Ongoing identification of cases of nCoV3,22 suggests continuing introduction of the virus to humans in the Middle East from an unknown source. Given the genetic relationship of hCoV-EMC/2012 to other bat coronaviruses5, one can speculate that bats may be the reservoir of this virus; however, additional host species should be considered. With documented human-to-human transmission in close contact situations and the first documented mild case22, there is a real concern that we could be observing the ‘tip of the iceberg’ and perhaps the start of an epidemic. Regardless, with a 65% case-fatality rate despite intensive medical intervention, therapeutic strategies are urgently needed. Despite the significant increase in research on coronaviruses since the discovery of SARS-CoV in 2003, there is no definitive antiviral or therapeutic treatment for coronavirus infections in humans. Pegylated interferon-α was shown to be an effective prophylactic treatment against infection with SARS-CoV in cynomolgus macaques, but was less effective when administered post exposure23. No other therapeutics have been tested for antiviral activity against SARS-CoV in a higher order animal model. In the SARS-CoV mouse model, poly IC:LC24 and mDEF201 (an adenovirus expressing mouse IFN-α)25 can protect mice from lethal disease; however, neither of these approaches yields an immediate therapeutic for use in humans. Poly IC:LC has been tested in numerous clinical trials, but is not currently approved for treatment of any human disease. Adenovirus-based therapy has multiple complicating factors, such as pre-existing immunity, that have not been adequately addressed, nor is it approved for use in humans26.

Here we identified a potential therapeutic approach against hCoV-EMC/2012 combining IFN-α2b and ribavirin. Either treatment alone reduced virus replication by at least 1-log or as much as 4-logs in susceptible cell lines. Moreover, when combined, efficacy was reached at lower concentrations. Thus, this combination may provide a benefit as a treatment in humans. Vero cells display a high level of resistance to the activity of ribavirin19,27. Thus, we also performed the same assay in LLC-MK2 cells, where sensitivity to ribavirin was observed at a much lower concentration.

Previous in vitro studies have demonstrated that SARS-CoV is sensitive to ribavirin28 and to various classes of interferon (α, β and γ)27,29,30,31,32,33,34. The sensitivity of SARS-CoV to ribavirin appears to be cell line dependent, with concentrations as low as 50 μg/ml ribavirin being reported as effective16. Unfortunately, this concentration is higher than the peak serum concentration reached in humans of approximately 24 μg/ml35. IFN-α2b was previously reported to inhibit growth of SARS-CoV starting at 1000 U/ml with a 1-log reduction at 2000 U/ml34. Following infection, only IFN-β (EC 50 560 IU/ml) has shown a dose dependent antiviral effect36. In this study we report a nearly 4-log reduction in virus titers for hCoV-EMC/2012 at comparable doses.

During the outbreak of SARS-CoV, different combinations of therapeutic interventions were attempted; however, none were implemented in a manner that allowed a critical assessment of their effectiveness. The most frequently administered therapeutics were broad-spectrum antibiotics, glucocorticoids and ribavirin37,38,39. The lack of a standard dosing regimen for ribavirin makes comparisons difficult18; however, low dose ribavirin (400–600 mg/day) therapy was shown to be ineffective likely due to an insufficient plasma concentration40. In contrast, when used at higher doses other studies have found that ribavirin alone reduced viral loads in over half of the patients and when combined with the viral protease inhibitors lopinavir/ritonavir, patients had a lower incidence of adverse outcomes16. Despite being used in a large number of patients, it still remains unclear whether ribavirin alone was effective against SARS-CoV41. Alfacon-1, a synthetic IFN-α, has also been suggested to be beneficial to patients17. Unfortunately, all of these studies suffer from the confounding use of corticosteroids in doses that vary among studies making a definitive treatment elusive. It has been suggested that combination of interferon and ribavirin treatment should be evaluated18. While ribavirin can result in reversible hemolytic anemia, this complication typically occurs following longer treatment protocols35,42. This suggests that short-course ribavirin therapy for an acute infection such as nCoV may not be a significant complication as mild anemia was the most frequently reported side effect during ribavirin treatment for Lassa virus infection43.

A synergistic effect of IFN-α and ribavirin has been previously reported in vitro for both SARS-CoV27,44 and feline infectious peritonitis virus45; however, we observed an additive effect against hCoV-EMC/2012 in this study. The levels of IFNα-2b and ribavirin required for inhibition of nCoV replication must be achievable in humans in order to be relevant for clinical use. In humans, an interferon concentration of 100-750 IU/ml has been observed after intravenous injection of up to 3 × 107 U46,47, while 24 μg/ml of ribavirin is achievable following a 1000 mg intravenous dose35. Here IFN-α2b and ribavirin alone were shown to have an antiviral effect against hCoV-EMC/2012; however, in Vero cells the concentrations required to achieve a beneficial effect are likely higher than what is achievable in humans. When combined, the inhibitory concentration of both IFNα-2b and ribavirin drops to ranges that are likely achievable in humans, suggesting that the combination is a potential treatment option. Used early in the course of infection or given prophylactically to close contacts of sick individuals (close contact transmission has been documented in infection chains) this combination may improve clinical outcomes. In addition, reduced viral load would also likely translate to reduced virus shedding; thus, reducing the risk of secondary transmission. As these two drugs are currently used together in the clinic, combination therapy including IFN-α2b and ribavirin should be considered for case patient management of new nCoV cases and possibly for prophylaxis in highly exposed individuals.