Nirsevimab is a monoclonal antibody that binds to the respiratory syncytial virus (RSV) fusion protein. During the Phase 2b (NCT02878330) and MELODY (NCT03979313) clinical trials, infants received one dose of nirsevimab or placebo before their first RSV season. In this pre-specified analysis, isolates from RSV infections were subtyped, sequenced and analyzed for nirsevimab binding site substitutions; subsequently, recombinant RSVs were engineered for microneutralization susceptibility testing. Here we show that the frequency of infections caused by subtypes A and B is similar across and within the two trials. In addition, RSV A had one and RSV B had 10 fusion protein substitutions occurring at >5% frequency. Notably, RSV B binding site substitutions were rare, except for the highly prevalent I206M:Q209R, which increases nirsevimab susceptibility; RSV B isolates from two participants had binding site substitutions that reduce nirsevimab susceptibility. Overall, >99% of isolates from the Phase 2b and MELODY trials retained susceptibility to nirsevimab.

Nirsevimab is an extended half-life monoclonal antibody specific for the prefusion conformation of the respiratory syncytial virus (RSV) F protein, which has been studied in preterm and full-term infants in the phase 2b and phase 3 MELODY trials. We analyzed serum samples collected from 2,143 infants during these studies to characterize baseline levels of RSV-specific immunoglobulin G antibodies and neutralizing antibodies (NAbs), duration of RSV NAb levels following nirsevimab administration, the risk of RSV exposure during the first year of life and the infant’s adaptive immune response to RSV following nirsevimab administration. Baseline RSV antibody levels varied widely; consistent with reports that maternal antibodies are transferred late in the third trimester, preterm infants had lower baseline RSV antibody levels than full-term infants. Nirsevimab recipients had RSV NAb levels >140-fold higher than baseline at day 31 and remained >50-fold higher at day 151 and >7-fold higher at day 361. Similar seroresponse rates to the postfusion form of RSV F protein in nirsevimab recipients (68–69%) compared with placebo recipients (63–70%; not statistically significant) suggest that while nirsevimab protects from RSV disease, it still allows an active immune response. In summary, nirsevimab provided sustained, high levels of NAb throughout an infant’s first RSV season and prevented RSV disease while allowing the development of an immune response to RSV.

Background Nirsevimab is an extended half-life monoclonal antibody to the respiratory syncytial virus (RSV) fusion protein that has been developed to protect infants for an entire RSV season. Previous studies have shown that the nirsevimab binding site is highly conserved. However, investigations of the geotemporal evolution of potential escape variants in recent (ie, 2015-2021) RSV seasons have been minimal. Here, we examine prospective RSV surveillance data to assess the geotemporal prevalence of RSV A and B, and functionally characterise the effect of the nirsevimab binding-site substitutions identified between 2015 and 2021. Methods We assessed the geotemporal prevalence of RSV A and B and nirsevimab binding-site conservation between 2015 and 2021 from three prospective RSV molecular surveillance studies (the US-based OUTSMART-RSV, the global INFORM-RSV, and a pilot study in South Africa). Nirsevimab binding-site substitutions were assessed in an RSV microneutralisation susceptibility assay. We contextualised our findings by assessing fusion-protein sequence diversity from 1956 to 2021 relative to other respiratory-virus envelope glycoproteins using RSV fusion protein sequences published in NCBI GenBank. Findings We identified 5675 RSV A and RSV B fusion protein sequences (2875 RSV A and 2800 RSV B) from the three surveillance studies (2015-2021). Nearly all (25 [100%] of 25 positions of RSV A fusion proteins and 22 [88%] of 25 positions of RSV B fusion proteins) amino acids within the nirsevimab binding site remained highly conserved between 2015 and 2021. A highly prevalent (ie, >40 & BULL;0% of all sequences) nirsevimab binding-site Ile206Met:Gln209Arg RSV B polymorphism arose between 2016 and 2021. Nirsevimab neutralised a diverse set of recombinant RSV viruses, including new variants containing binding-site substitutions. RSV B variants with reduced susceptibility to nirsevimab neutralisation were detected at low frequencies (ie, prevalence <1 & BULL;0%) between 2015 and 2021. We used 3626 RSV fusion-protein sequences published in NCBI GenBank between 1956 and 2021 (2024 RSV and 1602 RSV B) to show that the RSV fusion protein had lower genetic diversity than influenza haemagglutinin and SARS-CoV-2 spike proteins. Interpretation The nirsevimab binding site was highly conserved between 1956 and 2021. Nirsevimab escape variants were rare and have not increased over time. Funding AstraZeneca and Sanofi. Copyright & COPY; 2023 Elsevier Ltd. All rights reserved.

BACKGROUND:Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infection and hospitalization in infants. Nirsevimab is a monoclonal antibody to the RSV fusion protein that has an extended half-life. The efficacy and safety of nirsevimab in healthy late-preterm and term infants are uncertain.METHODS:We randomly assigned, in a 2:1 ratio, infants who had been born at a gestational age of at least 35 weeks to receive a single intramuscular injection of nirsevimab or placebo before the start of an RSV season. The primary efficacy end point was medically attended RSV-associated lower respiratory tract infection through 150 days after the injection. The secondary efficacy end point was hospitalization for RSV-associated lower respiratory tract infection through 150 days after the injection.RESULTS:A total of 1490 infants underwent randomization: 994 were assigned to the nirsevimab group and 496 to the placebo group. Medically attended RSV-associated lower respiratory tract infection occurred in 12 infants (1.2%) in the nirsevimab group and in 25 infants (5.0%) in the placebo group; these findings correspond to an efficacy of 74.5% (95% confidence interval [CI], 49.6 to 87.1; P<0.001) for nirsevimab. Hospitalization for RSV-associated lower respiratory tract infection occurred in 6 infants (0.6%) in the nirsevimab group and in 8 infants (1.6%) in the placebo group (efficacy, 62.1%; 95% CI, -8.6 to 86.8; P = 0.07). Among infants with data available to day 361, antidrug antibodies after baseline were detected in 58 of 951 (6.1%) in the nirsevimab group and in 5 of 473 (1.1%) in the placebo group. Serious adverse events were reported in 67 of 987 infants (6.8%) who received nirsevimab and in 36 of 491 infants (7.3%) who received placebo.CONCLUSIONS:A single injection of nirsevimab administered before the RSV season protected healthy late-preterm and term infants from medically attended RSV-associated lower respiratory tract infection. (Funded by MedImmune/AstraZeneca and Sanofi; MELODY ClinicalTrials.gov number, NCT03979313.).

SARS-CoV-2 variants harbor multiple substitutions in their spike trimers, potentially leading to breakthrough infections and clinical resistance to immune therapies. For this reason, a BLI assay was developed and qualified to evaluate the reliability of screening SARS-CoV-2 spike trimer variants against anti-SARS-CoV-2 monoclonal antibodies (MAbs) tixagevimab and cilgavimab, the components of AZD7442, prior to in vitro pseudovirus neutralization susceptibility verification testing.

Abstract Background AZD7442 is a combination of extended–half-life SARS-CoV-2–neutralizing monoclonal antibodies (tixagevimab/cilgavimab) that bind to distinct epitopes on the SARS-CoV-2 spike protein. In the PROVENT study, a single 300 mg intramuscular dose of AZD7442 demonstrated 77% efficacy for prevention of COVID-19 vs placebo at primary analysis, with 83% efficacy through 6-months follow-up, and was well-tolerated. We report conservation of AZD7442 binding sites and neutralizing activity against pseudotyped virus-like particles (VLPs) harboring spike substitutions identified in surveillance, and clinical SARS-CoV-2 isolates from the PROVENT study. Methods Consensus SARS-CoV-2 whole genome sequences were analyzed from open source databases to identify prevalent spike substitutions within the AZD7442 binding site. Phenotypic analyses determined neutralization susceptibility of pseudotyped VLPs with identified spike substitutions. Genotypic analyses were also performed on SARS-CoV-2 spike sequences from PROVENT study (NCT04625725) participants with RT-PCR-positive symptomatic illness. Results Most residues in tixagevimab (13/17) and cilgavimab (13/19) binding sites were >99% conserved among global SARS-CoV-2 isolates (N=8,373,740 through Apr 19, 2022). In 2021, AZD7442 binding site polymorphisms emerged among circulating strains (prevalence: R346K, 11%; N440K, 22%; G446S, 15%; S477N, 28%; L452R, 43%; T478K, 70%; E484A, 27%; E484K, 3%; Q493R, 27%), but these did not affect AZD7442 in vitro neutralization potency. AZD7442 retained neutralization activity against variants of concern or interest tested, including Omicron BA.2, with moderate reduction observed for Omicron BA.1. By median 6-months follow-up (Aug 29, 2021, data cut-off) in the PROVENT study, there were no AZD7442-resistant substitutions observed in breakthrough SARS-CoV-2 illness visits. Conclusion AZD7442 retained neutralization activity against all SARS-CoV-2 variants of concern or interest evaluated. Binding site substitutions identified in circulation, and in breakthrough SARS-CoV-2 infections following a single 300 mg dose of AZD7442 in the PROVENT study, were not associated with AZD7442 escape. Disclosures Kevin M. Tuffy, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Michael E. Abram, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Tiffany L. Roe, B.S., AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Bahar Ahani, BSC, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Tyler Brady, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Nicolette Schuko, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Lori Clarke, B.S., AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Carolina Caceres, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Tara Kenny, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Virginia Takahashi, B.S., AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Tianhui Zhang, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds David E. Tabor, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Gustavo H. Kijak, PharmD PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Elizabeth J. Kelly, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Katie Streicher, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds.

Despite the success of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines, there remains a need for more prevention and treatment options for individuals remaining at risk of coronavirus disease 2019 (COVID-19). Monoclonal antibodies (mAbs) against the viral spike protein have potential to both prevent and treat COVID-19 and reduce the risk of severe disease and death. Here, we describe AZD7442, a combination of two mAbs, AZD8895 (tixagevimab) and AZD1061 (cilgavimab), that simultaneously bind to distinct, nonoverlapping epitopes on the spike protein receptor binding domain to neutralize SARS-CoV-2. Initially isolated from individuals with prior SARS-CoV-2 infection, the two mAbs were designed to extend their half-lives and reduce effector functions. The AZD7442 mAbs individually prevent the spike protein from binding to angiotensin-converting enzyme 2 receptor, blocking virus cell entry, and neutralize all tested SARS-CoV-2 variants of concern. In a nonhuman primate model of SARS-CoV-2 infection, prophylactic AZD7442 administration prevented infection, whereas therapeutic administration accelerated virus clearance from the lung. In an ongoing phase 1 study in healthy participants (NCT04507256), a 300-mg intramuscular injection of AZD7442 provided SARS-CoV-2 serum geometric mean neutralizing titers greater than 10-fold above those of convalescent serum for at least 3 months, which remained threefold above those of convalescent serum at 9 months after AZD7442 administration. About 1 to 2% of serum AZD7442 was detected in nasal mucosa, a site of SARS-CoV-2 infection. Extrapolation of the time course of serum AZD7442 concentration suggests AZD7442 may provide up to 12 months of protection and benefit individuals at high-risk of COVID-19.

Abstract Background Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infection (LRTI) and hospitalization in infants. In two global pivotal placebo-controlled studies, nirsevimab, a monoclonal antibody to the RSV prefusion (F) protein with extended half-life, reduced medically attended (MA) RSV LRTI versus placebo throughout the RSV season (MELODY (Primary Cohort)/Study 3 (Proposed Dose) Pool, 79.5% efficacy). Here we summarize resistance analyses of all RT-PCR-confirmed RSV isolates from healthy term and preterm infants through 150 days post dose. Methods Infants were randomized 2:1 to receive one intramuscular injection of nirsevimab or placebo, prior to their first RSV season. RT-PCR-confirmed RSV isolates were reflexed for genotypic analyses of RSV F and phenotypic analyses of identified substitutions in a recombinant RSV neutralization susceptibility assay. Results In the pooled proposed dose analysis of Study 3 (50 mg nirsevimab if < 5 kg at dosing) and MELODY (50 or 100 mg nirsevimab if < 5 kg or ≥5 kg at dosing, respectively), no subject with MA RSV LRTI had an RSV isolate containing nirsevimab resistance-associated substitutions in either treatment group (nirsevimab, RSV A: 0/14 and RSV B: 0/5; placebo, RSV A: 0/35 and RSV B: 0/16). In Study 3 (50 mg nirsevimab if ≥5 kg at dosing), 2/18 subjects in the nirsevimab group and 0/20 subjects in the placebo group with MA RSV LRTI had an RSV isolate harbouring nirsevimab binding site substitutions I64T+K68E+I206M+Q209R (>447-fold) or N208S (>387-fold) that conferred reduced susceptibility to nirsevimab neutralization (nirsevimab, RSV A: 0/9 and RSV B: 2/9; placebo, RSV A: 0/10 and RSV B: 0/10). Subjects with RSV isolates harboring F protein sequence variations that maintained susceptibility to nirsevimab neutralization were balanced between treatment groups with no association with RSV disease severity. No subjects with non-protocol defined MA RSV LRTI cases or hospitalization due to any RSV respiratory illness had an RSV isolate conferring nirsevimab resistance. Conclusion Lack of nirsevimab resistance following immunization at the proposed dose supports efficacy and neutralization activity of nirsevimab against both RSV A and B strains throughout the RSV season. Disclosures Michael E. Abram, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Bahar Ahani, BSC, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds David E. Tabor, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Fiona Fernandes, PhD, AstraZeneca: Stocks/Bonds Deidre Wilkins, BSC, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Anastasia A. Aksyuk, PhD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Kevin M. Tuffy, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Hong Ji, BSc, AstraZeneca: Stocks/Bonds Christine Blaze, BSc, AstraZeneca: Stocks/Bonds Tyler Brady, MS, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Pamela Griffin, MD, AstraZeneca: Stocks/Bonds Amanda Leach, MD, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Tonya L. Villafana, PhD, MPH, AstraZeneca: Employee|AstraZeneca: Stocks/Bonds Mark T. Esser, PhD, AstraZeneca: Stocks/Bonds.

BACKGROUND:More than 2 million SARS-CoV-2 genome sequences have been generated and shared since the start of the COVID-19 pandemic and constitute a vital information source that informs outbreak control, disease surveillance, and public health policy. The Pango dynamic nomenclature is a popular system for classifying and naming genetically-distinct lineages of SARS-CoV-2, including variants of concern, and is based on the analysis of complete or near-complete virus genomes. However, for several reasons, nucleotide sequences may be generated that cover only the spike gene of SARS-CoV-2. It is therefore important to understand how much information about Pango lineage status is contained in spike-only nucleotide sequences. Here we explore how Pango lineages might be reliably designated and assigned to spike-only nucleotide sequences. We survey the genetic diversity of such sequences, and investigate the information they contain about Pango lineage status.RESULTS:Although many lineages, including the main variants of concern, can be identified clearly using spike-only sequences, some spike-only sequences are shared among tens or hundreds of Pango lineages. To facilitate the classification of SARS-CoV-2 lineages using subgenomic sequences we introduce the notion of designating such sequences to a "lineage set", which represents the range of Pango lineages that are consistent with the observed mutations in a given spike sequence.CONCLUSIONS:We find that many lineages, including the main variants-of-concern, can be reliably identified by spike alone and we define lineage-sets to represent the lineage precision that can be achieved using spike-only nucleotide sequences. These data provide a foundation for the development of software tools that can assign newly-generated spike nucleotide sequences to Pango lineage sets.

Background Respiratory syncytial virus (RSV) is a global cause of severe respiratory morbidity and mortality in infants. While preventive and therapeutic interventions are being developed, including antivirals, vaccines and monoclonal antibodies, little is known about the global molecular epidemiology of RSV. INFORM is a prospective, multicenter, global clinical study performed by ReSViNET to investigate the worldwide molecular diversity of RSV isolates collected from children less than 5 years of age. Methods The INFORM study is performed in 17 countries spanning all inhabited continents and will provide insight into the molecular epidemiology of circulating RSV strains worldwide. Sequencing of > 4000 RSV-positive respiratory samples is planned to detect temporal and geographical molecular patterns on a molecular level over five consecutive years. Additionally, RSV will be cultured from a subset of samples to study the functional implications of specific mutations in the viral genome including viral fitness and susceptibility to different monoclonal antibodies. Discussion The sequencing and functional results will be used to investigate susceptibility and resistance to novel RSV preventive or therapeutic interventions. Finally, a repository of globally collected RSV strains and a database of RSV sequences will be created.

Background Vesatolimod (VES; GS-9620) is a Toll-like receptor 7 (TLR7) agonist that directly activates human plasmacytoid dendritic cells (pDCs) and B lymphocytes resulting in direct and indirect production of cytokines and immune activation. VES is being evaluated in HIV-1-infected people as part of an HIV remission strategy. Here we investigated the potential of VES to trigger indirect activation of HIV-specific CD8 + T-cells using immune cell cultures derived from HIV+ donors. Methods Peripheral blood mononuclear cell (PBMC) cultures derived from HIV+ donors virologically suppressed on stable antiretroviral therapy ( n=31) were isolated and treated with VES or vehicle for 24 h. Cells were stained with surface and intracellular fluorescent conjugated antibodies and HIV-specific pentamers, and analysed by flow cytometry. Results Treatment of PBMCs with VES resulted in all 31 donors demonstrating a concentration dependent increase in CD8 + T-cell activation (CD69 + ) of up to 88%. Of these donors, 20 of 31 donors displayed a concentration-dependent increase in HIV-specific CD8 + T-cell activation due to VES with a maximum of 20.8%. Intracellular staining was performed in a subset of donors ( n=14), 5 of which displayed VES-induced activation of functional HIV-specific CD8 + T-cells as assessed by CD107a and/or tumour necrosis factor (TNF)-α upregulation. Conclusions This study demonstrates that VES treatment can induce the activation of functional HIV-specific CD8 + T-cells in donor derived PBMCs. These data support the potential use of VES to activate functional HIV-specific CD8 + T-cells as part of an HIV remission strategy.

Tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF) are prodrugs of the HIV-1 nucleotide reverse transcriptase inhibitor tenofovir (TFV). In vivo, TAF achieves >4-fold-higher intracellular levels of TFV diphosphate (TFV-DP) compared to TDF. Since thymidine analog-associated mutations (TAMs) in HIV-1 confer reduced TFV susceptibility, patients with TAM-containing HIV-1 may benefit from higher TFV-DP levels delivered by TAF. Moreover, the presence of the M184V mutation increases TFV susceptibility during TDF- or TAF-based therapy. The susceptibilities to antiviral drugs of site-directed mutants (SDMs) and patient-derived mutants containing combinations of TAMs (M41L, D67N, K70R, L210W, T215Y, and K219Q) with or without the M184V mutation (TAMsi - M184V) were evaluated using either 5-day multicycle (MC; n = 110) or 2-day single-cycle (SC; n = 96) HIV assays. The presence of M184V in TAM-containing HIV-1 SDMs (n = 48) significantly increased TAF sensitivity compared to SDMs without M184V (n = 48). The comparison of TAF and TDF resistance profiles was further assessed in viral breakthrough (VB) experiments mimicking clinically relevant drug concentrations. A total of 68 mutants were assayed at physiological concentration in VB experiments, with 15/68 mutants breaking through with TDF (TFV, the in vitro equivalent of TDF, was used in these experiments), and only 3 of 68 mutants breaking through under TAF treatment. Overall, in the VB assay mimicking the 4-fold-higher intracellular levels of TFV-DP observed clinically with TAF versus TDF, TAF inhibited viral breakthrough of most TAM-containing HIV-1, whereas TDF did not. These results indicate that TAF has a higher resistance threshold than TDF and suggest that higher resistance cutoffs should be applied for TAF compared to TDF in genotypic and phenotypic resistance algorithms.

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection among infants and young children, resulting in annual epidemics worldwide. INFORM-RSV is a multiyear clinical study designed to describe the global molecular epidemiology of RSV in children under 5 years of age by monitoring temporal and geographical evolution of current circulating RSV strains, F protein antigenic sites, and their relationships with clinical features of RSV disease. During the pilot season (2017–2018), 410 RSV G-F gene sequences were obtained from 476 RSV-positive nasal samples collected from 8 countries (United Kingdom, Spain, The Netherlands, Finland, Japan, Brazil, South Africa, and Australia).

The development of resistance to human immunodeficiency virus 1 (HIV-1) integrase strand-transfer inhibitors (INSTI) has been documented; however, knowledge of the impact of pre-existing integrase (IN) mutations on INSTI resistance (INSTI-R) is still evolving. The frequency of HIV-1 IN mutations in 2177 treatment-naïve subjects was investigated, along with the INSTI susceptibility of site-directed mutant viruses containing major and minor INSTI-R mutations. Total 6 of 39 minor INSTI-R mutations (M50I, S119P/G/T/R, and E157Q) were found in >1% of IN-treatment-naïve subjects with no impact on INSTI susceptibility. When each combined with major INSTI-R mutation, M50I, S119P, and E157Q led to decreased susceptibility to elvitegravir but remained sensitive to dolutegravir and bictegravir.

Background: FTC/TAF was shown to be noninferior to FTC/TDF with advantages in markers of renal and bone safety. Objective: To evaluate the efficacy and safety of switching to FTC/TAF from FTC/TDF by third agent (boosted protease inhibitor [PI] vs. unboosted third agent). Methods: We conducted a 48-week subgroup analysis based on third agent from a randomized, double blind study in virologically suppressed adults on a FTC/TDF-containing regimen who switched to FTC/TAF vs. continued FTC/TDF while remaining on the same third agent. Results: We randomized (1:1) 663 participants to either switch to FTC/TAF (N = 333) or continue FTC/TDF (N = 330), each with baseline third agent stratifying by class of third agent in the prior treatment regimen (boosted PI 46%, unboosted third agent 54%). At week 48, significant differences in renal biomarkers and bone mineral density were observed favoring FTC/TAF over FTC/TDF (p < 0.05 for all), with similar improvements in the FTC/TAF arm in those who received boosted PI vs. unboosted third agents. At week 48, virologic success rates were similar between treatment groups for those who received a boosted PI (FTC/TAF 92%, FTC/TDF 93%) and for those who received an unboosted third agent (97% vs. 93%). Conclusions: In virologically suppressed patients switching to FTC/TAF from FTC/TDF, high rates of virologic suppression were maintained, while renal and bone safety parameters improved, regardless of whether participants were receiving a boosted PI or an unboosted third agent. FTC/TAF offers safety advantages over FTC/TDF and can be an important option as an NRTI backbone given with a variety of third agents.