Toll-like receptor 7 (TLR7)-mediated antiviral response protects mice from lethal SARS-CoV-2 infection (preprint)

SARS-CoV-2-induced impaired antiviral and excessive inflammatory responses cause fatal pneumonia. However, the key pattern recognition receptors that elicit effective antiviral and lethal inflammatory responses in-vivo are not well defined. CoVs possess single-stranded RNA (ssRNA) that is abundantly produced during infection and stimulates both antiviral interferon (IFN) and inflammatory cytokine/ chemokine responses. Therefore, in this study, using wild-type control and TLR7 deficient BALB/c mice infected with a mouse-adapted SARS-COV-2 (MA-CoV-2), we evaluated the role of TLR7 signaling in MA-CoV-2-induced antiviral and inflammatory responses and disease outcome. We show that TLR7-deficient mice are more susceptible to MA-CoV-2 infection as compared to infected control mice. Further evaluation of MA-CoV-2 infected lungs showed significantly reduced mRNA levels of antiviral type I and type III IFNs, IFN stimulated genes (ISGs, ISG15 and CXCL10), and several pro-inflammatory cytokines/chemokines in TLR7 deficient compared to control mice. Reduced lung IFN/ISG levels and increased morbidity/mortality in TLR7 deficient mice correlated with high lung viral titer. Detailed examination of total cells from MA-CoV-2 infected lungs showed high neutrophil count in TLR7 deficient mice compared to control mice. Additionally, blocking TLR7 activity post-MA-CoV-2 infection using a specific inhibitor also enhanced disease severity. In summary, our results conclusively establish that TLR7 signaling is protective during SARS-CoV-2 infection, and despite robust inflammatory response, TLR7-mediated IFN/ISG responses likely protect the host from lethal disease. Given similar outcomes in control and TLR7 deficient humans and mice, these results show that MA-CoV-2 infected mice serve as excellent model to study COVID-19.

Safety and immunogenicity of SARS-CoV-2 self-amplifying RNA vaccine expressing anchored RBD: a randomised, observer-blind, phase 1 study (preprint)

BACKGROUND VLPCOV-01 is a lipid nanoparticle-encapsulated self-amplifying (sa) RNA vaccine that expresses a membrane-anchored receptor-binding domain (RBD) derived from the SARS-CoV-2 spike protein. METHODS A phase 1 study of VLPCOV-01 was conducted at Medical Corporation Heishinkai OPHAC Hospital, Japan. The investigational vaccines were administered to participants, between February 16, 2022, and March 17, 2022. Participants aged 18 to 55 or [≥]65 years who had completed two doses of the BNT162b2 mRNA vaccine 6 to 12 months previously were randomised to receive one intramuscular vaccination of 0{middle dot}3, 1{middle dot}0, or 3{middle dot}0 {micro}g VLPCOV-01, 30 {micro}g BNT162b2, or placebo. Solicited adverse events were collected up to 6 days post-administration, with follow-up on all adverse events until week 4. Interim immunogenicity analyses following data cutoff at day 29 included SARS-CoV-2 IgG and neutralising antibody titres. (The trial is registered: jRCT2051210164). FINDINGS 92 healthy adults were enrolled, with 60 participants receiving VLPCOV-01. No serious adverse events were reported up to 26 weeks, and no prespecified trial-halting events were met. VLPCOV-01 induced robust IgG titres against SARS-CoV-2 RBD protein that were maintained up to 26 weeks in non-elderly participants, with geometric means ranging from 5037 (95% CI 1272-19,940) at 0{middle dot}3 {micro}g to 12,873 (95% CI 937-17,686) at 3 {micro}g, in comparison to 3166 (95% CI 1619-6191) with 30 {micro}g BNT162b2. Among elderly participants, IgG titres at 26 weeks post-vaccination with 3 {micro}g VLPCOV-01 were 9865 (95% CI 4396-22138) compared to 4183 (95% CI 1436-12180) following vaccination with 30 {micro}g BNT162b2. Pseudovirus-Neutralising antibody responses were observed against multiple SARS-CoV-2 variants and strongly correlated with anti-SARS-CoV-2 IgG (r=0{middle dot}950, p<0{middle dot}001). INTERPRETATION VLPCOV-01 is immunogenic following low dose administration, with anti-SARS-CoV-2 immune responses comparable to BNT162b2. These findings support further development of VLPCOV-01 as a COVID-19 booster vaccine and the potential for saRNA vectors as a vaccine platform. FUNDING Supported by AMED, Grant No. JP21nf0101627.

Scoping Review of Existing Stroke Guidelines; Argument for a Value-Added Change (preprint)

Background and Purpose Stroke represents one of the most important causes of morbidity ( eighty million patients with disabling of ongoing effects of stroke at a given time, globally) and mortality (the second leading cause of death) worldwide. Innovative systems biology-based approach is likely to increase the understanding of the underpinning of acute stroke promise to enhance stroke prevention, acute treatment, and neurorehabilitation. Recent growing body of evidence with shared pathobiology with COVID-19 and the critically important role of inflammation in the context of stroke points to far-reaching consequences of acute stroke, just as in the case of COVID-19 ( post-acute event issues as well as long term issues). So far, stroke typically defined by late-appearing disease manifestation by the range of stroke subtypes as defined by the WHO or American Stroke Association. This definition neglects the underlying pathobiological mechanisms such as low-grade chronic inflammation and already compromised vascular system. Diseases such as stroke is hardly a simple result of a single problem, but rather a complex cascade of pathobiological processes and interactions in a complex biochemical environment. The evidence of changes in innate immunity and adaptive immunity during the index event of acute stroke and recovery over next 3-12 months can be easily elicited with simple bedside blood tests such as neutrophil-lymphocyte ratio (NLR) with well over 300 published papers including several systematic reviews and meta-analyses confirming this. Global standard operating procedures (SOP) of stroke care dictated by the national and international stroke guidelines at present. It is imperative to explore the evidence of systems biology approach in current stroke guidelines. This is likely to be a key turning point in managing stroke across the continuum (prevention, management of acute event and rehabilitation). Methods We systematically searched for guideline recommendation on the day-to-day use of peripheral inflammatory markers such as NLR published in the English language between January 1, 2005, and December 2020. Any other evidence of systems biology-based approach or recommendation was explored within the selected guidelines for this scoping review. Only the latest guideline per writing group was selected. Each guideline was analyzed independently by 2 to 4 authors to determine clinical scenarios explained/given, scientific evidence used, and recommendations presented in the context of systems biology. Results The scoping review found 3,830 (3830) titles with 119 guidelines from 46 countries included for this review ( Figure 1; PRISMA diagram). Stroke-related organizations wrote Sixty-five per cent of the guidelines while national ministries wrote a fewer number of guidelines. We were primarily interested in recommendations for acute management in AIS published in the English language. Fifteen eligible guidelines were identified from 15 different countries/regions. None of the guidelines recommended the routine use of peripheral markers of inflammation, such as NLR, among their acute assessment and management recommendations. None of the existing guidelines explored the systems biology approach to one of the most complex diseases affecting the human brain, stroke. Figure 1 Acute Ischemic Stroke Guidelines Worldwide Figure 2: PRISMA Diagram Conclusions This systematic review has identified a significant evidence-practice gap in all existing national stroke guidelines published in English medium as of October 2020. These guidelines included the only current “living stroke guidelines, Stroke Guidelines from Australia with a real opportunity to modernize the living stroke guidelines with systems biology approach and provide 2020 vision towards better stroke care globally. Investigation of complex disease such as stroke is best served through a systems biology approach. One of the easiest places to start is simple blood tests such as total white cell count and NLR. Systems biology approach point us towards simple tools such immune-inflammatory index (SII), Sunshine Prognostic Score (SPS) which should pave the way for the stroke physician community address the challenges in systems biology approach in stroke care. These challenges include translating bench research to the bedside, managing big data ( continuous pulse, blood pressure, sleep, Oxygen saturation, progressive changes in NLR, SII, SPS, etc.). Working with an interdisciplinary team is also provide a distinct advantage.

The majority of male patients with COVID-19 present low testosterone levels on admission to Intensive Care in Hamburg, Germany: a retrospective cohort study. (preprint)

Background. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread worldwide and pose a major public health burden. There is increasing evidence that men are more likely to die from SARS-CoV-2 infection than women. However, underlying factors that mediate the observed sex bias in coronavirus disease 2019 (COVID-19) remain unknown. Methods. In this retrospective cohort, we included COVID-19 patients who were admitted to an Intensive Care Unit at the University Hospital Hamburg-Eppendorf, Germany. We obtained demographic data of all patients who were discharged or had died by 29th April 2020. We systematically analyzed sex hormones as well as cytokine and chemokine responses in male and female patients with laboratory-confirmed SARS-CoV-2 infections upon hospital admission. We used uni- and multivariable linear regression methods to identify potential risk factors for disease severity in males and females. Findings. All enrolled patients (n=45; n=35 males and n=10 females) presented comorbidities with hypertension being the most common (45.7% in males; 40% in females), followed by cancer (35% in males; 40% in females), obesity (34.3% in males and 30% in females), type II diabetes (25.7% in males and 20% in females) and chronic heart diseases (8.6% in males and 0% in females). We detected that the vast majority of male COVID-19 patients present low testosterone (68.6%) and low dihydrotestosterone (48.6%) levels. In contrast, most female COVID-19 patients have elevated testosterone levels (60%) without alterations in dihydrotestosterone levels. Both, female and male COVID-19 patients may present elevated estradiol levels (45.7% in males and 40% in females). Disease severity defined by SOFA score correlates with elevated cytokine responses (e.g. IL-6) in males and IL-2 in females. In male COVID-19 patients, testosterone levels negatively correlate with inflammatory IL-2 and IFN-{gamma}, whereas estradiol levels positively correlate with the inflammatory cytokine IL-6. Vice versa, in female COVID-19 patients, testosterone levels positively correlate with inflammatory cytokines (e.g. IL-6). Interpretation. We here show that critically ill male COVID-19 patients suffer from severe testosterone and dihydrotestosterone deficiencies. Both androgens are required to mount antiviral immune responses to combat infection in males.