Cross-reactive MHC Class I T Cell Epitopes May Dictate Heterologous Immune Responses Between Respiratory Viruses and Food Allergens (preprint)

Respiratory virus infections play a major role in asthma inception, persistence, and exacerbations. There is also a close correlation between asthma and food allergy, and we hypothesize that food-allergen-induced T cell-mediated heterologous immunity likely plays a role in inducing asthma symptoms in sensitized individuals. In this study, we used two independent in silico pipelines for the identification of cross-reactive virus- and food allergen- derived T cell epitopes, considering individual peptide sequence similarity, MHC binding affinity and immunogenicity. We assessed the proteomes of human rhinovirus (RV1b), respiratory syncytial virus (RSVA2) and influenza-strains contained in the seasonal quadrivalent influenza vaccine 2019/2020 (QIV 2019/2020), as well as SARS-CoV-2 for the most frequent human HLA alleles, in addition to more than 200 most common food allergen protein sequences. All resulting allergen-derived peptide candidates were subjected to an elaborate scoring system considering multiple criteria, including clinical relevance. In both bioinformatics approaches, we found that shortlisted peptide pairs that are potentially binding to MHC class II molecules scored up to 10x lower compared to MHC class I candidate epitopes. For MHC class I food allergen epitopes, several candidate peptides from shrimp, kiwi, apple, soy bean and chicken were identified. Such allergen sources contained potentially cross-reactive epitopes to the aforementioned viruses. The shortlisted set of peptide pairs may be implicated as heterologous virus-mediated immune response to food allergens. Our findings may be translated to peptide immunization strategies with immunomodulatory properties.

Comorbidity defines risk of asthmatics for COVID-19 hospitalization: a global perspective (preprint)

Background: The global epidemiology of asthma among COVID-19 patients presents striking geographic differences defining high and low [asthma and COVID-19] co-occurrence prevalence zones (1). The objective of the present study was to compare asthma prevalence among hospitalized COVID-19 patients in major global hubs across the world with the application of common inclusion criteria and definitions. Methods: : We built a network of six academic hospitals in Stanford (Stanford University)/USA, Frankfurt (Goethe University), Giessen (Justus Liebig University) and Marburg (Philipps University)/Germany, and Moscow (Clinical Hospital 52 in collaboration with Sechenov University)/Russia. We collected clinical and laboratory data for patients hospitalized due to COVID-19. Comorbidities reported were based on the 2020 International Classification of Diseases-10th Revision codes. Results: : Asthmatics were overrepresented among hospitalized COVID-19 patients in Stanford and underrepresented in Moscow and Germany as compared to the prevalence among adults in the local community. Asthma prevalence was similar among ICU and hospital non-ICU patients, which implied that the risk for developing severe COVID-19 was not higher among asthmatics. The number of males and comorbidities was higher among COVID-19 patients in the Stanford cohort, and the most frequent comorbidities among these asthma patients were other chronic inflammatory airway disorders such as chronic obstructive pulmonary disease (COPD). Conclusion: Observed disparity in COVID-19-associated risk among asthmatics across countries and continents is connected to varying prevalence of underlying comorbidities, particularly COPD. Public health policies in the future will need to consider comorbidities with an emphasis on COPD for prioritization of vaccination and preemptive treatment.

Autoantibodies targeting cytokines and connective tissue disease autoantigens are common in acute non-SARS-CoV-2 infections. (preprint)

The widespread presence of autoantibodies in acute infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is increasingly recognized, but the prevalence of autoantibodies in infections with organisms other than SARS-CoV-2 has not yet been reported. We used protein arrays to profile IgG autoantibodies from 317 samples from 268 patients across a spectrum of non-SARS-CoV-2 infections, many of whom were critically ill with pneumonia. Anti-cytokine antibodies (ACA) were identified in > 50% of patients infected with non-SARS-CoV-2 viruses and other pathogens, including patients with pneumonia attributed to bacterial causes. In cell-based functional assays, some ACA blocked binding to surface receptors for type I interferons (Type I IFN), granulocyte-macrophage colony-stimulating factor (GM-CSF), and interleukin-6 (IL-6). Autoantibodies against traditional autoantigens associated with connective tissue diseases (CTDs) were also commonly observed in these cohorts, including newly-detected antibodies that emerged in longitudinal samples from patients infected with influenza. We conclude that autoantibodies, some of which are functionally active, may be much more prevalent than previously appreciated in patients who are symptomatically infected with diverse pathogens.

Single cell RNA-seq uncovers the nuclear decoy lincRNA PIRAT as a regulator of systemic monocyte immunity during COVID-19 (preprint)

The systemic immune response to viral infection is shaped by master transcription factors such as NF{kappa}B or PU.1. Although long non-coding RNAs (lncRNAs) have been suggested as important regulators of transcription factor activity, their contributions to the systemic immunopathologies observed during SARS-CoV-2 infection have remained unknown. Here, we employed a targeted single-cell RNA-seq approach to reveal lncRNAs differentially expressed in blood leukocytes during severe COVID-19. Our results uncover the lncRNA PIRAT as a major PU.1 feedback-regulator in monocytes, governing the production of the alarmins S100A8/A9 - key drivers of COVID-19 pathogenesis. Knockout and transgene expression, combined with chromatin-occupancy profiling characterized PIRAT as a nuclear decoy RNA, diverting the PU.1 transcription factor from alarmin promoters to dead-end pseudogenes in naive monocytes. NF{kappa}B-dependent PIRAT down-regulation during COVID-19 consequently releases a transcriptional brake, fueling alarmin production. Our results suggest a major role of nuclear noncoding RNA circuits in systemic antiviral responses to SARS-CoV-2 in humans.

New-Onset IgG Autoantibodies in Hospitalized Patients with COVID-19 (preprint)

Coronavirus Disease 2019 (COVID-19), caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), is associated with a wide range of clinical manifestations, including autoimmune features and autoantibody production. We developed three different protein arrays to measure hallmark IgG autoantibodies associated with Connective Tissue Diseases (CTDs), Anti-Cytokine Antibodies (ACA), and anti-viral antibody responses in 147 hospitalized COVID-19 patients in three different centers. Autoantibodies were identified in approximately 50% of patients, but in <15% of healthy controls. When present, autoantibodies largely targeted autoantigens associated with rare disorders such as myositis, systemic sclerosis and CTD overlap syndromes. Anti-nuclear antibodies (ANA) were observed in ~25% of patients. Patients with autoantibodies tended to demonstrate one or a few specificities whereas ACA were even more prevalent, and patients often had antibodies to multiple cytokines. Rare patients were identified with IgG antibodies against angiotensin converting enzyme-2 (ACE-2). A subset of autoantibodies and ACA developed de novo following SARS-CoV-2 infection while others were transient. Autoantibodies tracked with longitudinal development of IgG antibodies that recognized SARS-CoV-2 structural proteins such as S1, S2, M, N and a subset of non-structural proteins, but not proteins from influenza, seasonal coronaviruses or other pathogenic viruses. COVID-19 patients with one or more autoantibodies tended to have higher levels of antibodies against SARS-CoV-2 Nonstructural Protein 1 (NSP1) and Methyltransferase (ME). We conclude that SARS-CoV-2 causes development of new-onset IgG autoantibodies in a significant proportion of hospitalized COVID-19 patients and are positively correlated with immune responses to SARS-CoV-2 proteins.

Recombination and low-diversity confound homoplasy-based methods to detect the effect of SARS-CoV-2 mutations on viral transmissibility (preprint)

The SARS-CoV-2 variant carrying the Spike protein mutation G614 was first detected in late January 2020 and within a few months became the dominant form globally. In the months that followed, many studies, both in vitro and in animal models, showed that variants carrying this mutation were more infectious and more readily transmitted than the ancestral Wuhan form. Here we investigate why a recently published study by van Dorp et al. failed to detect such higher transmissibility of the G614 variant using homoplasy-based methods. We show that both low diversity and recombination confound the methods utilized by van Dorp et al. and significantly decrease their sensitivity. Furthermore, though they claim no evidence of recombination in their dataset, we and several other studies identify a subset of the sequences as recombinants, possibly enough to affect their statistic adversely.

The evolutionary making of SARS-CoV-2 (preprint)

Covid-19 is the most devastating pandemic of the past 100 years. A zoonotic transfer presumably at a wildlife market introduced the causative virus, SARS-CoV-2 (sarbecovirus; beta-coronavirus), to humans in late 2019. Meanwhile, the mechanistic details of the infection process have been largely elucidated, and structural models explain binding of the virial spike to the human cell surface receptor ACE2. Yet, the evolutionary trajectory that gave rise to this pathogen is poorly understood. Here we scan SARS-CoV-2 protein sequences in-silico for innovations along the evolutionary lineage starting with the last common ancestor of coronaviruses. Substantial differences in the sets of proteins encoded by SARS-CoV-2 and viruses outside sarbecovirus, and in their domain architectures, indicate divergent functional demands. By contrast, sarbecoviruses themselves are almost fully conserved at these levels of resolution. However, profiling spike evolution on the sub-domain level using predicted linear epitopes reveals that this protein was gradually reshaped within sarbecovirus. The only epitope that is private to SARS-CoV-2 overlaps with the furin cleavage site. This lends phylogenetic support to the hypothesis that a change in strategy facilitated the zoonotic transfer of SARS-CoV-2 and its success as a human pathogen. Upon furin cleavage, spike switches from a "stealth mode" where immunodominant ACE2 binding epitopes are largely hidden to an "attack mode" where these epitopes are exposed. The resulting reinforcement of ACE2 binding extends the window of opportunity for cell entry. SARS-CoV-2 variants fine-tuning this mode switch will be particularly threatening as they optimize immune evasion.

Information retrieval in an infodemic: the case of COVID-19 publications (preprint)

In the context of searching for COVID-19 related scientific literature, we present an information retrieval methodology for effectively finding relevant publications for different information needs. We discuss different components of our architecture consisting of traditional information retrieval models, as well as modern neural natural language processing algorithms. We present recipes to better adapt these components to the case of an infodemic, where, from one hand, the number of publications has an exponential growth and, from the other hand, the topics of interest evolve as the pandemic progresses. The methodology was evaluated in the TREC-COVID challenge, achieving competitive results with top ranking teams participating in the competition. In retrospect to this challenge, we provide additional insights with further useful impacts.

The SARS-CoV-2 Y453F mink variant displays a striking increase in ACE-2 affinity but does not challenge antibody neutralization (preprint)

SARS-CoV-2 transmission from humans to animals has been reported for many domesticated species, including cats, dogs and minks. Identification of novel spike gene mutations appearing in minks has raised major concerns about potential immune evasion and challenges for the global vaccine strategy. The genetic variant, known as cluster-five, arose among farmed minks in Denmark and resulted in a complete shutdown of the worlds largest mink production. However, the functional properties of this new variant are not established. Here we present functional data on the Y453F cluster-five receptor-binding domain (RBD) and show that it does not decrease established humoral immunity or affect the neutralizing response in a vaccine model based on wild-type RBD or spike. However, it binds the human ACE-2 receptor with a four-fold higher affinity suggesting an enhanced transmission capacity and a possible challenge for viral control.

Development of spike receptor-binding domain nanoparticle as a vaccine candidate against SARS-CoV-2 infection in ferrets (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a causative agent of COVID-19 pandemic, enters host cells via the interaction of its Receptor-Binding Domain (RBD) of Spike protein with host Angiotensin-Converting Enzyme 2 (ACE2). Therefore, RBD is a promising vaccine target to induce protective immunity against SARS-CoV-2 infection. In this study, we report the development of RBD protein-based vaccine candidate against SARS-CoV-2 using self-assembling H. pylori-bullfrog ferritin nanoparticles as an antigen delivery. RBD-ferritin protein purified from mammalian cells efficiently assembled into 24-mer nanoparticles. 16-20 months-old ferrets were vaccinated with RBD-ferritin nanoparticles (RBD-nanoparticles) by intramuscular or intranasal inoculation. All vaccinated ferrets with RBD-nanoparticles produced potent neutralizing antibodies against SARS-CoV-2. Strikingly, vaccinated ferrets demonstrated efficient protection from SARS-CoV-2 challenge, showing no fever, body weight loss and clinical symptoms. Furthermore, vaccinated ferrets showed rapid clearance of infectious viruses in nasal washes and lungs as well as viral RNA in respiratory organs. This study demonstrates the Spike RBD-nanoparticle as an effective protein vaccine candidate against SARS-CoV-2.

Homologies between SARS-CoV-2 and allergen proteins may direct T cell-mediated heterologous immune responses (preprint)

The outbreak of the new Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is a public health emergency. Asthma does not represent a risk factor for COVID-19 in several published cohorts. We hypothesized that the SARS-CoV-2 proteome contains T cell epitopes, which are potentially cross-reactive to allergen epitopes. We aimed at identifying homologous peptide sequences by means of two distinct complementary bioinformatics approaches. Pipeline 1 included prediction of MHC Class I and Class II epitopes contained in the SARS-CoV-2 proteome and allergens along with alignment and elaborate ranking approaches. Pipeline 2 involved alignment of SARS-CoV-2 overlapping peptides with known allergen-derived T cell epitopes. Our results indicate a large number of MHC Class I epitope pairs including known as well as de novo predicted allergen T cell epitopes with high probability for cross-reactivity. Allergen sources, such as Aspergillus fumigatus, Phleum pratense and Dermatophagoides species are of particular interest due to their association with multiple cross-reactive candidate peptides, independently of the applied bioinformatic approach. In contrast, peptides derived from food allergens, as well as MHC class II epitopes did not achieve high in silico ranking and were therefore not further investigated. Our findings warrant further experimental confirmation along with examination of the functional importance of such cross-reactive responses.

Review and methodological analysis of trials currently testing treatment and prevention options for the novel coronavirus disease (COVID-19) globally. (preprint)

Background: As novel coronavirus disease (COVID-19) cases continue to steeply rise globally within an unprecedented short period of time, solid evidence from large randomised controlled trials is still lacking. Currently, numerous trials testing potential treatment and preventative options are undertaken globally. Objectives: We summarised all currently registered clinical trials examining treatment and prevention options for COVID-19 pneumonia. Additionally, we evaluated the quality of the retrieved interventional studies. Data sources: The ClinicalTrials.gov, the Chinese Clinical Trial Registry and the European Union Clinical Trials Register were systematically searched. Study eligibility criteria: Registered clinical trials examining treatment and/or prevention options for COVID-19 were included. No language, country or study design restrictions were applied. Withdrawn, cancelled studies and trials not reporting therapeutic or preventative strategies for COVID-19 were excluded. Participants and interventions: No restrictions in terms of participants' age and medical background or type of intervention were enforced. Methods: The registries were searched using the term "coronavirus" or "COVID-19" from their inception until 26th March 2020. Additional manual search of the registries was also performed. Eligible studies were summarised and tabulated. Interventional trials were methodologically analysed, excluding expanded access studies and trials testing Traditional Chinese Medicine. Results: In total, 309 trials evaluating therapeutic management options, 23 studies assessing preventive strategies and 3 studies examining both were retrieved. Interventional treatment studies were mostly randomised (n=150, 76%) and open-label (n=73, 37%) with a median number of planned inclusions of 90 (IQR 40-200). Major categories of interventions that are currently being investigated are discussed. Conclusion: Numerous clinical trials have been registered since the onset of the COVID-19 pandemic. Summarised data on these trials will assist physicians and researchers to promote patient care and guide future research efforts for COVID-19 pandemic containment. However, up to the end of March, 2020, significant information concerning reported trials was lacking.