Nirsevimab is an extended half-life (M252Y/S254T/T256E [YTE]-modified) monoclonal antibody that binds to the prefusion (pre-F) conformation of the respiratory syncytial virus (RSV) fusion (F) protein and is licensed for the prevention of RSV lower respiratory tract disease in neonates, infants and medically vulnerable children. Previous analyses have shown that nirsevimab immunization does not impair the development of antibody (Ab) responses to the G attachment, nucleocapsid or postfusion (post-F) conformation of the RSV F protein. Pre-F-specific Abs determine the magnitude of RSV neutralizing activity. However, the impact of nirsevimab immunization on pre-F responses has been difficult to assess due to shared epitopes. Here we describe a method of depleting YTE-modified Abs from serum, and its application in assessing natural pre-F binding and neutralizing Ab (nAb) responses following nirsevimab immunization in participants from the placebo-controlled Phase 3 MELODY study (NCT03979313).Figure 1 Participants provided serum samples during site visits on Day 151 and 361. Nirsevimab was selectively depleted from serum using streptavidin beads coupled to an anti-YTE capture Ab. Pre-F Ab and nAb levels were respectively measured by multiplex serology and fluorescent focus-based microneutralization assay before and after YTE-depletion. Participant RSV exposure was determined by a diagnostic test or post-F seroresponse.Figure 2 Pre-F Ab responses were induced in nirsevimab-immunized participants with diagnostically confirmed RSV exposure or post-F seroresponses (Figure 1). The range of pre-F Ab responses in nirsevimab-immunized participants overlapped with those from placebo participants with RSV exposure, albeit with lower median concentrations, and was higher than responses in placebo participants without exposure. Similar trends were observed for nAb responses above the assay’s lower limit of detection. The range of nAb responses in nirsevimab-immunized participants was similar, albeit lower, than observed in placebo participants with RSV exposure, and higher than in placebo participants without exposure (Figure 2). Nirsevimab immunization does not prevent an infant’s immune system from developing natural pre-F binding and nAb responses. Ann Marie Stanley, PhD, AstraZeneca: Employement|AstraZeneca: Stocks/Bonds (Public Company) Vancheswaran Gopalakrishnan, PhD, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company) Carolina Caceres, MS, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company) Kevin M. Tuffy, MS, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company) Beth Kelly, PhD, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company)|Sanofi: Employment|Sanofi: Stocks/Bonds (Private Company) Mark T. Esser, PhD, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company) Tonya L. Villafana, PhD, MPH, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company) Anastasia A. Aksyuk, PhD, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company)|MesoScale Diagnostics: Intellectual Property/Patents Deidre Wilkins, BSC, AstraZeneca: Employment|AstraZeneca: Stocks/Bonds (Public Company)

Local intramuscular administration of synthetic plasmid DNA (pDNA) encoding monoclonal antibodies (mAbs) offers a promising alternative to traditional recombinant protein-based mAb delivery. This approach may enable durable in vivo expression of functional antibodies and overcome limitations related to cost, production, and cold-chain logistics. AZD5396 and AZD8076 are modified versions of the SARS-CoV-2 neutralizing antibody cocktail Evusheld, encoded as DNA-delivered monoclonal antibodies (DMAbs). CONSORT diagram and trial schematic Longitudinal serum concentration of in vivo-expressed DMAbs AZD5396 and AZD8076 In this Phase 1, dose-escalation study (ClinicalTrials.gov identifier: NCT05293249), we evaluated the safety, tolerability, and pharmacokinetics of a pDNA cocktail encoding AZD5396 and AZD8076 in healthy adults. Participants received up to four intramuscular doses of the pDNA cocktail delivered by CELLECTRA™ electroporation. The primary endpoints were safety and pharmacokinetics. Exploratory endpoints included anti-drug antibody (ADA) development and functional activity against SARS-CoV-2 variants. All 44 enrolled participants received at least one dose, and DMAbs were detected in 100% of evaluable participants (n=39). Serum DMAb concentrations reached a mean peak of 1.11 µg/mL, with sustained expression observed in all participants who completed 72 weeks of follow-up. The product was well tolerated, and no product-related serious adverse events were reported. Exploratory analyses demonstrated binding to multiple SARS-CoV-2 spike variants and neutralizing activity in pseudovirus assays. Across ∼1,000 serum samples, no ADAs were detected using validated tiered assays. These findings provide the first-in-human proof-of-concept that synthetic pDNA DMAb technology enables durable in vivo production of a functional mAb cocktail. The results highlight the critical role of optimized synthetic design, formulation, and delivery in achieving biologically relevant expression. DNA-delivered mAbs may represent a long-acting, scalable, cold-chain-independent platform for targeting a wide range of diseases treatable with antibody-based therapeutics. All Authors: No reported disclosures

Antibiotic combination therapy is often used to broaden the antimicrobial spectrum, limit resistance and improve treatment efficacy. Several antibiotics show collateral effects where resistance to one antibiotic increases susceptibility to another. In intensive care units (ICUs), antibiotic treatments are frequently adjusted based on patient outcomes, without considering collateral effects. This provides a setting to study these effects in Pseudomonas aeruginosa (PA), a highly adaptable, multidrug-resistant (MDR), nosocomial pathogen. We compared longitudinal PA isolates from twenty-five ventilated ICU patients receiving various antibiotics to laboratory strains undergoing in vitro adaptive evolution under four antipseudomonal monotherapies. Prolonged exposure to certain antibiotics produced resistance with collateral effects. In vitro, increasing antibiotic pressure drove distinct mutational trajectories. In patients, the number of antibiotics administered did not correlate with resistance changes to those antibiotics, suggesting that switching may reduce persistence of resistance. Notably, an inverse correlation between resistance to non-administered antibiotics and the number of different antibiotic classes administered, aligns with the principles of collateral susceptibility driven by multi-class exposure. This study provides s real-world evidence that empirical antibiotic mixing in ICU patients leverages evolutionary trade-offs. Consequently, diversifying antibiotic pressure via multi-class exposure may attenuate the fixation and persistence of MDR phenotypes in critical care.

The PROVENT study demonstrated the efficacy and safety of a single 300-mg dose of AZD7442 (tixagevimab/cilgavimab) for pre-exposure prophylaxis of COVID-19 in at-risk individuals. Here we report an analysis of repeat dosing of intramuscular AZD7442 300 and 600 mg from the PROVENT sub-study. The sub-study enrolled eligible participants from the parent study, creating four sub-study groups. Group 1 received AZD7442 300 mg in PROVENT followed by one 300-mg dose in the sub-study (10–14 months apart). Group 2 received placebo in PROVENT followed by two AZD7442 300-mg doses 6 months apart in the sub-study. Group 3a received AZD7442 300 mg in PROVENT followed by one 300-mg dose and two 600-mg doses 6 months apart in the sub-study. Group 3b received placebo in PROVENT followed by one 300-mg dose and two 600-mg doses 6 months apart in the sub-study. The primary endpoint was safety. Secondary endpoints included pharmacokinetics and anti-drug antibody (ADA) responses. Adverse events (AEs) and serious AEs (SAEs) were reported in 75.7–81.5

Monoclonal antibodies have potential as rapidly developable agents for treatment and prevention of emerging viruses. The ACTIV-2 trial randomized persons with mild-moderate COVID-19 to the monoclonal antibody combination tixagevimab/cilgavimab via intramuscular injection (600 mg IM) or infusion (300 mg IV) versus placebo. We present final safety and laboratory outcomes; primary outcomes were previously reported. The analyzed IM group included 214 participants, and the IV group, 106 participants. Adverse events were not different between treatment and placebo. The half-life of both components was >90 days for IM or 75 days for IV. New anti-drug antibodies were about 3 times more likely in active vs. placebo recipients. SARS-CoV-2 neutralizing antibodies increased 157-fold at 7 days and 127-fold at 1 month (IM-treated) but were less robust in IV participants. These data can inform future development of monoclonal antibodies against SARS-CoV-2 and other viruses, even if this intervention is of low utility for contemporary SARS-CoV-2 variants.

Most previously authorized clinical antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have lost neutralizing activity to recent variants due to rapid viral evolution. To mitigate such escape, we preemptively enhance AZD3152, an antibody authorized for prophylaxis in immunocompromised individuals. Using deep mutational scanning (DMS) on the SARS-CoV-2 antigen, we identify AZD3152 vulnerabilities at antigen positions F456 and D420. Through two iterations of computational antibody design that integrates structure-based modeling, machine-learning, and experimental validation, we co-optimize AZD3152 against 24 contemporary and previous SARS-CoV-2 variants, as well as 20 potential future escape variants. Our top candidate, 3152-1142, restores full potency (100-fold improvement) against the more recently emerged XBB.1.5+F456L variant that escaped AZD3152, maintains potency against previous variants of concern, and shows no additional vulnerability as assessed by DMS. This preemptive mitigation demonstrates a generalizable approach for optimizing existing antibodies against potential future viral escape.

Clinical development of monoclonal antibodies (mAbs) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is challenging due to rapid changes in the variant landscape. This study identified a threshold model for neutralising antibody (nAb) titres associated with clinically relevant protection against symptomatic COVID-19 for vulnerable populations. Using efficacy data from the phase 3 PROVENT pre-exposure prophylaxis trial of tixagevimab-cilgavimab (NCT04625725), individual nAb ID50 titres were predicted by dividing serum mAb concentration by prevalence-adjusted tixagevimab-cilgavimab potency (from in vitro IC50 values combined with viral surveillance data) and related to efficacy with a Cox model. The Threshold of Protection (ToP) Cox model was externally validated using data from the phase 3 SUPERNOVA trial (NCT05648110), which assessed sipavibart efficacy against symptomatic COVID-19 in immunocompromised participants. The PROVENT ToP model estimated the variant-specific observed efficacies from SUPERNOVA for 3 and 6 months post any dose with Lin's concordance of 0.86 and 0.75, respectively. This approach integrates predicted nAb ID50 titres against multiple SARS-CoV-2 variants into a ToP model that can be applied across different variants and could serve as a surrogate endpoint in immunobridging studies to expedite clinical evaluation and regulatory approval for mAbs targeting SARS-CoV-2.

Influenza virus pandemics and seasonal epidemics have claimed countless lives. Recurrent zoonotic spillovers of influenza viruses with pandemic potential underscore the need for effective countermeasures. In this study, we show that pre-exposure prophylaxis with broadly neutralizing antibody (bnAb) MEDI8852 is highly effective in protecting cynomolgus macaques from severe disease caused by aerosolized highly pathogenic avian influenza H5N1 virus infection. Protection was antibody dose-dependent yet independent of Fc-mediated effector functions at the dose tested. Macaques receiving MEDI8852 at 10 milligrams per kilogram or higher had negligible impairment of respiratory function after infection, whereas control animals were not protected from severe disease and fatality. Given the breadth of MEDI8852 and other bnAbs, we anticipate that protection from unforeseen pandemic influenza A viruses is achievable.

Abstract Background The SARS-CoV-2 antibody combination tixagevimab/cilgavimab (AZD7442) was efficacious in preventing COVID-19 over 6 months following a single 300 mg intramuscular (IM) dose in the PROVENT study. Participants included those that: (a) were immunocompromised and/or at increased risk for inadequate response to COVID-19 vaccination, or (b) were at increased severe COVID-19 risk. Following PROVENT, emergence of Omicron variants with reduced AZD7442 susceptibility prompted increase in the authorized dose and dosing frequency. Here, we report safety and pharmacokinetics (PK) of repeated doses of AZD7442 300–600 mg in participants who opted to continue in the PROVENT substudy. Methods Participants who entered the PROVENT open-label sub-study (NCT04625725) were assigned initially to 2 groups: Group 1 (n=234) received 1x AZD7442 300 mg in the parent study, then 1x 300 mg ∼10–14 months later (substudy Day 1). Group 2 (n=119) received placebo in the parent study, then 2x AZD7442 300 mg doses in the substudy. Group 3a (n=76) was a subset of Group 1 who received 2x 300 mg then 2x 600 mg doses. Group 3b (n=74) was a subset of Group 2 who received 1x 300 mg then 2x 600 mg doses (Table 1). The primary endpoint was safety; PK and antibodies to AZD7442 were also assessed. Results Baseline characteristics were overall similar across the groups. Adverse events (AEs) were reported in 75.7–81.5% and serious AEs in 13.2–16.8% of participants across the groups (Table 2). There were 14 deaths: 8 (3.4%) in Group 1, 5 (4.2%) in Group 2, and 1 (1.4%) in Group 3b. There was no clinically meaningful increase of AEs due to repeat doses of AZD7442 and no deaths were considered related to AZD7442. AZD7442 PK was consistent following redosing and between groups (Figure 1). The percentage of participants positive for treatment-emergent antibodies to AZD7442 was 10.5% (Group 1), 5.2% (Group 2), 10.7% (Group 3a), and 4.1% (Group 3b). Conclusion Safety and PK of AZD7442 were consistent after repeat dosing, regardless of whether participants initially received AZD7442 or placebo. These results will support future development of long-acting antibodies for pre-exposure prophylaxis against COVID-19. Disclosures Andrew Ustianowski, MD, PhD, AstraZeneca: Honoraria|Gilead: Honoraria|GSK: Honoraria|GSK: Speaker fees|Janssen: Honoraria|Janssen: Speaker fees|Merck: Honoraria|Merck: Speaker fees, Advisory Board|Pfiizer: Advisory Board|Sanofi: Honoraria|Sanofi: Speaker fees|ViiV Healthcare/GSK: Advisory Board Myron J. Levin, MD, CSL Seqirus USA: Advisor/Consultant|Curevo: Advisory Board|GSK: Grant/Research Support|GSK: Advisory Board|Moderna: Advisory Board|Pfizer: Advisory Board Stéphane De Wit, MD, AstraZeneca: Honoraria|AstraZeneca: Speaker fees|Gilead: Honoraria|Gilead: Speaker fees, Advisory Board|GSK: Honoraria|GSK: Speaker fees|Janssen: Honoraria|Janssen: Speaker fees|Merck: Honoraria|Merck: Speaker fees, Advisory Board|Pfizer: Advisory Board|Sanofi: Honoraria|Sanofi: Speaker fees|ViiV Healthcare/GSK: Advisory Board Odile Launay, MD, PhD, AstraZeneca: Principal investigator for the AstraZeneca-sponsored PROVENT study in France|AstraZeneca and other pharmaceutical companies: Honoraria|AstraZeneca and other pharmaceutical companies: Speaker fees Audrey Sharbaugh, PhD, AstraZeneca: Employee, holds or may hold stock Rohini Beavon, PhD, AstraZeneca: Employee, holds or may hold stock Jesse Thissen, MSc, AstraZeneca: Employee, holds or may hold stock Lauren Hirao, PhD, AstraZeneca: Employee, holds or may hold stock Vitalina Dzutseva, MD, PhD, AstraZeneca: Employee, holds or may hold stock Seth Seegobin, PhD, AstraZeneca: Employee, holds or may hold stock Katie Streicher, PhD, AstraZeneca: Employee of AstraZeneca and may own AstraZeneca stock or stock options. Alexandre Kiazand, MD, AstraZeneca: Employee, holds or may hold stock Mark T. Esser, PhD, AstraZeneca: Employee, holds or may hold stock Taylor Cohen, PhD, AstraZeneca: Employee, holds or may hold stock Lee-Jah Chang, MD, AstraZeneca: Employee of AstraZeneca John L. Perez, MD, AstraZeneca: Employee, holds or may hold stock

Objectives: Pseudomonas aeruginosa (PA) is a common causative pathogen of pneumonia acquired in the intensive care unit (ICU). The aim of this study was to determine the incidence of PA ICU pneumonia (PAIP) and to quantify its independent association with PA colonization at different body sites. Methods: Adult patients on mechanical ventilation at ICU admission were prospectively enrolled across 30 European ICUs. PA colonization in the perianal area and in the lower respiratory tract was assessed within 72 hours after ICU admission and twice weekly until ICU discharge. PAIP development was evaluated daily. Competing risk models with colonization as a time-varying exposure and ICU death and discharge as competing events were fitted and adjusted for confounders to investigate the association between PA carriage and PAIP. Results: A total of 1971 subjects were enrolled. The colonization prevalence with PA in the first 72 hours of ICU admission was 10.4% (179 perianal and 51 respiratory), whereas the acquisition incidence during the ICU stay was 7.0% (158 perianal and 47 respiratory). Of the 43 (1.8%) patients who developed PAIP, 11 (25.6%) were PA colonized on admission and 9 (20.9%) acquired colonization before PAIP onset. Both perianal (adjusted subdistribution hazard ratio, 4.4; 95% CI, 1.7-11.6) and respiratory colonization (adjusted subdistribution hazard ratio: 4.6, 95% CI, 1.9-11.1) were independently associated with PAIP development. Discussion: PAIP incidence was higher in PA colonized vs. non-colonized patients. Colonization of both the rectum and of the respiratory tract was associated with development of PAIP. The increased risk of PA colonization for subsequent infection provides an opportunity for targeted preventive interventions. (c) 2024 The Author(s). Published by Elsevier Ltd on behalf of European Society of Clinical Microbiology and Infectious Diseases. This is an open access article under the CC BY license (http://creativecommons. org/licenses/by/4.0/).

BACKGROUND:Nirsevimab is an extended half-life, highly potent, anti-respiratory syncytial virus (RSV) fusion protein neutralizing monoclonal antibody with efficacy against RSV-associated medically attended (MA) lower respiratory tract infection (LRTI) in infants and medically vulnerable children (aged ≤24 months). This post hoc exploratory analysis examined the incidence of LRTI from RSV and other respiratory pathogens during MELODY: a 2:1 randomized, double-blind, placebo-controlled, phase 3 study of nirsevimab in healthy term and late preterm (ie, gestational age ≥35 weeks) infants entering their first RSV season. METHODS:A total of 3012 participants were randomized to nirsevimab (n = 2009) or placebo (n = 1003). Nasopharyngeal swabs were collected from infants who presented with an LRTI and tested for 22 different respiratory pathogens using the BioFire® Respiratory 2.1 Panel. Incidence of RSV and non-RSV MA-LRTIs through day 511 and LRTI severity were assessed. RESULTS:A total of 852 nasopharyngeal swabs were collected from 561 participants through day 511: 519 swabs from 337 nirsevimab participants and 333 swabs from 224 placebo participants. RSV and non-RSV infections were detected in 193 of 852 (22.7%) and 55 of 852 (64.7%) swabs, respectively. RSV infection rates were lower with nirsevimab compared with placebo, including RSV-rhinovirus/enterovirus coinfections. Rates of other viral infections were similar between study arms. Approximately 70% of single RSV infections and RSV coinfections were adjudicated as mild, and 26.2% of single RSV infections and 24.5% of RSV coinfections required hospitalization. CONCLUSIONS:Nirsevimab protected against RSV single and coinfections, with no evidence of replacement of RSV with other respiratory viruses. Clinical Trials Registration. NCT03979313.

Supplementary Table S1. Representativeness of Study Participants Supplementary Table S2. Disease response as assessed by RECIST v1.1 by PD-L1 tumor cell expression (response-evaluable population). Supplementary Table S3. Disease response as assessed by RECIST v1.1 in prior line of therapy subgroups (response-evaluable population) and survival (as-treated population).

Supplementary Figure S1. Patient disposition. Supplementary Figure S2. Association of PD-L1 expression and HPV status with best antitumor response. Supplementary Figure S3. Kaplan-Meier distribution curves for (A) PFS and (B) OS in the as-treated population according to line of treatment and platinum-refractory status, including estimates of medians. Supplementary Figure S4. Peripheral HPV-18-specific T-cells on IFNγ ELISpot assay of PMBCs. Supplementary Figure S5. Peripheral HPV-16 (A, C) E6-specific and (B, D) E7-specific T-cell responses on IFNγ ELISpot assay of PMBCs. Supplementary Figure S6. Peripheral HPV-18 (A, C) E6-specific and (B, D) E7-specific T-cell responses on IFNγ ELISpot assay of PMBCs. Supplementary Figure S7. Baseline peripheral (A) HPV-16-specific and HPV-16 (B) E6-specific and (C) E7-specific T-cell counts on IFNγ ELISpot assay of PMBCs according to best response to treatment. Supplementary Figure S8. Baseline peripheral (A) HPV-18-specific and HPV-18 (B) E6-specific and (C) E7-specific T-cell counts on IFNγ ELISpot assay of PMBCs according to best response to treatment. Supplementary Figure S9. (A) HPV-16-specific and (B) HPV-18-specific T-cell count measured by IFNγ ELISpot assay of PMBCs over time per individual patient (n = 34), color-coded by best response, plotted on a log10 y-axis.

Staphylococcus aureus ST398 is a typical ‘One Health’ pathobiont exemplifying multiple-host tropisms. Here we traced the evolutionary trajectory of the global accessory genome (an assembly of accessory genes) of S. aureus ST398 over 20 years, with the aim of identifying the mechanisms linking accessory genomes with multiple-host tropisms and the phylogenomic traits associated with severe human infections. We analyzed 1079 high-quality genomes of ST398 from 13 host species, spanning 23 years (1998–2021) and 25 countries across 5 continents, and showed that accessory gene pools of ST398 substantially expanded in the early period before 2010, aligning with the increase in the host-species spectrum. The more recent shifts of accessory genomes were mainly driven by stochastic processes. Accessory genes transferred widely across ST398 from different host-species and barely formed host-specific accessory gene pools, indicating that a host-jump of ST398 was followed shortly by another host-switch rather than a long-term co-evolution with a new host species to generate host-specific gene pools. Human-ST398 was a major recipient of accessory gene transfer, with more common gene transfer with ST398 from pig than other animals. Life-threatening exotoxin genes separately encoding Panton-Valentine Leukocidin and the staphylococcal enterotoxin B were abundant and exclusive to human-ST398 that showed a higher evolution rate than animal-ST398. Both accessory and core genome analyses implied nutrient metabolism as a major force for ST398 evolution. Analyses of clinical data revealed a conserved evolution of ST398 along infection development within a patient, and identified a novel subtype ST398-9 (a relatively recent phylogenetic branch) and phages StauST398_5 and StauST398_1 to be closely associated with human infections. Our findings elucidate mechanisms underlying the distribution and evolution of accessory gene pools of ST398, which determine the development of multiple-host tropisms and pathogenicity.

In the phase 3 TACKLE study, outpatient treatment with AZD7442 (tixagevimab/cilgavimab) was well tolerated and significantly reduced progression to severe disease or death through day 29 in adults with mild-to-moderate coronavirus disease 2019 (COVID-19) at the primary analysis. Here, we report data from the final analysis of the TACKLE study, performed after approximately 15 months’ follow-up. Eligible participants were randomized 1:1 and dosed within 7 days of symptom onset with 600 mg intramuscular AZD7442 (n = 456; 300 mg tixagevimab/300 mg cilgavimab) or placebo (n = 454). Severe COVID-19 or death through day 29 occurred in 4.4 https://clinicaltrials.gov/study/NCT04723394 . The body’s immune system produces proteins called antibodies that specifically target foreign substances such as viruses. AZD7442 is a combination of two antibodies (called tixagevimab and cilgavimab) that bind to the severe acute respiratory syndrome coronavirus 2 virus spike protein, preventing it from causing coronavirus disease 2019 (COVID-19). AZD7442 was designed to be “long-acting” and therefore provide prolonged protection against COVID-19 lasting several months from a single dose. It was tested in a clinical trial (TACKLE) to see if it could prevent people who had recently developed symptoms of COVID-19 from getting sicker, being hospitalized, or dying. Around 900 adults took part in this clinical trial. Half of this group were treated with a dose of AZD7442, given as two injections. The other half received a placebo (injections that look like the AZD7442 injections but contain no medicine). The effect of AZD7442 treatment against COVID-19 was monitored over 6 months, and safety was monitored over 15 months. Around the same percentage of participants in the trial reported side effects with AZD7442 and placebo, suggesting there were no safety issues with AZD7442. AZD7442 treatment reduced the risk of participants getting severe COVID-19 or dying from COVID-19 by approximately half, compared with the placebo group. Participants receiving AZD7442 also had fewer hospitalizations due to COVID-19 complications, compared with the placebo group. These results showed the long-term safety of using long-acting antibodies such as AZD7442 as a treatment for COVID-19.

The evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in variants that can escape neutralization by therapeutic antibodies. Here, we describe AZD3152, a SARS-CoV-2–neutralizing monoclonal antibody designed to provide improved potency and coverage against emerging variants. AZD3152 binds to the back left shoulder of the SARS-CoV-2 spike protein receptor binding domain and prevents interaction with the human angiotensin-converting enzyme 2 receptor. AZD3152 potently neutralized a broad panel of pseudovirus variants, including the currently dominant Omicron variant JN.1 but has reduced potency against XBB subvariants containing F456L. In vitro studies confirmed F456L resistance and additionally identified T415I and K458E as escape mutations. In a Syrian hamster challenge model, prophylactic administration of AZD3152 protected hamsters from weight loss and inflammation-related lung pathologies and reduced lung viral load. In the phase 1 sentinel safety cohort of the ongoing SUPERNOVA study ( ClinicalTrials.gov : NCT05648110), a single 600-mg intramuscular injection of AZD5156 (containing 300 mg each of AZD3152 and cilgavimab) was well tolerated in adults through day 91. Observed serum concentrations of AZD3152 through day 91 were similar to those observed with cilgavimab and consistent with predictions for AZD7442, a SARS-CoV-2–neutralizing antibody combination of cilgavimab and tixagevimab, in a population pharmacokinetic model. On the basis of its pharmacokinetic characteristics, AZD3152 is predicted to provide durable protection against symptomatic coronavirus disease 2019 caused by susceptible SARS-CoV-2 variants, such as JN.1, in humans.

Given the continuous emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VoCs), immunotherapeutics that target conserved epitopes on the spike (S) glycoprotein have therapeutic advantages. Here, we report the crystal structure of the SARS-CoV-2 S receptor -binding domain (RBD) at 1.95 A and describe flexibility and distinct conformations of the angiotensin-converting enzyme 2 (ACE2)-binding site. We identify a set of SARS-CoV-2-reactive monoclonal antibodies (mAbs) with broad RBD cross -reactivity including SARS-CoV-2 Omicron subvariants, SARS-CoV-1, and other sarbecoviruses and determine the crystal structures of mAb-RBD complexes with Ab246 and CR3022 mAbs targeting the class IV site, WRAIR-2134, which binds the recently designated class V epitope, and WRAIR-2123, the class I ACE2-binding site. The broad reactivity of class IV and V mAbs to conserved regions of SARS-CoV-2 VoCs and other sarbecovirus provides a framework for long-term immunotherapeutic development strategies.

ABSTRACT AZD7442 is a combination of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-neutralizing antibodies, tixagevimab and cilgavimab, developed for pre-exposure prophylaxis (PrEP) and treatment of coronavirus disease 2019 (COVID-19). Using data from eight clinical trials, we describe a population pharmacokinetic (popPK) model of AZD7442 and show how modeling of “interim” data accelerated decision-making during the COVID-19 pandemic. The final model was a two-compartmental distribution model with first-order absorption and elimination, including standard allometric exponents for the effect of body weight on clearance and volume. Other covariates included were as follows: sex, age >65 years, body mass index ≥30 kg/m 2 , and diabetes on absorption rate; diabetes on clearance; Black race on central volume; and intramuscular (IM) injection site on bioavailability. Simulations indicated that IM injection site and body weight had > 20% effects on AZD7442 exposure, but no covariates were considered to have a clinically relevant impact requiring dose adjustment. The pharmacokinetics of AZD7442, cilgavimab, and tixagevimab were comparable and followed linear kinetics with extended half-lives (median 78.6 days for AZD7442), affording prolonged protection against susceptible SARS-CoV-2 variants. Comparison of popPK simulations based on “interim data” with a target concentration based on 80% viral inhibition and assuming 1.81% partitioning into the nasal lining fluid supported a decision to double the PrEP dosage from 300 mg to 600 mg to prolong protection against Omicron variants. Serum AZD7442 concentrations in adolescents weighing 40–95 kg were predicted to be only marginally different from those observed in adults, supporting authorization for use in adolescents before clinical data were available. In these cases, popPK modeling enabled accelerated clinical decision-making.

Single monoclonal antibodies (mAbs) can be expressed in vivo through gene delivery of their mRNA formulated with lipid nanoparticles (LNPs). However, delivery of a mAb combination could be challenging due to the risk of heavy and light variable chain mispairing. We evaluated the pharmacokinetics of a three mAb combination against Staphylococcus aureus first in single chain variable fragment scFv-Fc and then in immunoglobulin G 1 (IgG1) format in mice. Intravenous delivery of each mRNA/LNP or the trio (1 mg/kg each) induced functional antibody expression after 24 h (10-100 μg/mL) with 64%-78% cognate-chain paired IgG expression after 3 days, and an absence of non-cognate chain pairing for scFv-Fc. We did not observe reduced neutralizing activity for each mAb compared with the level of expression of chain-paired mAbs. Delivery of the trio mRNA protected mice in an S. aureus-induced dermonecrosis model. Intravenous administration of the three mRNA in non-human primates achieved peak serum IgG levels ranging between 2.9 and 13.7 μg/mL with a half-life of 11.8-15.4 days. These results suggest nucleic acid delivery of mAb combinations holds promise and may be a viable option to streamline the development of therapeutic antibodies.

COVID-19 remains a major public health concern. Monoclonal antibodies have received emergency use authorization (EUA) for pre-exposure prophylaxis against COVID-19 among high-risk groups for treatment of mild to moderate COVID-19. In addition to recombinant biologics, engineered synthetic DNA-encoded antibodies (DMAb) are an important strategy for direct in vivo delivery of protective mAb. A DMAb cocktail was synthetically engineered to encode the immunoglobulin heavy and light chains of two different two different Fc-engineered anti-SARS-CoV-2 antibodies. The DMAbs were designed to enhance in vivo expression and delivered intramuscularly to cynomolgus and rhesus macaques with a modified in vivo delivery regimen. Serum levels were detected in macaques, along with specific binding to SARS-CoV-2 spike receptor binding domain protein and neutralization of multiple SARS-CoV-2 variants of concern in pseudovirus and authentic live virus assays. Prophylactic administration was protective in rhesus macaques against signs of SARS-CoV-2 (USA-WA1/2020) associated disease in the lungs. Overall, the data support further study of DNA-encoded antibodies as an additional delivery mode for prevention of COVID-19 severe disease. These data have implications for human translation of gene-encoded mAbs for emerging infectious diseases and low dose mAb delivery against COVID-19.