from the day of infection or from the next day. (MPV = Molnupiravir 200 mg/kg)



Reducing transmission: only the contact hamsters were prophylactically treated with S-217622 or MPV (b.i.d.) n, o, Viral RNA levels (n) and virus titers (o) in lungs from the contact hamsters after 6 days of co-housing with the infected hamster






Ep 282-4: Uraki Antiviral susceptibility of omicron BA.2 Nature 7 July 2022




In hamsters Ensitrelvir at lower concentration more active than Molnupiravir or Nirmatrelvir against omicron BA.2








Ep 282-5: Mukae Phase 2a study in  medRxiv 17 May 2022


Set-Up: Three small groups of patients with mild-moderate COVID, treated for 5 days: either 125, 250 or 0 mg of Ensitrelvir per day. 


Patients included within 120 hours after positive PCR

Of 47 patients:

  • 40 had mild-moderate disease and 7 were asymptomatic;
  • 42 delta infected and 5 with omicron
  • 38 twice vaccinated


Results: Ensitrelvir faster and more pronounced decrease in viral titer (TCID50) and viral load (RNA) than placebo






But no significant difference in symptom score

Interpretation: favorable profile according to authors for 2 reasons:

  • Principal: orally available non-peptidic, non-covalent cysteine protease inhibitor, unlike the peptide-like nirmatrelvir : challenges remain for improving the target selectivity and PK profiles of peptide-like covalent inhibitors owing to the intrinsic nature of the reactivity, low membrane permeability and low metabolic stability.
  • Activity: Viral RNA reduction superior with Ensitrelvir =: -1.4 log10  on day 5, while Nirmatrelvir/Ritonavir = only – 0.868 log10 on day 5 (and Molnupiravir only  -0.547 log10)



Ep 282-6: Mukae Phase 2b study medRxiv 26 June 2022



Among the 428 patients in total (recruited within 120 hours after PCR+), 142, 143, and 143 were randomized to receive Ensitrelvir 125 mg (375 mg day 1), Ensitrelvir 250 mg (750 mg day 1), or placebo, respectively, within 120 hours after until day 5.  All patients had at least 1 symptom, with mild to moderate score;

Of those 341 patients (Ensitrelvir 125 mg: 114 pt; Ensitrelvir 250 mg: 116 pt; and 108 placebo: 111 pt) were included in the intention-to-treat (ITT) population

There were no differences in baseline characteristics (See Table 1) 

  • Mean age was around 36 in all groups, with very few > 65.
  • Nearly half of the patients were randomized within 72 hours of COVID-19 symptom onset and >80% had received at least 1 dose of mRNA SARS-CoV-2 vaccine




The change from baseline in viral titer (log10 TCID50/mL) on day 4, was significantly greater with Ensitrelvir 125 mg and 250 mg versus placebo (a = absolute; b = change from baseline This was irrespective of vaccination status and treatment initiation > or < 76 hours.  Differences were small, however.




The time of viral clearance was clearly shorter with Ensitrelvir




There was no very clear difference in symptoms, although respiratory symptoms and feverishness tended to improve faster with Ensitrelvir, but digestive or systemic symptoms not so.


Treatment-related adverse events (AEs) were slightly higher in Ensitrelvir groups (13.6 and 22.1 %) versus placebo (5 %):  HDL cholesterol showed a very temporary “dip”






  1. Improvement in viral load and symptoms seemed more pronounced in phase 2a than in phase 2b.  Possible explanation:  in 2a Delta was dominant, while in 2b the “milder” omicron was dominant e.g. the baseline viral load was higher in phase 2a than 2b?
  2. Nevertheless, still greater reduction in viral loads between baseline and day 4
    • Ensitrelvir  -1.25 log10
    • Molnupiravir  -0.55
    • Paxlovid  -0.87
    • Regeneron mAb cocktail -086
  3. No viral rebound with Ensitrelvir, while it has been described for Paxlovid (https://www.fda.gov/media/155194/download): because of longer half-life of Ensitrelvir (51 hrs) versus Paxlovid (6.5 hrs).
  4. No clear difference between 125 and 250 mg dose


Conclusion: Five-day, once daily, oral Ensitrelvir treatment demonstrated rapid and favorable antiviral efficacy with an acceptable safety profile in patients with mild-to-moderate COVID-19, a majority of whom had been vaccinated.  The results support further clinical development of Ensitrelvir for mild-to moderate COVID-19, but more attention should be given to 65+ high risk population.



Ep 282-7: Hiroshi Yotsuyanagi Protocol for phase 3 medRxiv 17 July 2022


Dosing and study population rather similar to phase 2b, but

  • Also subjects 65-70 yrs included, but not older.  
  • Should present at least 1 “moderate symptom”.
  • Really severe symptoms excluded.
  • Both 125 mg and 250 mg doses included (although no difference between both dosages in phase 2)






Ep 282-8: Moghadasi  Circulating SARS-CoV-2 variants with protease inhibitor resistance mutations bioRxiv 8 August 2022




T45I, D48Y and M49I selective resistance to Ensitrelvir

S144A and A173V/T selective resistance to Nirmatrelvir

Deletion of P168: low-level cross-resistance to Ensitrelvir and Nirmatrelvir

Combination of ∆P168 + A173V: high level resistance to Nirmatrelvir, but low level to Ensitrelvir



The mutations ∆P168,  A173V and M49I are present in circulating SARS-CoV-2 viruses, including BA.5



Ep 282-9:  Iketami Multiple pathways for SARS-CoV-2 resistance to Nirmatrelvir bioRxiv 8 August 2022


In vitro dose-escalating resistance induction with Nirmatrelvir shows multiple complex pathways.


  1. Single mutations: S144A, E166V, and T21I + S144A showed substantial (~7 to 25-156 fold) cross-resistance to Ensitrelvir




  1. Combination of mutations: L50F + F140L + L167F + T304I showed cross-resistance to Ensitrelvir (~25-fold)




  • Some circulating SARS-CoV-2 variants (including BA.5) have low level resistance to Ensitrelvir or Nirmatrelvir. Delta P168 confers low level cross-resistance to both protease inhibitors
  • After serial in vitro passage with Nirmatrelvir, some of the induced single mutations and combinations of mutations show rather high cross-resistance to Ensitrelvir.





  1. Uno S‑217622, a Noncovalent Oral SARS-CoV‑2 3CL Protease Inhibitor Clinical Candidate for Treating COVID-19   https://pubs.acs.org/doi/pdf/10.1021/acs.jmedchem.2c00117
  2. Haruaki Nobori Efficacy of ensitrelvir against SARS-CoV-2 in a 24 Hrs delayed-treatment mouse model  JAC August 2022: https://academic.oup.com/jac/advance-article/doi/10.1093/jac/dkac257/6652992
  3. Sasaki Oral administration of S-217622, a SARS-CoV-2 main protease inhibitor, decreases viral load and accelerates recovery from clinical aspects of COVID-19 in hamsters bioRxiv 15 Feb 2022 https://www.biorxiv.org/content/10.1101/2022.02.14.480338v1
  4. Uraki Characterization and antiviral susceptibility of SARS-CoV-2 Omicron BA.2 Nature 7 July 2022 https://www.nature.com/articles/s41586-022-04856-1
  5. Mukae Phase 2a study medRxiv 17 May 2022 https://www.medrxiv.org/content/10.1101/2022.05.17.22275027v1
  6. Mukae Phase 2b study 26 June 2022 https://www.medrxiv.org/content/10.1101/2022.06.22.22276792v1
  7. Hiroshi Yotsuyanagi Protocol for phase 3 study 17 July 2022 https://www.medrxiv.org/content/10.1101/2022.07.15.22277670v1
  8. Moghadasi Transmissible SARS-CoV-2 variants with resistance to clinical protease inhibitors bioRxiv 8 August https://www.biorxiv.org/content/10.1101/2022.08.07.503099v1
  9. Sho Iketani Multiple pathways for SARS-CoV-2 resistance to nirmatrelvir bioRxiv 8 August 2022 https://www.biorxiv.org/content/10.1101/2022.08.07.499047v1