Combination therapy with oral antiviral and anti-inflammatory drugs improves the efficacy of delayed treatment in severe COVID-19 (preprint)

Pulmonary infection with SARS-CoV-2 stimulates host immune responses and can also result in the progression of dysregulated and critical inflammation. Throughout the pandemic, the management and treatment of COVID-19 has been continuously updated with a range of antiviral drugs and immunomodulators. Monotherapy with oral antivirals has proven to be effective in the treatment of COVID-19. However, the treatment should be initiated in the early stages of infection to ensure beneficial therapeutic outcomes, and there is still room for further consideration on therapeutic strategies using antivirals. Here, we show that the oral antiviral ensitrelvir combined with the anti-inflammatory corticosteroid methylprednisolone has higher therapeutic effects and better outcomes in a delayed dosing model of SARS-CoV-2 infected hamsters compared to the monotherapy with ensitrelvir or methylprednisolone alone. Combination therapy with these drugs improved respiratory conditions and the development of pneumonia in hamsters even when the treatment was started after 2 days post infection. The combination therapy led to a differential histological and transcriptomic pattern in comparison to either of the monotherapies, with reduced lung damage and down-regulated expressions of genes involved in inflammatory response. Furthermore, we found that the combination treatment is effective in infection with both highly pathogenic delta and circulating omicron variants. Our results demonstrate the advantage of combination therapy with antiviral and corticosteroid drugs in COVID-19 treatment. Since both drugs are available as oral medications, this combination therapy could provide a clinical and potent therapeutic option for COVID-19.

Evidence for broad cross-reactivity of the SARS-CoV-2 NSP12-directed CD4+ T-cell response with pre-primed responses directed against common cold coronaviruses.

Introduction: The nonstructural protein 12 (NSP12) of the severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) has a high sequence identity with common cold coronaviruses (CCC). Methods: Here, we comprehensively assessed the breadth and specificity of the NSP12-specific T-cell response after in vitro T-cell expansion with 185 overlapping 15-mer peptides covering the entire SARS-CoV-2 NSP12 at single-peptide resolution in a cohort of 27 coronavirus disease 2019 (COVID-19) patients. Samples of nine uninfected seronegative individuals, as well as five pre-pandemic controls, were also examined to assess potential cross-reactivity with CCCs. Results: Surprisingly, there was a comparable breadth of individual NSP12 peptide-specific CD4+ T-cell responses between COVID-19 patients (mean: 12.82 responses; range: 0-25) and seronegative controls including pre-pandemic samples (mean: 12.71 responses; range: 0-21). However, the NSP12-specific T-cell responses detected in acute COVID-19 patients were on average of a higher magnitude. The most frequently detected CD4+ T-cell peptide specificities in COVID-19 patients were aa236-250 (37%) and aa246-260 (44%), whereas the peptide specificities aa686-700 (50%) and aa741-755 (36%), were the most frequently detected in seronegative controls. In CCC-specific peptide-expanded T-cell cultures of seronegative individuals, the corresponding SARS-CoV-2 NSP12 peptide specificities also elicited responses in vitro. However, the NSP12 peptide-specific CD4+ T-cell response repertoire only partially overlapped in patients analyzed longitudinally before and after a SARS-CoV-2 infection. Discussion: The results of the current study indicate the presence of pre-primed, cross-reactive CCC-specific T-cell responses targeting conserved regions of SARS-CoV-2, but they also underline the complexity of the analysis and the limited understanding of the role of the SARS-CoV-2 specific T-cell response and cross-reactivity with the CCCs.

Changes in Personal Protective Equipment Usage Among Healthcare Personnel From the Beginning of Pandemic to Intra-COVID-19 Pandemic in Thailand.

OBJECTIVES: Personal protective equipment (PPE) use is associated with reduced risk of SARS-CoV-2 infection among healthcare personnel (HCP). There are limited data on the impact of the novel coronavirus disease 2019 (COVID-19) pandemic on the PPE use of HCP. We describe the changes in PPE use from just before the widespread of community outbreaks ('pre-pandemic') to intra-pandemic time points, and examine factors associated with not changing in PPE use behavior among HCP in four Thai hospitals. METHODS: We performed a retrospective cohort evaluation using two-time points: (i) February-March 2020 (pre-pandemic period); and (ii) January-March 2021 (intra-pandemic period). Self-reported frequency of appropriate PPE use was measured by a Likert scale. We used multivariable logistic regression to identify factors associated with no increase in self-reported PPE use. RESULTS: Of 343 HCP, the proportion of participants reporting 'always' using PPE rose from 66% during the pre-pandemic period to 80% during the pandemic. Factors associated with HCP who did not increase in PPE use included having high baseline reported PPE, being a non-registered HCP (e.g. nurse assistants, dental assistants, porters), being male, and having a low perceived risk of becoming infected with any respiratory virus while working in the hospital. CONCLUSION: PPE education, training, and risk communication content should target all cadres of HCP, regardless of registered/non-registered status, with a focus on behavior change for improved prevention and control of SARS-CoV-2 and other respiratory viruses in healthcare settings.

Field-deployable multiplex detection method of SARS-CoV-2 and influenza virus using loop-mediated isothermal amplification and DNA chromatography.

A novel multiplex loop-mediated isothermal amplification (LAMP) method combined with DNA chromatography was developed for the simultaneous detection of three important respiratory disease-causing viruses: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, and influenza B virus. Amplification was performed at a constant temperature, and a positive result was confirmed by a visible colored band. An in-house drying protocol with trehalose was used to prepare the dried format multiplex LAMP test. Using this dried multiplex LAMP test, the analytical sensitivity was determined to be 100 copies for each viral target and 100-1000 copies for the simultaneous detection of mixed targets. The multiplex LAMP system was validated using clinical COVID-19 specimens and compared with the real-time qRT-PCR method as a reference test. The determined sensitivity of the multiplex LAMP system for SARS-CoV-2 was 71% (95% CI: 0.62-0.79) for cycle threshold (Ct) ≤ 35 samples and 61% (95% CI: 0.53-0.69) for Ct ≤40 samples. The specificity was 99% (95%CI: 0.92-1.00) for Ct ≤35 samples and 100% (95%CI: 0.92-1.00) for the Ct ≤40 samples. The developed simple, rapid, low-cost, and laboratory-free multiplex LAMP system for the two major important respiratory viral diseases, COVID-19 and influenza, is a promising field-deployable diagnosis tool for the possible future 'twindemic, ' especially in resource-limited settings.

Novel Therapeutic Target Critical for SARS-CoV-2 Infectivity and Induction of the Cytokine Release Syndrome.

We discovered a novel therapeutic target critical for SARS-CoV-2, cellular infectivity and the induction of the cytokine release syndrome. Here, we show that the mammalian enzyme neuraminidase-1 (Neu-1) is part of a highly conserved signaling platform that regulates the dimerization and activation of the ACE2 receptors and the Toll-like receptors (TLRs) implicated in the cytokine release syndrome (CRS). Activated Neu-1 cleaves glycosylated residues that provide a steric hindrance to both ACE2 and TLR dimerization, a process critical to both viral attachment to the receptor and entry into the cell and TLR activation. Blocking Neu-1 inhibited ACE2 receptor dimerization and internalization, TLR dimerization and activation, and the expression of several key inflammatory molecules implicated in the CRS and death from ARDS. Treatments that target Neu-1 are predicted to be highly effective against infection with SARS-CoV-2, given the central role played by this enzyme in viral cellular entry and the induction of the CRS.

An Early Evaluation of Remote Site Visits for ACGME Program Applications.

Background: Prior to the COVID-19 pandemic, accreditation site visit interviews occurred in-person. In response to the pandemic, the Accreditation Council for Graduate Medical Education (ACGME) developed a remote site visit protocol. Objective: To perform an early assessment of the remote accreditation site visits for programs applying for initial ACGME accreditation. Methods: A cohort of residency and fellowship programs that had remote site visits was evaluated from June to August 2020. Surveys were sent to program personnel, ACGME accreditation field representatives, and executive directors following the site visits. Comparison of accreditation decisions (Initial Accreditation or Accreditation Withheld) was completed for matched residency or fellowship programs having in-person site visits in 2019. Results: Surveys were sent to all program personnel from the 58 residency and fellowship programs that had remote site visits for new program applications, as well as the accreditation field representatives who performed the remote visits. The survey response rate was 58% (352 of 607). Ninety-one percent of all respondents were extremely or very confident that remote site visits provided a thorough assessment of proposed residency or fellowship programs. Fifty-four programs having remote site visits were matched by specialty to programs having had in-person program application site visits in 2019. Forty-six programs that had remote site visits received Initial Accreditation, and 52 programs that had in-person site visits in 2019 received Initial Accreditation (P=.093, 95% CI 0.91-22.38). Conclusions: Most program personnel and accreditation field representatives were confident that remote site visits conducted for program applications provided fair and thorough assessments of the program.

Designated antenatal clinic for pregnant women with COVID in the fifth wave of pandemic

We report our experience in setting up a designated antenatal clinic in the Kowloon East Cluster during the fifth wave of COVID pandemic for infected pregnant women.

Application of estimand framework in ICH E9 (R1) to safety evaluation.

Defining the right question of interest is important to a clinical study. ICH E9 (R1) introduces the framework of an estimand and its five attributes, which provide a basis for connecting different components of a study with its clinical questions. Most of the applications of the estimand framework focus on efficacy instead of safety assessment. In this paper, we expand the estimand framework into the safety evaluation and compare/contrast the similarity and differences between safety and efficacy estimand. Furthermore, we present and discuss applications of a safety estimand to oncology trials and pooled data analyses. At last, we also discuss the potential usage of safety estimand to handle the impacts of COVID-19 pandemic on safety assessment.

Evolution of Educational Commission for Foreign Medical Graduates Certification in the Absence of the USMLE Step 2 Clinical Skills Examination.

The United States Medical Licensing Examination Step 2 Clinical Skills (CS) was paused in 2020 because of the ongoing COVID-19 pandemic and discontinued in 2021. Step 2 CS was an important tool to assess readiness of international medical graduates (IMGs) to enter graduate medical education (GME) in the United States. This article describes the Educational Commission for Foreign Medical Graduates' (ECFMG's) response to the discontinuation of Step 2 CS. ECFMG certifies IMGs who seek eligibility for GME and licensure in the United States. Requirements for ECFMG certification include, among other factors, demonstration of adequate clinical skills and English proficiency, which were previously assessed as part of Step 2 CS. Beginning in June 2020 and during the next year, ECFMG modified the certification process with the introduction of 6 opportunities (pathways) for IMGs to demonstrate adequate clinical skills and a new test of English proficiency. In addition, permanent ECFMG certification is now granted only after the successful completion of the first year of residency, as determined by the program director. The COVID-19 pandemic and discontinuation of Step 2 CS caused a significant crisis for many IMGs who sought entrance into the United States, impacting the careers of those who had planned entry and those who would be eligible for U.S. training and the future workforce. Despite challenges due to the ongoing global pandemic, ECFMG certification continues to allow qualified physicians to enter U.S. GME and ensures that these individuals are ready to begin supervised training.

Asthma exacerbations: the Achilles heel of asthma care.

Asthma exacerbations significantly impact millions of patients worldwide to pose large disease burdens on affected patients, families, and health-care systems. Although numerous environmental factors cause asthma exacerbations, viral respiratory infections are the principal triggers. Advances in the pathophysiology of asthma have elucidated dysregulated protective immune responses and upregulated inflammation that create susceptibility and risks for exacerbation. Biologics for the treatment of severe asthma reduce rates of exacerbations and identify specific pathways of inflammation that contribute to altered pathophysiology, novel therapeutic targets, and informative biomarkers. Major steps to prevent exacerbations include the identification of molecular pathways whose blockage will prevent asthma attacks safely, predictably, and effectively.

Canadians' opinions towards COVID-19 data-sharing: a national cross-sectional survey.

OBJECTIVES: COVID-19 research has significantly contributed to pandemic response and the enhancement of public health capacity. COVID-19 data collected by provincial/territorial health authorities in Canada are valuable for research advancement yet not readily available to the public, including researchers. To inform developments in public health data-sharing in Canada, we explored Canadians' opinions of public health authorities sharing deidentified individual-level COVID-19 data publicly. DESIGN/SETTING/INTERVENTIONS/OUTCOMES: A national cross-sectional survey was administered in Canada in March 2022, assessing Canadians' opinions on publicly sharing COVID-19 datatypes. Market research firm Léger was employed for recruitment and data collection. PARTICIPANTS: Anyone greater than or equal to 18 years and currently living in Canada. RESULTS: 4981 participants completed the survey with a 92.3% response rate. 79.7% were supportive of provincial/territorial authorities publicly sharing deidentified COVID-19 data, while 20.3% were hesitant/averse/unsure. Datatypes most supported for being shared publicly were symptoms (83.0% in support), geographical region (82.6%) and COVID-19 vaccination status (81.7%). Datatypes with the most aversion were employment sector (27.4% averse), postal area (26.7%) and international travel history (19.7%). Generally supportive Canadians were characterised as being ≥50 years, with higher education, and being vaccinated against COVID-19 at least once. Vaccination status was the most influential predictor of data-sharing opinion, with respondents who were ever vaccinated being 4.20 times more likely (95% CI 3.21 to 5.48, p=0.000) to be generally supportive of data-sharing than those unvaccinated. CONCLUSIONS: These findings suggest that the Canadian public is generally favourable to deidentified data-sharing. Identifying factors that are likely to improve attitudes towards data-sharing are useful to stakeholders involved in data-sharing initiatives, such as public health agencies, in informing the development of public health communication and data-sharing policies. As Canada progresses through the COVID-19 pandemic, and with limited testing and reporting of COVID-19 data, it is essential to improve deidentified data-sharing given the public's general support for these efforts.

The DataHarmonizer: a tool for faster data harmonization, validation, aggregation and analysis of pathogen genomics contextual information.

Pathogen genomics is a critical tool for public health surveillance, infection control, outbreak investigations as well as research. In order to make use of pathogen genomics data, they must be interpreted using contextual data (metadata). Contextual data include sample metadata, laboratory methods, patient demographics, clinical outcomes and epidemiological information. However, the variability in how contextual information is captured by different authorities and how it is encoded in different databases poses challenges for data interpretation, integration and their use/re-use. The DataHarmonizer is a template-driven spreadsheet application for harmonizing, validating and transforming genomics contextual data into submission-ready formats for public or private repositories. The tool's web browser-based JavaScript environment enables validation and its offline functionality and local installation increases data security. The DataHarmonizer was developed to address the data sharing needs that arose during the COVID-19 pandemic, and was used by members of the Canadian COVID Genomics Network (CanCOGeN) to harmonize SARS-CoV-2 contextual data for national surveillance and for public repository submission. In order to support coordination of international surveillance efforts, we have partnered with the Public Health Alliance for Genomic Epidemiology to also provide a template conforming to its SARS-CoV-2 contextual data specification for use worldwide. Templates are also being developed for One Health and foodborne pathogens. Overall, the DataHarmonizer tool improves the effectiveness and fidelity of contextual data capture as well as its subsequent usability. Harmonization of contextual information across authorities, platforms and systems globally improves interoperability and reusability of data for concerted public health and research initiatives to fight the current pandemic and future public health emergencies. While initially developed for the COVID-19 pandemic, its expansion to other data management applications and pathogens is already underway.

A pseudovirus system enables deep mutational scanning of the full SARS-CoV-2 spike.

A major challenge in understanding SARS-CoV-2 evolution is interpreting the antigenic and functional effects of emerging mutations in the viral spike protein. Here, we describe a deep mutational scanning platform based on non-replicative pseudotyped lentiviruses that directly quantifies how large numbers of spike mutations impact antibody neutralization and pseudovirus infection. We apply this platform to produce libraries of the Omicron BA.1 and Delta spikes. These libraries each contain ∼7,000 distinct amino acid mutations in the context of up to ∼135,000 unique mutation combinations. We use these libraries to map escape mutations from neutralizing antibodies targeting the receptor-binding domain, N-terminal domain, and S2 subunit of spike. Overall, this work establishes a high-throughput and safe approach to measure how ∼105 combinations of mutations affect antibody neutralization and spike-mediated infection. Notably, the platform described here can be extended to the entry proteins of many other viruses.

Shark nanobodies with potent SARS-CoV-2 neutralizing activity and broad sarbecovirus reactivity.

Despite rapid and ongoing vaccine and therapeutic development, SARS-CoV-2 continues to evolve and evade, presenting a need for next-generation diverse therapeutic modalities. Here we show that nurse sharks immunized with SARS-CoV-2 recombinant receptor binding domain (RBD), RBD-ferritin (RFN), or spike protein ferritin nanoparticle (SpFN) immunogens elicit a set of new antigen receptor antibody (IgNAR) molecules that target two non-overlapping conserved epitopes on the spike RBD. Representative shark antibody variable NAR-Fc chimeras (ShAbs) targeting either of the two epitopes mediate cell-effector functions, with high affinity to all SARS-CoV-2 viral variants of concern, including the divergent Omicron strains. The ShAbs potently cross-neutralize SARS-CoV-2 WA-1, Alpha, Beta, Delta, Omicron BA.1 and BA.5, and SARS-CoV-1 pseudoviruses, and confer protection against SARS-CoV-2 challenge in the K18-hACE2 transgenic mouse model. Structural definition of the RBD-ShAb01-ShAb02 complex enabled design and production of multi-specific nanobodies with enhanced neutralization capacity, and picomolar affinity to divergent sarbecovirus clade 1a, 1b and 2 RBD molecules. These shark nanobodies represent potent immunotherapeutics both for current use, and future sarbecovirus pandemic preparation.

Early detection of SARS-CoV-2 variants using traveler-based genomic surveillance at four US airports, September 2021- January 2022.

We enrolled arriving international air travelers in SARS-CoV-2 genomic surveillance, using molecular testing of pooled nasal swabs, and sequencing positive samples for viral sublineage. Traveler-based genomic surveillance provided early warning variant detection; we reported the first U.S. Omicron BA.2 and first BA.3 in North America, weeks before next reported detection.

S-217622, a SARS-CoV-2 main protease inhibitor, decreases viral load and ameliorates COVID-19 severity in hamsters.

In parallel with vaccination, oral antiviral agents are highly anticipated to act as countermeasures for the treatment of the coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Oral antiviral medication demands not only high antiviral activity, but also target specificity, favorable oral bioavailability, and high metabolic stability. Although a large number of compounds have been identified as potential inhibitors of SARS-CoV-2 infection in vitro, few have proven to be effective in vivo. Here, we show that oral administration of S-217622 (ensitrelvir), an inhibitor of SARS-CoV-2 main protease (Mpro, also known as 3C-like protease), decreases viral load and ameliorates disease severity in SARS-CoV-2-infected hamsters. S-217622 inhibited viral proliferation at low nanomolar to sub-micromolar concentrations in cells. Oral administration of S-217622 demonstrated favorable pharmacokinetic properties and accelerated recovery from acute SARS-CoV-2 infection in hamster recipients. Moreover, S-217622 exerted antiviral activity against SARS-CoV-2 variants of concern (VOCs), including the highly pathogenic Delta variant and the recently emerged Omicron BA.5 and BA.2.75 variants. Overall, our study provides evidence that S-217622, an antiviral agent that is under evaluation in a phase 3 clinical trial (clinical trial registration no. jRCT2031210350), possesses remarkable antiviral potency and efficacy against SARS-CoV-2 and is a prospective oral therapeutic option for COVID-19.

Exploring the intersection of structural racism and ageism in healthcare.

The American Geriatrics Society (AGS) has consistently advocated for a healthcare system that meets the needs of older adults, including addressing impacts of ageism in healthcare. The intersection of structural racism and ageism compounds the disadvantage experienced by historically marginalized communities. Structural racism and ageism have long been ingrained in all aspects of US society, including healthcare. This intersection exacerbates disparities in social determinants of health, including poor access to healthcare and poor outcomes. These deeply rooted societal injustices have been brought into the forefront of the collective public consciousness at different points throughout history. The COVID-19 pandemic laid bare and exacerbated existing inequities inflicted on historically marginalized communities. Ageist rhetoric and policies during the COVID-19 pandemic further marginalized older adults. Although the detrimental impact of structural racism on health has been well documented in the literature, generative research on the intersection of structural racism and ageism is limited. The AGS is working to identify and dismantle the healthcare structures that create and perpetuate these combined injustices and, in so doing, create a more just US healthcare system. This paper is intended to provide an overview of important frameworks and to guide future efforts to both identify and eliminate bias within healthcare delivery systems and health professions training with a particular focus on the intersection of structural racism and ageism.

Influenza vaccination reveals sex dimorphic imprints of prior mild COVID-19.

Acute viral infections can have durable functional impacts on the immune system long after recovery, but how they affect homeostatic immune states and responses to future perturbations remain poorly understood1-4. Here we use systems immunology approaches, including longitudinal multimodal single-cell analysis (surface proteins, transcriptome and V(D)J sequences) to comparatively assess baseline immune statuses and responses to influenza vaccination in 33 healthy individuals after recovery from mild, non-hospitalized COVID-19 (mean, 151 days after diagnosis) and 40 age- and sex-matched control individuals who had never had COVID-19. At the baseline and independent of time after COVID-19, recoverees had elevated T cell activation signatures and lower expression of innate immune genes including Toll-like receptors in monocytes. Male individuals who had recovered from COVID-19 had coordinately higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared with healthy male individuals and female individuals who had recovered from COVID-19, in part because male recoverees had monocytes with higher IL-15 responses early after vaccination coupled with elevated prevaccination frequencies of 'virtual memory'-like CD8+ T cells poised to produce more IFNγ after IL-15 stimulation. Moreover, the expression of the repressed innate immune genes in monocytes increased by day 1 to day 28 after vaccination in recoverees, therefore moving towards the prevaccination baseline of the healthy control individuals. By contrast, these genes decreased on day 1 and returned to the baseline by day 28 in the control individuals. Our study reveals sex-dimorphic effects of previous mild COVID-19 and suggests that viral infections in humans can establish new immunological set-points that affect future immune responses in an antigen-agnostic manner.

Lay theories about emotion recognition explain cultural differences in willingness to wear facial masks during the COVID-19 pandemic.

Given that mask-wearing proved to be an important tool to slow the spread of infection during the COVID-19 pandemic, investigating the psychological and cultural factors that influence norms for mask wearing across cultures is exceptionally important. One factor that may influence mask wearing behavior is the degree to which people believe masks potentially impair emotion recognition. Based on previous research suggesting that there may be cultural differences in facial regions that people in Japan and the United States attend to when inferring a target's emotional state, we predicted that Americans would perceive masks (which cover the mouth) as more likely to impair emotion recognition, whereas Japanese would perceive facial coverings that conceal the eye region (sunglasses) to be more likely to impair emotion recognition. The results showed that Japanese participants reported wearing masks more than Americans. Americans also reported higher expected difficulty in interpreting emotions of individuals wearing masks (vs. sunglasses), while Japanese reported the reverse effect. Importantly, expectations about the negative impact of facial masks on emotion recognition explained cultural differences in mask-wearing behavior, even accounting for existing social norms.