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.

Cryo-EM structures and binding of mouse ACE2 to SARS-CoV-2 variants of concern (preprint)

Investigation of potential hosts of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is crucial to understanding future risks of spillover and spillback. SARS-CoV-2 has been reported to be transmitted from humans to various animals after requiring relatively few mutations. Mice are well adapted to human environments, frequently come in contact with humans, are used widely as infection models, and may act as reservoirs for SARS-CoV-2. Structural and binding data of the mouse ACE2 receptor with the Spike protein of newly identified SARS-CoV-2 variants are needed to better understand the impact of variants of concern (VOC). Previous studies have developed mouse-adapted variants and have identified some determinants of binding. Here we report the cryo-EM structures of mouse ACE2 bound to Spike ectodomains of four different VOC: Beta, Omicron BA.1, Omicron BA.2.12.1 and Omicron BA.4/5. These variants represent the oldest to the newest variants that are able to bind the mouse ACE2 receptor. Our high-resolution structural data complemented with bio-layer interferometry (BLI) binding assays reveal a requirement for a combination of mutations in the Spike protein to enable the binding to mouse ACE2.

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.

Structural analysis of the Spike of the Omicron SARS-COV-2 variant by cryo-EM and implications for immune evasion (preprint)

ABSTRACT The Omicron (B.1.1.529) SARS-COV-2 was reported on November 24, 2021 and declared a variant of concern a couple of days later. 1,2 With its constellation of mutations acquired by this variant on its Spike glycoprotein and the speed at which this new variant has replaced the previously dominant variant Delta in South Africa and the United Kingdom, it is crucial to have atomic structural insights to reveal the mechanism of its rapid proliferation. Here we present a high-resolution cryo-EM structure of the Spike protein of the Omicron variant.

Riding the Wave: Maintaining Safe Lung Cancer Surgery During the COVID-19 Pandemic in a Global City (preprint)

Objectives: SARS-CoV-2 has challenged health service provision worldwide. This work evaluates safe surgical pathways and standard operating procedures implemented in the high volume, global city of London during the first wave of SARS-CoV-2 infection. We also assess the safety of minimally invasive surgery(MIS) for anatomical lung resection. Methods: This multicentre cohort study was conducted across all London thoracic surgical units, covering a catchment area of approximately 14.8 Million. A Pan-London Collaborative was created for data sharing and dissemination of protocols. All patients undergoing anatomical lung resection 1st March-1st June 2020 were included. Primary outcomes were SARS-CoV-2 infection, access to minimally invasive surgery, post-operative complication, length of intensive care and hospital stay (LOS), and death during follow up. Results: 352 patients underwent anatomical lung resection with an median age of 69 (IQR: 35-86) years. Self-isolation and pre-operative screening were implemented following the UK national lockdown. Pre-operative SARS-CoV-2 swabs were performed in 63.1% and CT imaging in 54.8%. 61.7% of cases were performed minimally invasively. Median LOS was 6 days. Significant complications developed in 7.3% of patients (Clavien-Dindo Grade 3-4). There were 6 deaths(1.7%) and 12 re-admissions(3.4%). Seven patients(2.0%) were diagnosed with SARS-CoV-2 infection, two of whom died (28.5%). Conclusions: Major elective surgery can be safely undertaken via open and MIS approaches at the peak of a viral pandemic if precautionary measures are implemented. High volume surgery should continue during further viral peaks to minimise health service burden and potential harm to cancer patients.Funding Statement: Nil.Declaration of Interests: None of the authors have conflict of interests to declare.Ethics Approval Statement: Institutional approval was obtained from all Trusts contributing to the collaborative. Approval was granted by each hospital trust for data sharing, collaborative work and retrospective review. Formal individual informed consent was not required due to the retrospective nature of the study.

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).

Structural investigation of ACE2 dependent disassembly of the trimeric SARS-CoV-2 Spike glycoprotein (preprint)

The human membrane protein Angiotensin-converting enzyme 2 (hACE2) acts as the main receptor for host cells invasion of the new coronavirus SARS-CoV-2. The viral surface glycoprotein Spike binds to hACE2, which triggers virus entry into cells. As of today, the role of hACE2 for virus fusion is not well understood. Blocking the transition of Spike from its prefusion to post-fusion state might be a strategy to prevent or treat COVID-19. Here we report a single particle cryo-electron microscopy analysis of SARS-CoV-2 trimeric Spike in presence of the human ACE2 ectodomain. The binding of purified hACE2 ectodomain to Spike induces the disassembly of the trimeric form of Spike and a structural rearrangement of its S1 domain to form a stable, monomeric complex with hACE2. This observed hACE2 dependent dissociation of the Spike trimer suggests a mechanism for the therapeutic role of recombinant soluble hACE2 for treatment of COVID-19.

Genomic Similarity of Nucleotides in SARS CoronaVirus using K-Means Unsupervised Learning Algorithm (preprint)

The drastic increase in the number of coronaviruses discovered and coronavirus genomes being sequenced have given us a great opportunity to perform genomics and bioinformatics analysis on this family of viruses. Coronaviruses possess the largest genomes (26.4 to 31.7 kb) among all known RNA viruses, with G + C contents varying from 32% to 43%. Phylogenetically, three genera, Alphacoronavirus, Betacoronavirus and Gammacoronavirus, with Betacoronavirus consisting of subgroups A, B, C were known to exist but now a new genus D also exists,namely the Deltacoronavirus. In such a situation, it becomes highly important for efficient classification of all virus data so that it helps us in suitable planning,containment and treatment. The objective of this paper is to classify SARS corona-virus nucleotide sequences based on parameters such as sequence length, percentage similarity between the sequence information,open and closed gaps in the sequence due to multiple mutations and many others. By doing this, we will be able to predict accurately the similarity of SARS CoV-2 virus with respect to other corona-viruses like the Wuhan corona-virus, the bat corona-virus and the pneumonia virus and would help us better understand about the taxonomy of the corona-virus family.

Immune response to vaccine candidates based on different types of nanoscaffolded RBD domain of the SARS-CoV-2 spike protein (preprint)

Effective and safe vaccines against SARS-CoV-2 are highly desirable to prevent casualties and societal cost caused by Covid-19 pandemic. The receptor binding domain (RBD) of the surface-exposed spike protein of SARS-CoV-2 represents a suitable target for the induction of neutralizing antibodies upon vaccination. Small protein antigens typically induce weak immune response while particles measuring tens of nanometers are efficiently presented to B cell follicles and subsequently to follicular germinal center B cells in draining lymph nodes, where B cell proliferation and affinity maturation occurs. Here we prepared and analyzed the response to several DNA vaccines based on genetic fusions of RBD to four different scaffolding domains, namely to the foldon peptide, ferritin, lumazine synthase and {beta}-annulus peptide, presenting from 6 to 60 copies of the RBD on each particle. Scaffolding strongly augmented the immune response with production of neutralizing antibodies and T cell response including cytotoxic lymphocytes in mice upon immunization with DNA plasmids. The most potent response was observed for the 24-residue {beta}-annulus peptide scaffold that forms large soluble assemblies, that has the advantage of low immunogenicity in comparison to larger scaffolds. Our results support the advancement of this vaccine platform towards clinical trials.

Qualitative Changes in the SARS-CoV-2 Antibody Response in the Post-Infection Phase Impact the estimates of infections in Population-Based Seroprevalence Studies (preprint)

We have determined SARS-CoV-2-specific antibody responses in a cohort of 96 individuals with acute infection and in 578 individuals enrolled in a seroprevalence population study in Switzerland including three groups, i.e. subjects with previous RT-PCR confirmed SARS-CoV-2 infections (n=90), positive patient contacts (n=177) and random selected subjects (n=311). SARS-CoV-2 antibody responses specific to the Spike (S), in the monomeric and native trimeric forms, and/or the nucleocapsid (N) proteins were equally sensitive in the acute infection phase. Interestingly, as compared to anti-S antibody responses, those against the N protein appear to wane in the post-infection and substantially underestimated the proportion of SARS-CoV-2 infections in the groups of patient positive contacts, i.e. 10.9 to 32.2% reduction and in the random selected general population, i.e. up to 45% reduction. The overall reduction in seroprevalence targeting only anti-N IgG antibodies for the total cohort ranged from 9.4 to 31%. Of note, the use of the S protein in its native trimer form was more sensitive as compared to monomeric S proteins. These results indicate that the assessment of anti-S IgG antibody responses against the native trimeric S protein should be implemented to estimate SARS-CoV-2 infections in population-based seroprevalence studies.

Population-wide evolution of SARS-CoV-2 immunity tracked by a ternary immunoassay (preprint)

We describe the evolution of severe acute respiratory coronavirus 2 (SARS-CoV-2) seroprevalence in the greater area of Zurich, Switzerland, a region that has been only mildly hit by the pandemic in spite of hosting an international airport hub and a highly mobile population. Seroprevalence studies in low-prevalence settings require large sample sizes and high-specificity methodologies. To address this particular challenge, we developed a Tripartite Automated Blood Immunoassay to assess the IgG response against three SARS-CoV-2 proteins on approximately 40'000 samples from university hospital patients and healthy blood donors. The seroprevalence increased in March 2020 (0.3%; CI95%: 0.1% - 0.5%) in the cohort of the hospital patients but rapidly plateaued in April at 1.1-1.3%, with a slight drop in June, then decreased in July to 0.3-0.7%. Seropositive samples were confirmed with Western Blotting and liquid-phase binding assays. Employing a dynamic transmission model that describes SARS-CoV-2 transmission and seroconversion in the general population of the Canton of Zurich, we estimated an infection fatality ratio of 0.6% (CI95%: 0.4%-0.8%). We conclude that a small proportion of the population in the greater area of Zurich has been exposed to SARS-CoV-2, with an IFR that is similar to that of other European areas. The evolution of seroprevalence points to a high effectiveness of containment measures and/or to rapid loss of humoral responses.