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Rapidly evolving influenza A viruses (IAVs) and influenza B viruses (IBVs) are major causes of recurrent lower respiratory tract infections. Current influenza vaccines elicit antibodies predominantly to the highly variable head region of haemagglutinin and their effectiveness is limited by viral drift1 and suboptimal immune responses2. Here we describe a neuraminidase-targeting monoclonal antibody, FNI9, that potently inhibits the enzymatic activity of all group 1 and group 2 IAVs, as well as Victoria/2/87-like, Yamagata/16/88-like and ancestral IBVs. FNI9 broadly neutralizes seasonal IAVs and IBVs, including the immune-evading H3N2 strains bearing an N-glycan at position 245, and shows synergistic activity when combined with anti-haemagglutinin stem-directed antibodies. Structural analysis reveals that D107 in the FNI9 heavy chain complementarity-determinant region 3 mimics the interaction of the sialic acid carboxyl group with the three highly conserved arginine residues (R118, R292 and R371) of the neuraminidase catalytic site. FNI9 demonstrates potent prophylactic activity against lethal IAV and IBV infections in mice. The unprecedented breadth and potency of the FNI9 monoclonal antibody supports its development for the prevention of influenza illness by seasonal and pandemic viruses.

Background Sotrovimab (VIR-7831) is an engineered human monoclonal antibody targeting a conserved region of the SARS-CoV-2 spike protein; it has been shown to have a favorable safety profile and be effective for early treatment of high-risk COVID-19 patients. The COMET-TAIL phase 3 study evaluated sotrovimab administered intravenously (IV) or intramuscularly (IM) for the treatment of participants with mild to moderate COVID-19 who are at high risk of disease progression. Methods Between June to August 2021, 973 participants were randomized and received sotrovimab by 500 mg IV infusion or by 500 or 250 mg IM injection. Deep sequencing of the spike gene was performed on nasopharyngeal samples. Baseline (BL; Day 1 or Day 3), post-BL (Day 5 or later), treatment-emergent (TE) substitutions at sotrovimab epitope positions, and presence of variants of concern/interest (VOC/VOI), were evaluated at a ≥5% allelic frequency. Phenotypic analyses were conducted using a pseudotyped virus assay. Results Sequences were available from 764 participants (500 mg IV: 314/393; 500 mg IM: 302/387; 250 mg IM: 148/193). Consistent with VOC circulation during enrollment, the Delta variant was detected in 88.2% (674/764) of participants. Alpha and Mu variants were also seen at >2% prevalence. Of the 764 participants, 26 met the primary endpoint for clinical progression to hospitalization >24 hours or death due to any cause through day 29 and were infected with Delta (500 mg IV: 4; 500 mg IM: 9; 250 mg IM: 11), Alpha (500 mg IM: 1), or Mu (500 mg IV: 1) variants. Substitutions at sotrovimab epitope positions were similar across arms and were detected in 82/764 (10.7%) participants at any visit (500 mg IV: 42/314; 500 mg IM: 27/302; 250 mg IM: 13/148). Of these, 2 participants experienced clinical progression: 1 participant infected with the Mu variant (500 mg IV) carried the characteristic R346K substitution at BL; 1 participant infected with the Delta variant (500 mg IM) had P337L and E340K substitutions detected at Day 3 and P337L was enriched at Day 8. The predominant TE epitope substitutions included P337L and E340A/K/V, which confer reduced susceptibility to sotrovimab in vitro. Conclusion Overall, TE epitope substitutions were not associated with clinical progression. Funding Vir & GSK (NCT04913675) Disclosures Maria L. Agostini, PhD, Vir Biotechnology, Inc: Employee|Vir Biotechnology, Inc: Stocks/Bonds Gretja Schnell, PhD, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds Julia di Iulio, PharmD, PhD, MBA, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds Anita Kohli, MD, GlaxoSmithKline: Third party funding to Vir|Vir Biotechnology Inc: Support as a trial site paid to my institution and non-financial support for medical writing support Adrienne E. Shapiro, MD, PhD, Vir Biotechnology: Support as a trial site paid to my institution and non-financial support for medical writing GlaxoSmithKline third party funding to Vir support Elias H. Sarkis, MD, Abbvie: Grant/Research Support|Abbvie: Speakers Bureau|Eisai: Grant/Research Support|GlaxoSmithKline: Third party funding to Vir|Ironshore: Grant/Research Support|Janssen: Speakers Bureau|Lilly: Grant/Research Support|Otsuka: Grant/Research Support|Teva: Speakers Bureau|Vir Biotechnology Inc: Support as a trial site paid to my institution and non-financial support for medical writing support Dave Inman, MSc, GlaxoSmithKline: Employee|GlaxoSmithKline: Stocks/Bonds Amanda Peppercorn, MD, GlaxoSmithKline: Employee|GlaxoSmithKline: Stocks/Bonds Andrew Skingsley, MD, GlaxoSmithKline: Employee|GlaxoSmithKline: Stocks/Bonds Leah A. Gaffney, PhD, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds Melissa Aldinger, PharmD, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds Christy M. Hebner, PhD, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds Andrea L. Cathcart, PhD, Vir Biotechnology: Employee|Vir Biotechnology: Stocks/Bonds.

During the COVID-19 pandemic, SARS-CoV-2 surveillance efforts integrated genome sequencing of clinical samples to identify emergent viral variants and to support rapid experimental examination of genome-informed vaccine and therapeutic designs. Given the broad range of methods applied to generate new viral genomes, it is critical that consensus and variant calling tools yield consistent results across disparate pipelines. Here we examine the impact of sequencing technologies (Illumina and Oxford Nanopore) and 7 different downstream bioinformatic protocols on SARS-CoV-2 variant calling as part of the NIH Accelerating COVID-19 Therapeutic Interventions and Vaccines (ACTIV) Tracking Resistance and Coronavirus Evolution (TRACE) initiative, a public-private partnership established to address the COVID-19 outbreak. Our results indicate that bioinformatic workflows can yield consensus genomes with different single nucleotide polymorphisms, insertions, and/or deletions even when using the same raw sequence input datasets. We introduce the use of a specific suite of parameters and protocols that greatly improves the agreement among pipelines developed by diverse organizations. Such consistency among bioinformatic pipelines is fundamental to SARS-CoV-2 and future pathogen surveillance efforts. The application of analysis standards is necessary to more accurately document phylogenomic trends and support data-driven public health responses.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron sublineages carry distinct spike mutations resulting in escape from antibodies induced by previous infection or vaccination. We show that hybrid immunity or vaccine boosters elicit plasma-neutralizing antibodies against Omicron BA.1, BA.2, BA.2.12.1, and BA.4/5, and that breakthrough infections, but not vaccination alone, induce neutralizing antibodies in the nasal mucosa. Consistent with immunological imprinting, most antibodies derived from memory B cells or plasma cells of Omicron breakthrough cases cross-react with the Wuhan-Hu-1, BA.1, BA.2, and BA.4/5 receptor-binding domains, whereas Omicron primary infections elicit B cells of narrow specificity up to 6 months after infection. Although most clinical antibodies have reduced neutralization of Omicron, we identified an ultrapotent pan-variant-neutralizing antibody that is a strong candidate for clinical development. ispartof: SCIENCE vol:378 issue:6620 pages:619-+ ispartof: location:United States status: published

SARS-CoV-2 from alpha to epsilon As battles to contain the COVID-19 pandemic continue, attention is focused on emerging variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus that have been deemed variants of concern because they are resistant to antibodies elicited by infection or vaccination or they increase transmissibility or disease severity. Three papers used functional and structural studies to explore how mutations in the viral spike protein affect its ability to infect host cells and to evade host immunity. Gobeil et al . looked at a variant spike protein involved in transmission between minks and humans, as well as the B1.1.7 (alpha), B.1.351 (beta), and P1 (gamma) spike variants; Cai et al . focused on the alpha and beta variants; and McCallum et al . discuss the properties of the spike protein from the B1.1.427/B.1.429 (epsilon) variant. Together, these papers show a balance among mutations that enhance stability, those that increase binding to the human receptor ACE2, and those that confer resistance to neutralizing antibodies. —VV

mRNA levels are determined by the balance between mRNA synthesis and decay. Protein factors that mediate both processes, including the 5'-3' exonuclease Xrn1, are responsible for a cross-talk between the two processes that buffers steady-state mRNA levels. However, the roles of these proteins in transcription remain elusive and controversial. Applying native elongating transcript sequencing (NET-seq) to yeast cells, we show that Xrn1 functions mainly as a transcriptional activator and that its disruption manifests as a reduction of RNA polymerase II (Pol II) occupancy downstream of transcription start sites. By combining our sequencing data and mathematical modeling of transcription, we found that Xrn1 modulates transcription initiation and elongation of its target genes. Furthermore, Pol II occupancy markedly increased near cleavage and polyadenylation sites in xrn1Δ cells, whereas its activity decreased, a characteristic feature of backtracked Pol II. We also provide indirect evidence that Xrn1 is involved in transcription termination downstream of polyadenylation sites. We noted that two additional decay factors, Dhh1 and Lsm1, seem to function similarly to Xrn1 in transcription, perhaps as a complex, and that the decay factors Ccr4 and Rpb4 also perturb transcription in other ways. Interestingly, the decay factors could differentiate between SAGA- and TFIID-dominated promoters. These two classes of genes responded differently to XRN1 deletion in mRNA synthesis and were differentially regulated by mRNA decay pathways, raising the possibility that one distinction between these two gene classes lies in the mechanisms that balance mRNA synthesis with mRNA decay.

SARS-CoV-2 evolution threatens vaccine- and natural infection-derived immunity as well as the efficacy of therapeutic antibodies. To improve public health preparedness, we sought to predict which existing amino acid mutations in SARS-CoV-2 might contribute to future variants of concern. We tested the predictive value of features comprising epidemiology, evolution, immunology, and neural network-based protein sequence modeling, and identified primary biological drivers of SARS-CoV-2 intra-pandemic evolution. We found evidence that ACE2-mediated transmissibility and resistance to population-level host immunity has waxed and waned as a primary driver of SARS-CoV-2 evolution over time. We retroactively identified with high accuracy (area under the receiver operator characteristic curve, AUROC=0.92-0.97) mutations that will spread, at up to four months in advance, across different phases of the pandemic. The behavior of the model was consistent with a plausible causal structure wherein epidemiological covariates combine the effects of diverse and shifting drivers of viral fitness. We applied our model to forecast mutations that will spread in the future and characterize how these mutations affect the binding of therapeutic antibodies. These findings demonstrate that it is possible to forecast the driver mutations that could appear in emerging SARS-CoV-2 variants of concern. We validate this result against Omicron, showing elevated predictive scores for its component mutations prior to emergence, and rapid score increase across daily forecasts during emergence. This modeling approach may be applied to any rapidly evolving pathogens with sufficiently dense genomic surveillance data, such as influenza, and unknown future pandemic viruses.

Understanding who is at risk of progression to severe COVID-19 is key to effective treatment. We studied correlates of disease severity in the COMET-ICE clinical trial that randomized 1:1 to placebo or to sotrovimab, a monoclonal antibody for the treatment of SARS-CoV-2 infection. Several laboratory parameters identified study participants at greater risk of severe disease, including a high neutrophil-lymphocyte ratio (NLR), a negative SARS-CoV-2 serologic test and whole blood transcriptome profiles. Sotrovimab treatment in these groups was associated with normalization of NLR and the transcriptomic profile, and with a decrease of viral RNA in nasopharyngeal samples. Transcriptomics provided the most sensitive detection of participants who would go on to be hospitalized or die. To facilitate timely measurement, we identified a 10-gene signature with similar predictive accuracy. In summary, we identified markers of risk for disease progression and demonstrated that normalization of these parameters occurs with antibody treatment of established infection.

SUMMARYThe recently emerged SARS-CoV-2 Omicron variant harbors 37 amino acid substitutions in the spike (S) protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody therapeutics. Here, we show that the Omicron RBD binds to human ACE2 with enhanced affinity relative to the Wuhan-Hu-1 RBD and acquires binding to mouse ACE2. Severe reductions of plasma neutralizing activity were observed against Omicron compared to the ancestral pseudovirus for vaccinated and convalescent individuals. Most (26 out of 29) receptor-binding motif (RBM)-directed monoclonal antibodies (mAbs) lost in vitro neutralizing activity against Omicron, with only three mAbs, including the ACE2-mimicking S2K146 mAb1, retaining unaltered potency. Furthermore, a fraction of broadly neutralizing sarbecovirus mAbs recognizing antigenic sites outside the RBM, including sotrovimab2, S2X2593 and S2H974, neutralized Omicron. The magnitude of Omicron-mediated immune evasion and the acquisition of binding to mouse ACE2 mark a major SARS-CoV-2 mutational shift. Broadly neutralizing sarbecovirus mAbs recognizing epitopes conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers.

A gene is considered essential if loss of function results in loss of viability, fitness or in disease. This concept is well established for coding genes; however, non-coding regions are thought less likely to be determinants of critical functions. Here we train a machine learning model using functional, mutational and structural features, including new genome essentiality metrics, 3D genome organization and enhancer reporter data to identify deleterious variants in non-coding regions. We assess the model for functional correlates by using data from tiling-deletion-based and CRISPR interference screens of activity of cis-regulatory elements in over 3 Mb of genome sequence. Finally, we explore two user cases that involve indels and the disruption of enhancers associated with a developmental disease. We rank variants in the non-coding genome according to their predicted deleteriousness. The model prioritizes non-coding regions associated with regulation of important genes and with cell viability, an in vitro surrogate of essentiality., Whole genome sequencing (WGS) holds promise to solve a subset of Mendelian disease cases for which exome sequencing did not provide a genetic diagnosis. Here, Wells et al. report a supervised machine learning model trained on functional, mutational and structural features for rank-scoring and interpreting variants in non-coding regions from WGS.

The spillovers of betacoronaviruses in humans and the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants highlight the need for broad coronavirus countermeasures. We describe five monoclonal antibodies (mAbs) cross-reacting with the stem helix of multiple betacoronavirus spike glycoproteins isolated from COVID-19 convalescent individuals. Using structural and functional studies, we show that the mAb with the greatest breadth (S2P6) neutralizes pseudotyped viruses from three different subgenera through the inhibition of membrane fusion, and we delineate the molecular basis for its cross-reactivity. S2P6 reduces viral burden in hamsters challenged with SARS-CoV-2 through viral neutralization and Fc-mediated effector functions. Stem helix antibodies are rare, oftentimes of narrow specificity, and can acquire neutralization breadth through somatic mutations. These data provide a framework for structure-guided design of pan-betacoronavirus vaccines eliciting broad protection. ispartof: SCIENCE vol:373 issue:6559 pages:1109-1115 ispartof: location:United States status: published

The recent emergence of SARS-CoV-2 variants of concern1-10 and the recurrent spillovers of coronaviruses11,12 into the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here we describe a human monoclonal antibody designated S2X259, which recognizes a highly conserved cryptic epitope of the receptor-binding domain and cross-reacts with spikes from all clades of sarbecovirus. S2X259 broadly neutralizes spike-mediated cell entry of SARS-CoV-2, including variants of concern (B.1.1.7, B.1.351, P.1, and B.1.427/B.1.429), as well as a wide spectrum of human and potentially zoonotic sarbecoviruses through inhibition of angiotensin-converting enzyme 2 (ACE2) binding to the receptor-binding domain. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2X259 possesses an escape profile that is limited to a single substitution, G504D. We show that prophylactic and therapeutic administration of S2X259 protects Syrian hamsters (Mesocricetus auratus) against challenge with the prototypic SARS-CoV-2 and the B.1.351 variant of concern, which suggests that this monoclonal antibody is a promising candidate for the prevention and treatment of emergent variants and zoonotic infections. Our data reveal a key antigenic site that is targeted by broadly neutralizing antibodies and will guide the design of vaccines that are effective against all sarbecoviruses.

An ideal therapeutic anti-SARS-CoV-2 antibody would resist viral escape1–3, have activity against diverse sarbecoviruses4–7, and be highly protective through viral neutralization8–11 and effector functions12,13. Understanding how these properties relate to each other and vary across epitopes would aid the development of therapeutic antibodies and guide vaccine design. Here we comprehensively characterize escape, breadth and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD). Despite a trade-off between in vitro neutralization potency and breadth of sarbecovirus binding, we identify neutralizing antibodies with exceptional sarbecovirus breadth and a corresponding resistance to SARS-CoV-2 escape. One of these antibodies, S2H97, binds with high affinity across all sarbecovirus clades to a cryptic epitope and prophylactically protects hamsters from viral challenge. Antibodies that target the angiotensin-converting enzyme 2 (ACE2) receptor-binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency. Nevertheless, we also characterize a potent RBM antibody (S2E128) with breadth across sarbecoviruses related to SARS-CoV-2 and a high barrier to viral escape. These data highlight principles underlying variation in escape, breadth and potency among antibodies that target the RBD, and identify epitopes and features to prioritize for therapeutic development against the current and potential future pandemics. A survey of SARS-CoV-2 RBD antibodies identifies those with activity against diverse SARS-CoV-2 variants and SARS-related coronaviruses, highlighting epitopes and features to prioritize in antibody and vaccine development.