In a recent study posted to the Research Square* preprint server, researchers demonstrated that impaired type I interferon (IFN-I) immunity is associated with increasing coronavirus disease 2019 (COVID-19) severity.
The response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is highly variable, resulting in diverse clinical outcomes, ranging from asymptomatic infection to severe disease and even death. A better understanding of such heterogeneity in responses to IFN-based COVID-19 treatment could help identify alternative therapeutic strategies.
Studies have shown that while blood type I interferon (IFN-I) responses are critical in early acute infection, they get impaired preceding clinical worsening of COVID-19. Hence, researchers have observed a dichotomous clinical response to IFN-I-based treatments in late-stage COVID-19.
Consistent with these observations, studies have also identified mutations in genetic loci that govern toll-like receptor (TLR)3- and interferon regulatory factor (IRF)7-dependent IFN-I immunity. Likewise, autoantibodies against IFNα, IFNω, or IFNβ, and TLR7 pathway are critical in conferring early immunity to SARS-CoV-2 infection. Yet, exogenous IFN-I treatment does not improve clinical outcomes for COVID-19.
Here, it is also important to note the importance of deploying sensitive immunoassays when studying type I IFN directly from patient samples. As is known, IFNα protein levels remain below the detection limits of conventional enzyme-linked immunosorbent assay (ELISA) and Luminex assays. More importantly, plasma IFN levels at the time of patient sampling do not depict how patients’ cells respond to viral exposure.
About the study
In the current study, researchers enrolled healthy controls and SARS-CoV-2 infected participants at St. James’s Hospital in Dublin, Ireland, between March and June 2020. The control group comprised 61 healthy individuals who matched the study cohort in age, ±5 years.
At the time of enrollment, the team assigned all the study participants with disease severity grades based on hospital admission and supplemental oxygen requirements. The patients who did not require hospitalization were classified as moderately ill, while those hospitalized and given supplemental oxygen via nasal cannula were considered severe. Those classified as critical required more than six liters of oxygen per minute, delivered via high-flow nasal oxygen or a venturi mask.
The team characterized all 13 IFN-alpha subtypes, especially IFNα2 proteins using highly sensitive assays, IFN function, interferon-stimulated genes (ISGs), and autoantibodies neutralizing IFN-I. The researchers also selected a patient subset and stimulated their whole blood samples with relevant viral antagonists to better assess the functionality of their immune system to external stimuli. Likewise, they obtained plasma samples from 342 and 212 for cytokine and autoantibody analysis, respectively. Further, they used plasma samples from a small subset of 31 patients for cellular phenotyping.
The authors observed multiple disturbances in the IFN-I response with increasing COVID-19 severity. Late stimulation with exogenous IFN-I did not benefit patients; moreover, it compromised any apparent clinical benefit of viral agonists. Notably, IFNβ secretion was more disturbed than IFNα, which was unanticipated given its low levels in patient plasma. This finding further indicated the challenges in accurately detecting and quantifying IFNβ in the blood.
Gyros assay identified four male patients as anti-IFNα autoantibody-positive in the hospitalized group, with two moderately, one severely, and one critically ill. Since the majority of patients remained negative for anti-IFN autoantibodies, this factor could not have driven severe disease in patients.
The authors also observed that stimulating the whole blood from hospitalized COVID-19 patients with ex vivo IFN-I induced a non-canonical inflammatory response. Further, TLR gene expression data from whole blood revealed similar TLR7 expression in all groups, lower TLR3 expression, and higher TLR4 and TLR8 expression in hospitalized patients. In addition, the authors found eight additional cytokines with significant differences in hospitalized patients, primarily after TLR3 stimulation.
The IFN-I response to stimulations, including polyinosinic: polycytidylic acid (Poly: IC), lipopolysaccharide (LPS), and R848 (a TLR7/8 agonist) was disturbed in critically ill patients. In fact, Poly: IC stimulation significantly reduced the IFNβ response in moderate and severe disease groups but reduced after R848- and LPS-stimulation in the hospitalized groups.
The blunting of broad IFN responses in severe COVID-19 patients remains unexplained. Studies showed that this phenotype was not attributable to SARS-CoV-2 interference with the host’s immunity. Consequently, with increasing severity, pDCs remained capable of producing IFNα intracellularly. Future works should investigate the functionality of intracellular pathways in cells isolated from severe COVID-19 patients. It could help better understand this phenotype and provide targets for new treatment strategies.
The study highlighted the critical role IFN-I mediated immunity plays in dictating COVID-19 outcomes. Other studies have implicated IFNβ in long COVID-19, thus further emphasizing the need to understand the regulation of IFN-I during SARS-CoV-2 infection.
The current study also identified two main reasons for an impaired IFN-I response in severe COVID-19. They showed decreased circulating plasmacytoid dendritic cells (pDC) and dysregulated monocytes which secreted fewer IFNα. Second, IFN-I stimulation of leukocytes promoted an inflammatory response in severe COVID-19 patients but not in moderately ill patients.
Furthermore, physiological concentrations of IFNα proteins are often below pg/mL levels. The study results, therefore, highlighted the significance of using sensitive assays to measure circulating IFNα levels in severe COVID-19 patients. The authors also observed differences in IFNα2 protein plasma levels between moderately and severely ill COVID-19 patients.
As new experimental tools become available, they will help the researchers fully understand the functions of all 13 IFNα subtypes in COVID-19 patients, which might also be relevant for other viral infections. Most importantly, clinical studies should test IFN-I intervention strategies early, i.e., before the disease symptoms appear, and target them toward patients with known risk factors.
Research Square publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.