Increased resistance to human interferons could be a potential driver of SARS-CoV-2 evolution

In a recent study published in PNAS, researchers compared the potency of 17 human interferons (IFNs) with multiple variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Recent SARS-CoV-2 variants of concern (VOCs) showed increased IFN resistance, indicating that evasion of innate immunity or human IFNs may be a driving force for SARS-CoV-2 evolution.

Study: Interferon resistance of emerging SARS-CoV-2 variants.  Image Credit: WhiteDragon/Shutterstock
Study: Interferon resistance of emerging SARS-CoV-2 variants. Image Credit: WhiteDragon/Shutterstock


Previous studies have linked the emergence of SARS-CoV-2 variants with spike (S) mutations. However, studies have not established the role of other viral protein mutations that can inhibit the human IFNs, e.g. IFNα2 and IFNβ, which are being repurposed for the treatment of coronavirus disease 2019 (COVID-19).

About the study

In the present study, researchers obtained representative SARS-CoV-2 isolates of the B, B.1, B.1.1.7, and B.1.351 lines from They incubated human alveolar type II epithelial cell line (A549)-angiotensin-converting enzyme 2 (ACE2) cells with 17 recombinant IFNs overnight in parallel and in triplicate. They infected these cells with a non-saturating dose of virus for 2 hours.

They used a quantitative polymerase chain reaction (qPCR) test to determine the amounts of virus produced 24 hours after infection. The investigators also evaluated the qPCR assay against a VeroE6 plaque assay using three-fold serial dilutions of SARS-CoV-2 isolate B.1.351, detecting one plaque-forming unit by this assay, corresponding to ∼900 SARS-CoV -2 nucleocapsid 1 (N1) copies.

Furthermore, the team used immunofluorescence to quantify viral copy numbers in primary airway epithelial cells infected with different SARS-CoV-2 variants. Finally, they generated a heat map to visualize the antiviral potency of all 17 IFNs against the five SARS-CoV-2 isolates.

Study findings

Although the IFNβ and IFNλ1 doses did not substantially affect cell viability, a 2-picomolar (pM) concentration maximally distinguished the antiviral activities of IFNβ and IFNλ1, two widely divergent IFNs. Although the authors observed a strong correlation between the virus titers obtained with both assays, compared to the VeroE6 plaque assay, the qPCR assay strongly distinguished between the antiviral activity of the different IFNs.

All 17 IFNs exhibited antiviral activities against SARS-CoV-2, with an observed correlation between IFN potencies against all five isolates. The three IFNλ subtypes showed no to less than two-fold antiviral activities compared to most IFN-Is. IFNα8, IFNβ and IFNω were the most potent, the type III (λ) IFNs were the least potent, and IFNα5, IFNα17 and IFNα14 showed moderate potency.

The heatmap data confirmed that the IFN resistance of emerging SARS-CoV-2 variants was especially striking compared to the ancestral USA-WA1/2020 strain, showing 25 to 322-fold higher IFN-I resistance . However, the research experiments did not reveal how different IFN-I doses affected the replication of all SARS-CoV-2 strains. It also remained unclear whether the emerging variants were resistant to IFN-IIIs. Inhibition curves for five IFNs showed that IFNλ1 had a 100-fold higher half-maximal inhibitory concentration (IC50) than IFNα2, showing a 14-fold higher IC50 than IFNβ, IFNα5 and IFNα8. Both IFNβ and IFNλ1 similarly inhibited lineage A and B isolates.

Interestingly, both IFNβ and IFNλ1 failed to achieve maximal inhibition against the alpha-VOC, saturating at 15- to 20-fold higher levels than the line B isolate. Notably, Omicron had the highest levels of residual virus replication at the highest doses of one of the most potent IFNs, IFNβ.


The study demonstrated antiviral potency for the various human IFNs in a specific sequence, indicating how SARS-CoV-2 evolved after switching hosts to become more resistant to human IFNs. The study results may help identify IFN subtypes that may be most helpful in treating early SARS-CoV-2 infections. For example, SARS-CoV-2 required higher doses of IFNλ to achieve a comparable antiviral effect in vivo as the IFN-Is and IFNβ showed potential as a therapeutic agent against COVID-19.

Among the IFNα subtypes, IFNα8 exhibited anti-SARS-CoV-2 potency comparable to that of IFNβ. It also showed high antiviral activity against human immunodeficiency virus 1 (HIV-1), providing a potential treatment approach for both viruses. While IFNα6 restricted HIV-1, IFNα subtypes were the weakest against SARS-CoV-2. Conversely, IFNα5 strongly inhibited SARS-CoV-2 but weakly inhibited HIV-1. The genes limiting IFN responses in emerging SARS-CoV-2 variants suggested that IFNs may also influence the evolution of SARS-CoV-2. Similarly, previous studies with HIV-1 suggested that IFNs may determine the evolution of pandemic viruses.

Overall, the present study reinforced the importance of continued genomic surveillance of SARS-CoV-2 to assess emerging variants that evade host IFN responses. Indeed, the role of the innate immune response in driving SARS-CoV-2 evolution could have implications for IFN-based therapies.

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