Antiviral efficacy of favipiravir against Zika and SARS-CoV-2 viruses in non-human primates.

The COVID-19 pandemic has exemplified that rigorous evaluation in large animal models is key for translation from promising in vitro results to successful clinical implementation. Among the drugs that have been largely tested in clinical trials but failed so far to bring clear evidence of clinical efficacy is favipiravir, a nucleoside analogue with large spectrum activity against several RNA viruses in vitro and in small animal models. Here, we evaluate the antiviral activity of favipiravir against Zika or SARS-CoV-2 virus in cynomolgus macaques. In both models, high doses of favipiravir are initiated before infection and viral kinetics are evaluated during 7 to 15 days after infection. Favipiravir leads to a statistically significant reduction in plasma Zika viral load compared to untreated animals. However, favipiravir has no effects on SARS-CoV-2 viral kinetics, and 4 treated animals have to be euthanized due to rapid clinical deterioration, suggesting a potential role of favipiravir in disease worsening in SARS-CoV-2 infected animals. To summarize, favipiravir has an antiviral activity against Zika virus but not against SARS-CoV-2 infection in the cynomolgus macaque model. Our results support the clinical evaluation of favipiravir against Zika virus but they advocate against its use against SARS-CoV-2 infection.

Vaccine-associated enhanced disease in humans and animal models: Lessons and challenges for vaccine development.

The fight against infectious diseases calls for the development of safe and effective vaccines that generate long-lasting protective immunity. In a few situations, vaccine-mediated immune responses may have led to exacerbated pathology upon subsequent infection with the pathogen targeted by the vaccine. Such vaccine-associated enhanced disease (VAED) has been reported, or at least suspected, in animal models, and in a few instances in humans, for vaccine candidates against the respiratory syncytial virus (RSV), measles virus (MV), dengue virus (DENV), HIV-1, simian immunodeficiency virus (SIV), feline immunodeficiency virus (FIV), severe acute respiratory syndrome coronavirus 1 (SARS-CoV-1), and the Middle East respiratory syndrome coronavirus (MERS-CoV). Although alleviated by clinical and epidemiological evidence, a number of concerns were also initially raised concerning the short- and long-term safety of vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is causing the ongoing COVID-19 pandemic. Although the mechanisms leading to this phenomenon are not yet completely understood, the individual and/or collective role of antibody-dependent enhancement (ADE), complement-dependent enhancement, and cell-dependent enhancement have been highlighted. Here, we review mechanisms that may be associated with the risk of VAED, which are important to take into consideration, both in the assessment of vaccine safety and in finding ways to define models and immunization strategies that can alleviate such concerns.

Modelling the response to vaccine in non-human primates to define SARS-CoV-2 mechanistic correlates of protection.

The definition of correlates of protection is critical for the development of next-generation SARS-CoV-2 vaccine platforms. Here, we propose a model-based approach for identifying mechanistic correlates of protection based on mathematical modelling of viral dynamics and data mining of immunological markers. The application to three different studies in non-human primates evaluating SARS-CoV-2 vaccines based on CD40-targeting, two-component spike nanoparticle and mRNA 1273 identifies and quantifies two main mechanisms that are a decrease of rate of cell infection and an increase in clearance of infected cells. Inhibition of RBD binding to ACE2 appears to be a robust mechanistic correlate of protection across the three vaccine platforms although not capturing the whole biological vaccine effect. The model shows that RBD/ACE2 binding inhibition represents a strong mechanism of protection which required significant reduction in blocking potency to effectively compromise the control of viral replication.

Detection of SARS-CoV-2 in subcutaneous fat but not visceral fat, and the disruption of fat lymphocyte homeostasis in both fat tissues in the macaque.

The well documented association between obesity and the severity of SARS-CoV-2 infection raises the question of whether adipose tissue (AT) is impacted during this infection. Using a model of SARS-CoV-2 infection in cynomolgus macaques, we detected the virus within subcutaneous AT (SCAT) but not in visceral AT (VAT) or epicardial AT on day 7 post-infection. We sought to determine the mechanisms responsible for this selective detection and observed higher levels of angiotensin-converting-enzyme-2 mRNA expression in SCAT than in VAT. Lastly, we evaluated the immunological consequences of SARS-CoV-2 infection on AT: both SCAT and VAT T cells showed a drastic reduction in CD69 expression, a standard marker of resident memory T cell in tissue, that is also involved in the migratory and metabolic properties of T cells. Our results demonstrate that in a model of mild infection, SCAT is selectively infected by SARS-CoV-2 although changes in the immune properties of AT are observed in both SCAT and VAT.

A Case Study to Dissect Immunity to SARS-CoV-2 in a Neonate Nonhuman Primate Model.

Most children are less severely affected by coronavirus-induced disease 2019 (COVID-19) than adults, and thus more difficult to study progressively. Here, we provide a neonatal nonhuman primate (NHP) deep analysis of early immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in blood and mucosal tissues. In addition, we provide a comparison with SARS-CoV-2-infected adult NHP. Infection of the neonate resulted in a mild disease compared with adult NHPs that develop, in most cases, moderate lung lesions. In concomitance with the viral RNA load increase, we observed the development of an early innate response in the blood, as demonstrated by RNA sequencing, flow cytometry, and cytokine longitudinal data analyses. This response included the presence of an antiviral type-I IFN gene signature, a persistent and lasting NKT cell population, a balanced peripheral and mucosal IFN-γ/IL-10 cytokine response, and an increase in B cells that was accompanied with anti-SARS-CoV-2 antibody response. Viral kinetics and immune responses coincided with changes in the microbiota profile composition in the pharyngeal and rectal mucosae. In the mother, viral RNA loads were close to the quantification limit, despite the very close contact with SARS-CoV-2-exposed neonate. This pilot study demonstrates that neonatal NHPs are a relevant model for pediatric SARS-CoV-2 infection, permitting insights into the early steps of anti-SARS-CoV-2 immune responses in infants.

Durable immunogenicity, adaptation to emerging variants, and low-dose efficacy of an AAV-based COVID-19 vaccine platform in macaques.

The COVID-19 pandemic continues to have devastating consequences on health and economy, even after the approval of safe and effective vaccines. Waning immunity, the emergence of variants of concern, breakthrough infections, and lack of global vaccine access and acceptance perpetuate the epidemic. Here, we demonstrate that a single injection of an adenoassociated virus (AAV)-based COVID-19 vaccine elicits at least 17-month-long neutralizing antibody responses in non-human primates at levels that were previously shown to protect from viral challenge. To improve the scalability of this durable vaccine candidate, we further optimized the vector design for greater potency at a reduced dose in mice and non-human primates. Finally, we show that the platform can be rapidly adapted to other variants of concern to robustly maintain immunogenicity and protect from challenge. In summary, we demonstrate this class of AAV can provide durable immunogenicity, provide protection at dose that is low and scalable, and be adapted readily to novel emerging vaccine antigens thus may provide a potent tool in the ongoing fight against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).

Durable immunogenicity, adaptation to emerging variants and low dose efficacy of AAV-based COVID19 platform in macaques

The COVID19 pandemic continues to have devastating consequences on health and economy, even after the approval of safe and effective vaccines. Waning immunity, the emergence of variants of concern, breakthrough infections, and lack of global vaccine access and acceptance perpetuate the epidemic. Here, we demonstrate that a single injection of an AAV-based COVID19 vaccine elicits at least 17-month-long neutralizing antibody responses in non-human primates at levels that were previously shown to protect from viral challenge. To improve the scalability of this durable vaccine candidate, we further optimized the vector design for greater potency at a reduced dose in mice and nonhuman primates. Finally, we show that the platform can be rapidly adapted to other variants of concern to robustly maintain immunogenicity and protect from challenge. In summary, we demonstrate this class of AAV can provide durable immunogenicity, provide protection at dose that is low and scalable, and be adapted readily to novel emerging vaccine antigens thus may provide potent tool in the ongoing fight against SARS-CoV-2. Graphical This manuscript characterizes and optimizes an AAV-based vaccine platform for several COVID-19 development candidates: durability of humoral response at high level for over 20 months, the ability to reduce the dose and protect from challenge in NHP and the versatility and robustness of the platform across different variant of concern antigens.

Computed tomography and [18F]-FDG PET imaging provide additional readouts for COVID-19 pathogenesis and therapies evaluation in non-human primates.

Non-human primates (NHPs) are particularly relevant as preclinical models for SARS-CoV-2 infection and nuclear imaging may represent a valuable tool for monitoring infection in this species. We investigated the benefit of computed X-ray tomography (CT) and [18F]-FDG positron emission tomography (PET) to monitor the early phase of the disease in a large cohort (n = 76) of SARS-CoV-2 infected macaques. Following infection, animals showed mild COVID-19 symptoms including typical lung lesions. CT scores at the acute phase reflect the heterogeneity of lung burden following infection. Moreover, [18F]-FDG PET revealed that FDG uptake was significantly higher in the lungs, nasal cavities, lung-draining lymph nodes, and spleen of NHPs by 5 days postinfection compared to pre-infection levels, indicating early local inflammation. The comparison of CT and PET data from previous COVID-19 treatments or vaccines we tested in NHP, to this large cohort of untreated animals demonstrated the value of in vivo imaging in preclinical trials.

Validation of the Performance of A1HPV6, a Triage Blood Test for the Early Diagnosis and Prognosis of SARS-CoV-2 Infection.

BACKGROUND AND AIMS: Apolipoprotein A1 (A1) and haptoglobin (HP) serum levels are associated with the spread and severity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We have constructed and validated a multivariable risk calculator (A1HPV6) integrating A1, HP, alpha2-macroglobulin, and gamma glutamyl transferase to improve the performances of virological biomarkers. METHODS: In a prospective observational study of hospitalized patients with nonsevere SARS-CoV-2 infection, A1HPV6 was constructed in 127 patients and validated in 116. The specificity was assessed in 7482 controls representing the general population. The primary diagnostic endpoint was the area under the receiver operating characteristic curve in patients with positive SARS-CoV-2 PCR. The primary prognostic endpoint was the age-and sex-adjusted risk of A1HPV6 to predict patients with WHO-stage > 4 (W > 4) severity. We assessed the kinetics of the A1HPV6 components in a nonhuman primate model (NHP), from baseline to 7 days (D7) after SARS-CoV-2 infection. RESULTS: The area under the receiver operating characteristic curve for A1HPV6 was 0.99 (95% CI 0.97-0.99) in the validation subset, which was not significantly different from that in the construction subset, 0.99 (0.99-0.99; P = .80), like for sensitivity 92% (85-96) vs 94% (88-97; P = .29). A1HPV6 was associated with W > 4, with a significant odds ratio of 1.3 (1.1-1.5; 0.002). In NHP, A1 levels decreased (P < .01) at D2 and normalized at D4; HP levels increased at D2 and peaked at D4. In patients, A1 concentration was very low at D2 vs controls (P < .01) and increased at D14 (P < .01) but was still lower than controls; HP increased at D2 and remained elevated at D14. CONCLUSION: These results validate the diagnostic and prognostic performances of A1HPV6. Similar kinetics of apolipoprotein A1, HP, and alpha-2-macroglobulin were observed in the NHP model. ClinicalTrials.gov number, NCT01927133.

Immunization with synthetic SARS-CoV-2 S glycoprotein virus-like particles protects macaques from infection.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has caused an ongoing global health crisis. Here, we present as a vaccine candidate synthetic SARS-CoV-2 spike (S) glycoprotein-coated lipid vesicles that resemble virus-like particles. Soluble S glycoprotein trimer stabilization by formaldehyde cross-linking introduces two major inter-protomer cross-links that keep all receptor-binding domains in the "down" conformation. Immunization of cynomolgus macaques with S coated onto lipid vesicles (S-LVs) induces high antibody titers with potent neutralizing activity against the vaccine strain, Alpha, Beta, and Gamma variants as well as T helper (Th)1 CD4+-biased T cell responses. Although anti-receptor-binding domain (RBD)-specific antibody responses are initially predominant, the third immunization boosts significant non-RBD antibody titers. Challenging vaccinated animals with SARS-CoV-2 shows a complete protection through sterilizing immunity, which correlates with the presence of nasopharyngeal anti-S immunoglobulin G (IgG) and IgA titers. Thus, the S-LV approach is an efficient and safe vaccine candidate based on a proven classical approach for further development and clinical testing.

A third SARS-CoV-2 spike vaccination improves neutralization of variants-of-concern.

The emergence of SARS-CoV-2 variants that are more resistant to antibody-mediated neutralization pose a new hurdle in combating the COVID-19 pandemic. Although vaccines based on the original Wuhan sequence have been shown to be effective at preventing COVID-19, their efficacy is likely to be decreased against more neutralization-resistant variants-of-concern (VOC), in particular, the Beta variant originating in South Africa. We assessed, in mice, rabbits, and non-human primates, whether a third vaccination with experimental Wuhan-based Spike vaccines could alleviate this problem. Our data show that a third immunization improves neutralizing antibody titers against the variants-of-concern, Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), and Delta (B.1.617.2). After three vaccinations, the level of neutralization against Beta was similar to the level of neutralization against the original strain after two vaccinations, suggesting that simply providing a third immunization could nullify the reduced activity of current vaccines against VOC.

Cross-reactive antibodies after SARS-CoV-2 infection and vaccination.

Current SARS-CoV-2 vaccines are losing efficacy against emerging variants and may not protect against future novel coronavirus outbreaks, emphasizing the need for more broadly protective vaccines. To inform the development of a pan-coronavirus vaccine, we investigated the presence and specificity of cross-reactive antibodies against the spike (S) proteins of human coronaviruses (hCoV) after SARS-CoV-2 infection and vaccination. We found an 11- to 123-fold increase in antibodies binding to SARS-CoV and MERS-CoV as well as a 2- to 4-fold difference in antibodies binding to seasonal hCoVs in COVID-19 convalescent sera compared to pre-pandemic healthy donors, with the S2 subdomain of the S protein being the main target for cross-reactivity. In addition, we detected cross-reactive antibodies to all hCoV S proteins after SARS-CoV-2 vaccination in macaques and humans, with higher responses for hCoV more closely related to SARS-CoV-2. These findings support the feasibility of and provide guidance for development of a pan-coronavirus vaccine.

COVA1-18 neutralizing antibody protects against SARS-CoV-2 in three preclinical models.

Effective treatments against Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) are urgently needed. Monoclonal antibodies have shown promising results in patients. Here, we evaluate the in vivo prophylactic and therapeutic effect of COVA1-18, a neutralizing antibody highly potent against the B.1.1.7 isolate. In both prophylactic and therapeutic settings, SARS-CoV-2 remains undetectable in the lungs of treated hACE2 mice. Therapeutic treatment also causes a reduction in viral loads in the lungs of Syrian hamsters. When administered at 10 mg kg-1 one day prior to a high dose SARS-CoV-2 challenge in cynomolgus macaques, COVA1-18 shows very strong antiviral activity in the upper respiratory compartments. Using a mathematical model, we estimate that COVA1-18 reduces viral infectivity by more than 95% in these compartments, preventing lymphopenia and extensive lung lesions. Our findings demonstrate that COVA1-18 has a strong antiviral activity in three preclinical models and could be a valuable candidate for further clinical evaluation.

Targeting SARS-CoV-2 receptor-binding domain to cells expressing CD40 improves protection to infection in convalescent macaques.

Achieving sufficient worldwide vaccination coverage against SARS-CoV-2 will require additional approaches to currently approved viral vector and mRNA vaccines. Subunit vaccines may have distinct advantages when immunizing vulnerable individuals, children and pregnant women. Here, we present a new generation of subunit vaccines targeting viral antigens to CD40-expressing antigen-presenting cells. We demonstrate that targeting the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein to CD40 (αCD40.RBD) induces significant levels of specific T and B cells, with long-term memory phenotypes, in a humanized mouse model. Additionally, we demonstrate that a single dose of the αCD40.RBD vaccine, injected without adjuvant, is sufficient to boost a rapid increase in neutralizing antibodies in convalescent non-human primates (NHPs) exposed six months previously to SARS-CoV-2. Vaccine-elicited antibodies cross-neutralize different SARS-CoV-2 variants, including D614G, B1.1.7 and to a lesser extent B1.351. Such vaccination significantly improves protection against a new high-dose virulent challenge versus that in non-vaccinated convalescent animals.

An AAV-based, room-temperature-stable, single-dose COVID-19 vaccine provides durable immunogenicity and protection in non-human primates.

The SARS-CoV-2 pandemic has affected more than 185 million people worldwide resulting in over 4 million deaths. To contain the pandemic, there is a continued need for safe vaccines that provide durable protection at low and scalable doses and can be deployed easily. Here, AAVCOVID-1, an adeno-associated viral (AAV), spike-gene-based vaccine candidate demonstrates potent immunogenicity in mouse and non-human primates following a single injection and confers complete protection from SARS-CoV-2 challenge in macaques. Peak neutralizing antibody titers are sustained at 1 year and complemented by functional memory T cell responses. The AAVCOVID vector has no relevant pre-existing immunity in humans and does not elicit cross-reactivity to common AAVs used in gene therapy. Vector genome persistence and expression wanes following injection. The single low-dose requirement, high-yield manufacturability, and 1-month stability for storage at room temperature may make this technology well suited to support effective immunization campaigns for emerging pathogens on a global scale.

Predictive Markers of Immunogenicity and Efficacy for Human Vaccines.

Vaccines represent one of the major advances of modern medicine. Despite the many successes of vaccination, continuous efforts to design new vaccines are needed to fight "old" pandemics, such as tuberculosis and malaria, as well as emerging pathogens, such as Zika virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Vaccination aims at reaching sterilizing immunity, however assessing vaccine efficacy is still challenging and underscores the need for a better understanding of immune protective responses. Identifying reliable predictive markers of immunogenicity can help to select and develop promising vaccine candidates during early preclinical studies and can lead to improved, personalized, vaccination strategies. A systems biology approach is increasingly being adopted to address these major challenges using multiple high-dimensional technologies combined with in silico models. Although the goal is to develop predictive models of vaccine efficacy in humans, applying this approach to animal models empowers basic and translational vaccine research. In this review, we provide an overview of vaccine immune signatures in preclinical models, as well as in target human populations. We also discuss high-throughput technologies used to probe vaccine-induced responses, along with data analysis and computational methodologies applied to the predictive modeling of vaccine efficacy.

[COVID-19: Warp Speed vaccines]. / COVID-19, des vaccins à la vitesse de l'éclair.

A vaccine is required to effectively control the COVID-19 pandemic in the mid and long term. The development of vaccines against SARS-CoV-2 was initiated as soon as the genetic sequence of the virus was published, and has evolved at an unprecedented speed, with a first clinical trial launched in March 2020. One year later, more than a dozen of vaccines based on different concepts, with some having been evaluated only in clinical trials so far, are authorized under emergency procedures. Here, we review these vaccines, compare their properties and discuss the challenges they face, including the emergence of viral variants of concern.

TITLE: COVID-19, des vaccins à la vitesse de l'éclair. ABSTRACT: Un vaccin est nécessaire pour endiguer efficacement, à moyen et long terme, une pandémie comme celle de la COVID-19 (coronavirus disease 2019). Le développement de vaccins contre le virus responsable de la maladie, le SARS-CoV-2 (severe acute respiratory syndrome-coronavirus 2), a été débuté dès la publication de la séquence du génome viral. Ce développement a progressé à une vitesse sans précédent, avec un premier essai clinique réalisé peu de temps après, en mars 2020. Un an plus tard, une dizaine de vaccins reposant sur des concepts différents, dont certains n'avaient été testés que dans des essais cliniques, sont autorisés dans le cadre de procédures d'urgence. Dans cet article, nous passons en revue ces différents vaccins, nous comparons leurs propriétés et nous discutons les défis auxquels ils sont confrontés, en particulier l'émergence de nouveaux variants viraux.

Non-human primate models of human respiratory infections.

Respiratory pathogens represent a great burden for humanity and a potential source of new pandemics, as illustrated by the recent emergence of coronavirus disease 2019 (COVID-19). In recent decades, biotechnological advances have led to the development of numerous innovative therapeutic molecules and vaccine immunogens. However, we still lack effective treatments and vaccines against many respiratory pathogens. More than ever, there is a need for a fast, predictive, preclinical pipeline, to keep pace with emerging diseases. Animal models are key for the preclinical development of disease management strategies. The predictive value of these models depends on their ability to reproduce the features of the human disease, the mode of transmission of the infectious agent and the availability of technologies for monitoring infection. This review focuses on the use of non-human primates as relevant preclinical models for the development of prevention and treatment for human respiratory infections.

SARS-CoV-2 viral dynamics in non-human primates.

Non-human primates infected with SARS-CoV-2 exhibit mild clinical signs. Here we used a mathematical model to characterize in detail the viral dynamics in 31 cynomolgus macaques for which nasopharyngeal and tracheal viral load were frequently assessed. We identified that infected cells had a large burst size (>104 virus) and a within-host reproductive basic number of approximately 6 and 4 in nasopharyngeal and tracheal compartment, respectively. After peak viral load, infected cells were rapidly lost with a half-life of 9 hours, with no significant association between cytokine elevation and clearance, leading to a median time to viral clearance of 10 days, consistent with observations in mild human infections. Given these parameter estimates, we predict that a prophylactic treatment blocking 90% of viral production or viral infection could prevent viral growth. In conclusion, our results provide estimates of SARS-CoV-2 viral kinetic parameters in an experimental model of mild infection and they provide means to assess the efficacy of future antiviral treatments.