Antiviral activity of mycophenolic acid against influenza viruses and MERS coronavirus

Abstract

Influenza virusand Middle East Respiratory Syndrome Coronavirus(MERS-CoV) cause life-threatening respiratory disease. There are 3 to 5million severe cases and 250,000 to 500,000 fatal cases caused by seasonal influenza virus A(H1N1)virus, A(H3N2) virus and influenza B virus every year. Pandemic influenza, which is associated with higher mortality, has once every few decades. Among various influenza viruses, the avian-origin A(H5N1)virus and A(H7N9) virus are the most virulent in humans. MERS-CoV was first reported to cause severe human infections in2012, and now there are over 800 cases worldwide with a case-fatality rate of about 30%.

Antiviral is one of the armamentaria in the treatment of influenza virus and MERS-CoV infection. For influenza viruses, the neuraminidase inhibitors oseltamivir and zanamivir have been used in the treatment of influenza A and B infection. However, the clinical efficacy has been questioned, and delayed administration of NAIs has been associated with poor outcome. Favipiravir, a viral RNA polymerase inhibitor, has been approved only in Japan. Novelantivirals are still under development and unlikely to be available in the near future. No antivirals against MERS-CoV have been approved for clinical use yet.

Since effective antivirals against influenza virus and MERS-CoV are urgently needed, we investigated whether an approved drug, mycophenolic acid (MPA),can have antiviral activity. In this thesis, we investigated the antiviral effect of MPA against the A(H1N1)pdm09 virus, influenza B virus, A(H7N9) virus, and MERS-CoV. MPA, which is available for clinical use as mycophenolate mofetil (MMF)or mycophenolate sodium, has been used as immunosuppressant for transplant recipients and in the treatment of autoimmune diseases.MPA suppresses the synthesis of guanosine in the de novo pathway. MPA has been reported to have antimicrobial activities against several viruses and fungi.

Time-of-addition experiment showed that earlier addition of MPA led to better cell protectionon influenza infection. The EC50of MPA added 2 hours before virus inoculation was 1.510 µM. The EC50of MPA was >12.5 µM when MPA was added 7 hours after virus inoculation. Cell protection assay showed that MPA protected cells infected with influenza viruses(EC50, 0.208-1.510µM)or MERS-CoV(EC50, 0.531 µM). Virus yield reduction assay showed that the growth of influenza viruses and MERS-CoV were reduced in dose-dependent manner in the presence of MPA. Plaque reduction assay also showed that MPA inhibited the growth of influenza virus(EC50, 0.528-0.872 µM) and MERS-CoV (EC50, 0.395µM). Among influenza viruses, the activity of MPA against influenza B virus was most potent. Assay assessing nucleoprotein expression revealed that nucleoprotein expression in influenza A virus was completely inhibited in the presence of 12.5 µM of MPA. Addition of guanosine, but not adenosine, revertedthe antiviral activity of MPA on influenza infection completely, suggesting that the antiviral activity is dependent on the depletion of guanosine. In conclusion, our study showed that MPA was active against influenza virus and MERS-CoV. Further studies are necessary to investigate the use of MPA for influenza infection or MERS in clinical practice.