ACE2 mimetic antibody potently neutralizes all SARS-CoV-2 variants and fully protects in XBB.1.5 challenged monkeys (preprint)

The rapid evolution of SARS-CoV-2 to variants with improved transmission efficiency and reduced sensitivity to vaccine-induced humoral immunity has abolished the protective effect of licensed therapeutic human monoclonal antibodies (mAbs). To fill this unmet medical need and protect vulnerable patient populations, we isolated the P4J15 mAb from a previously infected, vaccinated donor, with <20 ng/ml neutralizing activity against all Omicron variants including the latest XBB.2.3 and EG.1 sub-lineages. Structural studies of P4J15 in complex with Omicron XBB.1 Spike show that the P4J15 epitope shares ~93% of its buried surface area with the ACE2 contact region, consistent with an ACE2 mimetic antibody. Although SARS-CoV-2 mutants escaping neutralization by P4J15 were selected in vitro, these displayed lower infectivity, poor binding to ACE2, and the corresponding "escape" mutations are accordingly rare in public sequence databases. Using a SARS-CoV-2 XBB.1.5 monkey challenge model, we show that P4J15 confers complete prophylactic protection. We conclude that the P4J15 mAb has potential as a broad-spectrum anti-SARS-CoV-2 drug.

SARS-CoV-2 Omicron potently neutralized by a novel antibody with unique Spike binding properties (preprint)

The SARS-CoV-2 Omicron variant exhibits very high levels of transmission, pronounced resistance to authorized therapeutic human monoclonal antibodies and reduced sensitivity to vaccine-induced immunity. Here we describe P2G3, a human monoclonal antibody (mAb) isolated from a previously infected and vaccinated donor, which displays picomolar-range neutralizing activity against Omicron BA.1, BA.1.1, BA.2 and all other current variants, and is thus markedly more potent than all authorized or clinically advanced anti-SARS-CoV-2 mAbs. Structural characterization of P2G3 Fab in complex with the Omicron Spike demonstrates unique binding properties to both down and up spike trimer conformations at an epitope that partially overlaps with the receptor-binding domain (RBD), yet is distinct from those bound by all other characterized mAbs. This distinct epitope and angle of attack allows P2G3 to overcome all the Omicron mutations abolishing or impairing neutralization by other anti-SARS-COV-2 mAbs, and P2G3 accordingly confers complete prophylactic protection in the SARS-CoV-2 Omicron monkey challenge model. Finally, although we could isolate in vitro SARS-CoV2 mutants escaping neutralization by P2G3 or by P5C3, a previously described broadly active Class 1 mAb, we found these viruses to be lowly infectious and their key mutations extremely rare in the wild, and we could demonstrate that P2G3/P5C3 efficiently cross-neutralized one another's escapees. We conclude that this combination of mAbs has great prospects in both the prophylactic and therapeutic settings to protect from Omicron and other VOCs.

Design, immunogenicity, and efficacy of a pan-sarbecovirus dendritic-cell targeting vaccine (preprint)

The emergence of SARS-CoV-2 variants of concern (VOCs) that escape pre-existing antibody neutralizing responses increases the need for vaccines that target conserved epitopes and induce cross-reactive B- and T-cell responses. We used a computational approach and sequence alignment analysis to design a new-generation subunit vaccine targeting conserved sarbecovirus B- and T-cell epitopes from Spike (S) and Nucleocapsid (N) to antigen-presenting cells expressing CD40 (CD40.CoV2). We demonstrate the potency of CD40.CoV2 to elicit high levels of cross-neutralizing antibodies against SARS-CoV-2, VOCs, and SARS-CoV-1 in K18-hACE2 transgenic mice, associated with improved viral control and survival after challenge. In addition, we demonstrate the potency of CD40.CoV2 in vitro to recall human multi-epitope, functional, and cytotoxic SARS-CoV-2 S- and N-specific T-cell responses that are unaffected by VOC mutations and cross-reactive with SARS-CoV-1 and, to a lesser extent, MERS epitopes. Overall, these findings provide a framework for a pan-sarbecovirus vaccine. Keywords: COVID-19, SARS-CoV-2, vaccine, pre-clinical model, sarbecoviruses

A Highly Potent Antibody Effective Against SARS-CoV-2 Variants of Concern (preprint)

Control of the ongoing SARS-CoV-2 pandemic is endangered by the emergence of viral variants with increased transmission efficiency, resistance to marketed therapeutic antibodies and reduced sensitivity to vaccine-induced immunity. Here, we screened B cells from COVID-19 donors and identified P5C3, a highly potent and broadly neutralizing monoclonal antibody with picomolar neutralizing activity against all SARS-CoV-2 variants of concern (VOC) identified to date. Structural characterization of P5C3 Fab in complex with the Spike demonstrates a neutralizing activity defined by a large buried surface area, highly overlapping with the receptor-binding domain (RBD) surface necessary for ACE2 interaction. We further demonstrate that P5C3 showed complete prophylactic protection in the SARS-CoV-2 infected hamster challenge model. These results indicate that P5C3 opens exciting perspectives either as a prophylactic agent in immunocompromised individuals with poor response to vaccination or as combination therapy in SARS-CoV-2-infected individuals.Funding: This CARE project has received funding from the Innovative MedicinesInitiative 2 Joint Undertaking (JU) under grant agreement No 101005077. The JU receives support from the European Union’s Horizon 2020 research and innovation program and EFPIA and BILL & MELINDA GATES FOUNDATION, GLOBAL HEALTH DRUG DISCOVERYINSTITUTE, UNIVERSITY OF DUNDEE. Furthermore, funding was also provided through the Lausanne University Hospital, through the Swiss Vaccine Research Institute to G.P., and through the EPFL COVID fund to D.T.Conflict of Interest: None to declare. Ethical Approval: Study design and use of subject samples were approved by the Institutional Review Board of the Lausanne University Hospital and the ‘Commission d’éthique du Canton de Vaud’ (CER-VD).

HGF and CXCL13, Two Antagonizing Cytokines in Lung Inflammation and Fibrosis, Predict the Severity and the Mortality of COVID-19 (preprint)

The objective of the present study was to identify biological signatures of severe COVID-19 predictive of admission in the intensive care unit (ICU). Over 170 immunological markers were investigated in a ‘discovery’ cohort (n=98 patients) of the Lausanne University Hospital (LUH-1). While cellular immunological markers lacked power in discriminating between ICU and non-ICU patients, 13 out of 49 cytokines were significantly associated with ICU admission in the three cohorts (P<0.05 to P<0.001). The cytokine results were confirmed in two ‘validation’ cohorts, i.e. the French COVID-19 Study (FCS; n=62) and a second LUH-2 cohort (n=47). Of note, HGF is a pleiotropic cytokine with anti-inflammatory properties playing a fundamental role in lung tissue repair, and CXCL13, a pro-inflammatory chemokine associated with pulmonary fibrosis and regulating the maturation of B cell response. The two cytokines in combination were the best predictors of ICU admission (positive and negative predictive values ranging from 81.8% to 93.1% and 85.2% to 94.4% in the 3 cohorts) and occurrence of death during patient follow-up (8.8 fold higher likelihood of death when both cytokines were increased). Up-regulation of HGF reflects the most powerful counter-regulatory mechanism of the host immune response to antagonize the pro-inflammatory cytokines including CXCL13 and to prevent lung fibrosis in COVID-19 patients.

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

Controlling the circulation of the recently emerged SARS-CoV-2 in the human populations requires massive vaccination campaigns. Achieving sufficient worldwide vaccination coverage will require additional approaches to first generation of approved viral vector and mRNA vaccines. Subunit vaccines have excellent safety and efficacy records and may have distinct advantages, in particular when immunizing individuals with vulnerabilities or when considering the vaccination of children and pregnant women.. We have developed a new generation of subunit vaccines with enhanced immunogenicity by the targeting of viral antigens to CD40-expressing antigen-presenting cells, thus harnessing their intrinsic immune-stimulant properties. Here, 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 a long-term memory phenotype, in a humanized mouse model. In addition, 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. Such vaccination thus significantly improved protection against a new high-dose virulent challenge versus that in non-vaccinated convalescent animals. Viral dynamics modelling showed the high efficiency of the vaccine at controlling the viral dissemination.

CD177, a specific marker of neutrophil activation, is a hallmark of COVID-19 severity and death (preprint)

COVID-19 SARS-CoV-2 infection exhibits wide inter-individual clinical variability, from silent infection to severe disease and death. The identification of high-risk patients is a continuing challenge in routine care. We aimed to identify factors that influence clinical worsening. We analyzed 52 cell populations, 71 analytes, and RNA-seq gene expression in the blood of severe patients from the French COVID cohort upon hospitalization (n = 61). COVID-19 patients showed severe abnormalities of 27 cell populations relative to healthy donors (HDs). Forty-two cytokines, neutrophil chemo-attractants, and inflammatory components were elevated in COVID-19 patients. Supervised gene expression analyses showed differential expression of genes for neutrophil activation, interferon signaling, T- and B-cell receptors, EIF2 signaling, and ICOS-ICOSL pathways in COVID-19 patients. Unsupervised analysis confirmed the prominent role of neutrophil activation, with a high abundance of CD177, a specific neutrophil activation marker. CD177 was the most highly differentially-expressed gene contributing to the clustering of severe patients and its abundance correlated with CD177 protein serum levels. CD177 levels were higher in COVID-19 patients from both the French and "confirmatory" Swiss cohort (n = 203) than in HDs (P< 0.01) and in ICU than non-ICU patients (P< 0.001), correlating with the time to symptoms onset (P = 0.002). Longitudinal measurements showed sustained levels of serum CD177 to discriminate between patients with the worst prognosis, leading to death, and those who recovered (P = 0.01). These results highlight neutrophil activation as a hallmark of severe disease and CD177 assessment as a reliable prognostic marker for routine care.

Asymptomatic and symptomatic SARS-CoV-2 infections elicit polyfunctional antibodies. (preprint)

A large proportion of SARS-CoV-2 infected individuals remains asymptomatic. Little is known about the extent and quality of their antiviral humoral response. Here, we analyzed antibody functions in 52 asymptomatic infected individuals, 119 mild and 21 hospitalized COVID-19 patients. We measured anti-Spike antibody levels with the S-Flow assay and mapped SARS-CoV-2 Spike- and N-targeted regions by Luminex. Neutralization, complement deposition and Antibody-Dependent Cellular Cytotoxicity (ADCC) were evaluated using replication-competent SARS-CoV-2 or reporter cell systems. We show that COVID-19 sera mediate complement deposition and kill infected cells by ADCC. Sera from asymptomatic individuals neutralize the virus, activate ADCC and trigger complement deposition. Antibody levels and activities are slightly lower in asymptomatic individuals. The different functions of the antibodies are correlated, independently of disease severity. Longitudinal samplings show that antibody functions follow similar kinetics of induction and contraction, with minor variations. Overall, asymptomatic SARS-CoV-2 infection elicits polyfunctional antibodies neutralizing the virus and targeting infected cells. - Sera from convalescent COVID-19 patients activate the complement and kill infected cells by ADCC. - Asymptomatic and symptomatic SARS-CoV-2-infected individuals harbor polyfunctional antibodies. - Antibody levels and functions are slightly lower in asymptomatic individuals - The different antiviral activities of anti-Spike antibodies are correlated regardless of disease severity. - Functions of anti-Spike antibodies have similar kinetics of induction and contraction.

Immune alterations during SARS-CoV-2-related acute respiratory distress syndrome (preprint)

We report a longitudinal analysis of the immune response associated with a fatal case of COVID-19. This patient exhibited a rapid evolution towards multiorgan failure. SARS-CoV-2 was detected in multiple nasopharyngeal, blood, and pleural samples, despite antiviral and immunomodulator treatment. Clinical evolution in the blood was marked by an increase (2-3 fold) in differentiated effector T cells expressing exhaustion (PD-1) and senescence (CD57) markers, an expansion of antibody-secreting cells, a 15-fold increase in {gamma}{delta} T-cell and proliferating NK-cell populations, and the total disappearance of monocytes, suggesting lung trafficking. In the serum, waves of a proinflammatory cytokine storm, Th1 and Th2 activation, and markers of T-cell exhaustion, apoptosis, cell cytotoxicity, and endothelial activation were observed until the fatal outcome. This case underscores the need for well-designed studies to investigate complementary approaches to control viral replication, the source of the hyperinflammatory status, and immunomodulation to target the pathophysiological response.