Since the start of the SARS-CoV-2 pandemic back in early 2020, immunologists across the globe have been striving to identify the underlying cause of severe Covid-19. How is it possible that while some patients are asymptomatic, others are incredibly sick and even die as a result of the infection?

We all now know that there are several risk factors associated with poor patient outcome from Covid-19 including being male, being over 65, and having an underlying medical condition. In addition, the CDC has published a comprehensive list of high-risk groups for severe Covid-19 symptoms. 

https://www.cdc.gov/coronavirus/2019-ncov/need-extra-precautions/people-with-medical-conditions.html

But we have yet to understand the foundation of these risk factors, and how best to mitigate them in a clinical setting.

The Correlation Between Covid-19 infection and Autoimmune Disease

Over the past 10 months, epidemiology studies have identified ‘at risk’ groups within the population, and there appears to be a significant number of autoimmune conditions associated with severe Covid-19 symptoms. Whereas this increased risk of Covid-19 severity might be apparent in some conditions such as autoimmune myocarditis and Lupus, others such as the link to rheumatoid arthritis are less obvious. Remarkably, recent studies appear to indicate that SARS-CoV-2 infection may actually trigger autoimmune diseases in some people. This is not an isolated phenomenon, and in 2016, J. R. Kerr demonstrated that parvovirus B19 infection had the potential to give rise to the development of a spectrum of autoimmune diseases. All of this highlights that we have a long way to go before we can begin to understand why this virus is so dangerous for a sector of the population. 

One question now being raised by many in the medical field is ‘Could it be that the source of disease severity might not actually be the SARS-CoV-2 virus itself, but rather the unrestrained response of the immune system to it?’  The proposed theory is that hyper-response to SARS-CoV-2 infection may be primed by underlying dysfunction in the immune system associated with autoimmune disease, resulting in a misguided response to the virus. This hypothesis appears to be supported by recent multiparameter flow cytometry studies that describe the protracted impact of SARS-CoV-2 on both the adaptive and innate immune systems (Kratzer, et. al. 2020). It was reported that even 10 weeks after acute infection, convalescent Covid-19 patients display prolonged activation of CD3, CD4 and CD8 T-cells, coupled with increases in CD4 and CD3 effector memory cells. However, these same patients also display markedly reduced neutrophils and Tregs. The underlying mechanism for this neutrophil reduction from Covid-19 is not understood but may be either exhaustion of the neutrophil pool or the possible production of complement fixing anti-neutrophil antibodies Nevertheless, the loss of Tregs and neutrophils is a dangerous combination of immunological changes that have the potential to trigger autoimmune diseases (Kratzer, et. al. 2020). This study highlights the need to help patients manage long term issues of Covid-19, by providing immune monitoring and access to immunomodulatory treatments to help establish the re-balance of neutrophils and T-cells in these individuals.

Profile of Autoimmune Responses to SARS-CoV-2

One key indicator of a misguided immune response to the virus has been brought to light by the finding that critically ill patients with Covid-19 display proinflammatory immune profiles and signatures of a hypercoagulable state associated with pulmonary embolism and cerebral infarction.  

While much emphasis was placed in cytokine profiling of critically sick Covid-19 patients, compelling data has emerged around the development of autoreactive antibodies that instead of targeting the virus, target the tissues of individuals infected with SARS-CoV-2. 

Initial studies implicated these autoantibodies in the formation of blood clots, which if left untreated were associated with poor patient outcome. Clinicians noted similarity between this characteristic of severe Covid-19 and the autoimmune condition Antiphospholipid Syndrome (APS). APS is characterized by the presence of antiphospholipid antibodies (aPLs) contributing to a hypercoagulable state and the development of blood clots. A study from China appears to confirm this link, by identifying aPLs in a significant number of critically ill Covid-19 patients (Xiao et. al. 2020). For many of these patients these autoantibodies were only transiently expressed and titers fell over time, but for others it appears that the infection has resulted in Covid-19-induced APS-like Syndrome, the long term consequences of which have yet to be determined. 

Another study has linked Covid-19 disease severity with the generation of antibodies targeting type I IFNs (Bastard et. al. 2020). This is not a new concept since autoantibodies against IFN-γ, IL-6 and IL-17A have been linked to infections with mycobacterium, staphylococcus and mucocutaneous candidiasis respectively. However, this study provided evidence for increased rates of autoantibody against type I IFNs in males, indicating that this is one of the reasons for increased risk of severe Covid-19 symptoms in males. Significantly, this study also brings to light clinical implications, and proposes that Covid-19 patients should be screened for auto-antibodies in order to identify individuals at high-risk of death, and to exclude patients with these auto-antibodies from donating convalescent plasma. Furthermore, clinical management of these patients with plasmapheresis, monoclonal Abs depleting plasmablasts, and the targeted inhibition of type I IFN-reactive B cells might be significantly more effective than current go-to therapeutic interventions.

It isn’t clear if autoantibodies play a direct pathogenic role in Covid-19 or not, however it is important to note that during the SARS-CoV-1 outbreak, researches identified cross-reactivity between anti-spike antibodies and Annexin 2, expressed in the epithelial cells of the respiratory system (Fang et. al. 2010), so it is possible that a similar occurrence is taking place in some patients infected with SARS-CoV-2.  

Researchers at Emory University (Woodruff et. al. 2020a) demonstrated a clear link between these rogue antibodies and the development of severe Covid-19 symptoms by inactivating critical components of the virus immune defense systems. This study identified a strong correlation of disease severity with the expression of autoantibodies and high levels of C Reactive Protein (CRP). Significantly, the most frequent autoantibodies identified in this study were antinuclear antibodies and rheumatoid factor, that are both commonly screened for by rheumatologists, indicating that this routine clinical screen, coupled with dexamethasone treatment may represent a cost effective way to identify high risk patients and provide a means of immunosuppression to dampen these potentially lethal autoimmune responses. 

So how could this type of autoimmunity develop so rapidly?  

Antibodies represent powerful tools in the humoral immune response, serving as beacons for the immune system to react efficiently to infection. However, if a B-cell starts making an antibody that targets ‘self’, this can result in the development of an autoimmune disease. Under normal circumstances, B-cells undergo a training process that enables antibodies to be fine tuned to the virus, and during this process B-cells generating antibodies to self would be destroyed. But this process takes time and during a virulent infection such as Covid-19, the time taken for the immune system to respond effectively could be the difference between life and death; under these circumstances, the immune system uses a process called extrafollicular activation that appears to bypass many of the checkpoints associated with a more precise and targeted response. Extrafollicular responses develop rapidly and by design are short-lived; but there is growing evidence that this may not always be the case, and prolonged extrafollicular immune system responses are being linked to many autoimmune conditions, particularly Lupus. Indeed, extrafollicular B cell signatures are now being identified in severe Covid-19 patients.  

Flow cytometry studies by Woodruff et al. (2020b, c) have shown that disease severity and poor patient outcome closely correlates with intense activation of the extrafollicular response of B cells, even if high titers of neutralizing antibodies may be detected. Antibody dependent functions are well described for both infection and autoimmunity and are mediated through several interacting mechanisms that include the generation of proinflammatory cytokines (IFN-γ, IL-6, GM-CSF and TNF-α). In the case of Covid-19, the origins of antibody secreting cells and the possible role of extrafollicular cells in this response is currently being researched. What has been demonstrated in patients with severe Covid-19 is the expansion of a unique population of CD11c+ activated naïve B cells that differentiate into effector cells lacking naïve (IgD) and memory (CD27) markers. These cells are induced through a process driven by IFN-γ, IL-21 in a TLR7-dependent manner and linked with elevated IL-6 and IP-10 serum levels. Significantly, both elevated IL-6 and IP-10 serum levels, as well as elevated CRP are strongly associated with poorer patient prognosis. Is it possible that extrafollicular B cell responses are responsible for some of the long term effects of Covid-19 on patients, that results in the perpetuation of symptoms, and may result in long term tissue damage or even other autoimmune disorders?

Final Thoughts

As scientific research is focused on the development of therapeutics, it is important to acknowledge  that in some cases of Covid-19, controlling the patient’s immune response to the virus might be as critical as controlling the virus itself. But understanding the possible immune responses to the virus may have significant implications not only on therapeutic strategies, but also on the engineering of safe vaccines to target the virus. Effector B-cells and T-cell responses provide the foundation of the development of specific immunological memory. Whereas we have an extensive understanding of T-cell memory, human effector B-cell responses and ASCs expansion are both poorly understood. How this will influence vaccine initiatives has yet to be flushed out, but the overall safety of vaccines must remain a priority.  

A very diverse clinical spectrum has become the hallmark of the SARS-CoV-2 pandemic, and although the immunologic foundations remain undefined, deep-diving into the immune system responses using high dimensional flow cytometry is revealing distinct correlations in response profiles. Hopefully this profiling will support more effective stratification and treatment of patients based on their risk of severe Covid-19 symptoms, and ultimately save lives.  

References

Kerr J.R. (2016) The role of parvovirus B19 in the pathogenesis of autoimmunity and autoimmune disease. J. Clin. Pathol. 69(4):279–291

Xiao, M. et. al. (2020) Antiphospholipid Antibodies in Critically Ill Patients with Covid-19. Arthritis & Rheumatology. 

https://doi.org/10.1002/art.41425

Bastard, P. et, al. (2020) Autoantibodies against type I IFNs in patients with life-threatening Covid-19. Science vol 370, issue 6515.

DOI: 10.1126/science.abd4585

Fang, Y. T. et al. (2010) Annexin A2 on lung epithelial cell surface is recognized by severe acute respiratory syndrome-associated coronavirus spike domain 2 antibodies. Mol. Immunol. 47, 1000–1009 

Woodruff, M. C. et. al. (2020a) Clinically Identifiable Autoreactivity is Common in Severe SARS-CoV-2 infection. (under submission posted October 28, 2020). 

https://doi.org/10.1101/2020.10.21.20216192d

Woodruff, M. C., et. al. (2020b) Extrafollicular B cell responses correlate with neutralizing antibodies and morbidity in Covid-19. Nature Immunology. Oct. 7^th 2020.

Woodruff, M. C., et. al. (2020c) Dominant extrafollicular B cell responses in severe Covid-19 disease correlate with robust viral-specific antibody production but poor outcomes. https://doi.org/10.1101/2020.04.29.20083717

Dr. Julie Bick is a medicinal biochemist who has spent close to 7 years with FlowMetric Life Sciences. After receiving her doctorate in Biochemistry at Southampton University in the UK, she began her career as Associate Professor at Rutgers University, NJ, before moving to the west coast to perform biomedical research with Syngenta and Novartis at the Torrey Mesa Research Institute in San Diego. Dr. Bick specializes in biomedical engineering of cells and proteins in order to provide innovative therapeutic and diagnostic solutions. She brings to FlowMetric a clinical expertise across a wide range of therapeutic areas from autoimmunity to oncology and chronic inflammatory conditions, acquired over 25 years of research experience in academic, biotechnology and pharmaceutical laboratories. In leading FlowMetric Life Sciences’ innovation initiatives, Dr. Bick has been collaborating with BurstIQ to implement Block Chain solutions into the company’s Contract Research Organization division, with a focus on enhanced big data analytics and process control solutions in the regulated clinical environment. Dr. Bick is committed to working with local Community Colleges to support STEM programs for the next generation of scientists.