Validation of a severe acute respiratory syndrome coronavirus 2 microneutralization assay for evaluation of vaccine immunogenicity.

As variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continue to emerge, assessment of vaccine immunogenicity remains a critical factor to support continued vaccination. To this end, an in vitro microneutralization (MN50) assay was validated to quantitate SARS-CoV-2 neutralizing antibodies against prototype and variant strains (Beta, Delta, Omicron BA.1, Omicron BA.5, and XBB.1.5) in human serum. For the prototype strain, the MN50 assay met acceptance criteria for inter-/intra-assay precision, specificity, linearity, and selectivity. The assay was robust against changes to virus/serum incubation time, cell seeding density, virus content per well, cell passage number, and serum interference. Analyte in serum samples was stable up to five freeze/thaw cycles and for up to 12 months of storage at -80 ± 10 °C. Similar results were observed for the variant-adapted MN50 assays. The conversion factor to convert assay result units to WHO international standard units (IU/mL) was determined to be 0.62 for the prototype strain. This MN50 assay will be useful for vaccine immunogenicity analyses in clinical trial samples, enabling assessment of vaccine immunogenicity for ancestral and variant strains as variant-adapted vaccines are developed.

A Pseudovirus-Based Neutralization Assay for SARS-CoV-2 Variants: A Rapid, Cost-Effective, BSL-2-Based High-Throughput Assay Useful for Vaccine Immunogenicity Evaluation.

Neutralizing antibody responses from COVID-19 vaccines are pivotal in conferring protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Effective COVID-19 vaccines and assays measuring neutralizing antibodies against emerging variants (i.e., XBB.1.5, XBB.1.16, and XBB.2.3) are needed. The use of biosafety level (BSL)-3 laboratories for live virus assays results in higher costs and a longer turnaround time; therefore, a BSL-2-based pseudovirus neutralization assay (PNT) was developed. The pseudoviruses were produced by cotransfecting cells with plasmids encoding a lentiviral backbone-expressing luciferase reporter; non-surface proteins for lentiviral production; and ancestral or Omicron (BA.1 and BA.5) SARS-CoV-2 spike (S) proteins. The PNT was developed and optimized in dose and kinetics experiments. The representative serum samples (COVID-19-convalescent or NVX-CoV2373-vaccinated participants enrolled in the 2019nCoV-101 trial) demonstrated a wide dynamic range. The neutralization data showed robust correlation with validated anti-recombinant spike IgG levels and angiotensin-converting enzyme 2 inhibition titers (ancestral). This assay is suitable for measurement of the neutralization ability in clinical samples from individuals infected with SARS-CoV-2 or immunized with a COVID-19 vaccine. The results suggest that this PNT provides a lower cost, high-throughput, rapid turnaround alternative to BSL-3-based microneutralization assays and enables the discovery and development of effective vaccines against emerging variants.

Immunogenicity and safety of a bivalent (omicron BA.5 plus ancestral) SARS-CoV-2 recombinant spike protein vaccine as a heterologous booster dose: interim analysis of a phase 3, non-inferiority, randomised, clinical trial.

BACKGROUND: SARS-CoV-2 variants evade immunity despite vaccination with prototype COVID-19 vaccines or previous infection. The 2019nCoV-311 (part 2) study is evaluating immune responses after two booster doses of a vaccine containing the omicron BA.5 subvariant spike protein in adults previously vaccinated with a prototype mRNA vaccine. This interim analysis reports on day 28 immunogenicity and safety outcomes after one booster dose. METHODS: In this phase 3, randomised, observer-blinded study conducted at 35 sites in Australia, medically stable, previously COVID-19-vaccinated (mRNA-based; ≥three doses) adults aged 18 years or older were enrolled and randomly allocated (1:1:1; via an interactive web response system) to receive two doses of bivalent (NVX-CoV2373 + NVX-CoV2540; bivalent group), authorised prototype (NVX-CoV2373; prototype group), or BA.5 (NVX-CoV2540; BA.5 group) vaccine. Only blinded personnel performed study assessments or had participant contact to collect data after study vaccination. Participants received vaccines containing 5 µg SARS-CoV-2 recombinant spike protein and 50 µg Matrix-M adjuvant, administered via a 0·5 mL intramuscular injection (2·5 µg of NVX-CoV2373 plus 2·5 µg of NVX-CoV2540 for the bivalent vaccine, prepared on-site as a 1:1 mixture). The coprimary endpoints include day 28 neutralising antibody geometric mean titre (GMT) ratios (GMTRs) to omicron BA.5 and the ancestral strain, and seroresponse rates to BA.5, in the bivalent and prototype groups. These endpoints were calculated in the per-protocol analysis set, which was defined as participants who had received a vaccine dose, had baseline and day 28 immunogenicity data, and were PCR-negative for SARS-CoV-2, with no major protocol deviations. The primary objective was to determine the primary outcome (antibody responses), which consisted of three comparisons: superiority of the bivalent versus prototype vaccine for neutralising antibody GMT to BA.5 (ie, lower bound of the GMTR 95% CI >1·0); non-inferiority of neutralising antibody seroresponse rate to BA.5 (ie, lower bound of the seroresponse rate 95% CI >-5%); and non-inferiority of neutralising antibody GMT to the ancestral strain (ie, lower bound of GMTR 95% CI >0·67). This trial was registered at ClinicalTrials.gov, number NCT05372588. FINDINGS: Between March 22, 2023 and May 2, 2023, 837 participants were screened for eligibility and 766 were randomly allocated to receive the BA.5 (n=255), prototype (n=252), or bivalent (n=259) vaccine. After accounting for exclusions due to participants being baseline SARS-CoV-2-positive, having previous infection, or protocol deviations, the per-protocol analysis set included 694 participants (236 in BA.5 group, 227 in prototype group, and 231 in bivalent group). In this interim analysis (maximum follow-up 35 days after the first dose), the bivalent group, compared with the prototype group, had superior neutralising antibody responses to BA.5 (GMT 1017·8 [95% CI 891·0-1162·6] vs 515·1 [450·4-589·0]; GMTR 2·0 [1·69-2·33]) and a non-inferior seroresponse rate to BA.5 at day 28 (39·8% [33·5-46·5] vs 12·3% [8·4-17·3]; difference 27·5% [19·8-35·0]). The bivalent group also had non-inferior neutralising antibody responses to the ancestral strain (GMTR 1·0 [0·84-1·20]), compared with the prototype group. All vaccines were similarly well tolerated. INTERPRETATION: All three coprimary endpoints were met in part 2 of the ongoing 2019nCoV-311 study. These data support the development of monovalent and/or bivalent vaccines for the most currently circulating variants, to optimise protection. With no new safety findings, further investigation of omicron-based subvariant vaccines is supported by the evidence. FUNDING: Novavax.

Altered IgG4 Antibody Response to Repeated mRNA versus Protein COVID Vaccines.

Repeated mRNA SARS-CoV-2 vaccination has been associated with increases in the proportion of IgG4 in spike-specific antibody responses and concurrent reductions in Fcγ-mediated effector functions that may limit control of viral infection. Here, we assessed anti-Spike total IgG, IgG1, IgG2, IgG3 and IgG4, and surrogate markers for antibody-dependent cellular phagocytosis (ADCP, FcγRIIa binding), antibody-dependent cellular cytotoxicity (ADCC, FcγRIIIa binding), and antibody-dependent complement deposition (ADCD, C1q binding) associated with repeated SARS-CoV-2 vaccination with NVX-CoV2373 (Novavax Inc., Gaithersburg, MD). The NVX-CoV2373 protein vaccine did not induce notable increases in spike-specific IgG4 or negatively impact surrogates for Fcγ effector responses. Conversely, repeated NVX-CoV2373 vaccination uniquely enhanced IgG3 responses which are known to exhibit strong affinity for FcγRIIIa and have previously been linked to potent neutralization of SARS-CoV-2. Subsequent investigations will help to understand the immunological diversity generated by different SARS-CoV-2 vaccine types and have the potential to reshape public health strategies.

Immunogenicity and safety of heterologous Omicron BA.1 and bivalent SARS-CoV-2 recombinant spike protein booster vaccines: a phase 3, randomized, clinical trial.

BACKGROUND: Mutations present in emerging SARS-CoV-2 variants permit evasion of neutralization with prototype vaccines. A novel Omicron BA.1 subvariant-specific vaccine (NVX-CoV2515) was tested alone, or as a bivalent preparation in combination with the prototype vaccine (NVX-CoV2373), to assess antibody responses to SARS-CoV-2. METHODS: Participants aged 18 to 64 years immunized with 3 doses of prototype mRNA vaccines were randomized 1:1:1 to receive a single dose of NVX-CoV2515, NVX-CoV2373, or bivalent mixture in a phase 3 study investigating heterologous boosting with SARS-CoV-2 recombinant spike protein vaccines. Immunogenicity was measured 14 and 28 days after vaccination for the SARS-CoV-2 Omicron BA.1 sublineage and ancestral strain. Safety profiles of vaccines were assessed. RESULTS: Of participants who received trial vaccine (N = 829), those administered NVX-CoV2515 (n = 286) demonstrated superior neutralizing antibody response to BA.1 versus NVX-CoV2373 (n = 274) at Day 14 (geometric mean titer ratio [95% CI]: 1.6 [1.33, 2.03]). Seroresponse rates [n/N; 95% CI] were 73.4% [91/124; 64.7, 80.9] for NVX-CoV2515 versus 50.9% [59/116; 41.4, 60.3] for NVX-CoV2373. All formulations were similarly well-tolerated. CONCLUSIONS: NVX-CoV2515 elicited a superior neutralizing antibody response against the Omicron BA.1 subvariant compared with NVX-CoV2373 when administered as a fourth dose. Safety data were consistent with the established safety profile of NVX-CoV2373.

A Phase 3, randomized, non-inferiority study of a heterologous booster dose of SARS CoV-2 recombinant spike protein vaccine in adults.

Due to waning immunity following primary immunization with COVID-19 vaccines, booster doses may be required. The present study assessed a heterologous booster of SII-NVX-CoV2373 (spike protein vaccine) in adults primed with viral vector and inactivated vaccines. In this Phase 3, observer-blind, randomized, active controlled study, a total of 372 adults primed with two doses of ChAdOx1 nCoV-19 (n = 186) or BBV152 (n = 186) at least six months ago, were randomized to receive a booster of SII-NVX-CoV2373 or control vaccine (homologous booster of ChAdOx1 nCoV-19 or BBV152). Anti-S IgG and neutralizing antibodies (nAbs) were assessed at days 1, 29, and 181. Non-inferiority (NI) of SII-NVX-CoV2373 to the control vaccine was assessed based on the ratio of geometric mean ELISA units (GMEU) of anti-S IgG and geometric mean titers (GMT) of nAbs (NI margin > 0.67) as well as seroresponse (≥ 2 fold-rise in titers) (NI margin -10%) at day 29. Safety was assessed throughout the study period. In both the ChAdOx1 nCoV-19 prime and BBV152 prime cohorts, 186 participants each received the study vaccines. In the ChAdOx1 nCoV-19 prime cohort, the GMEU ratio was 2.05 (95% CI 1.73, 2.43) and the GMT ratio was 1.89 (95% CI 1.55, 2.32) whereas the difference in the proportion of seroresponse was 49.32% (95% CI 36.49, 60.45) for anti-S IgG and 15% (95% CI 5.65, 25.05) for nAbs on day 29. In the BBV152 prime cohort, the GMEU ratio was 5.12 (95% CI 4.20, 6.24) and the GMT ratio was 4.80 (95% CI 3.76, 6.12) whereas the difference in the proportion of seroresponse was 74.08% (95% CI 63.24, 82.17) for anti-S IgG and 24.71% (95% CI 16.26, 34.62) for nAbs on day 29. The non-inferiority of SII-NVX-CoV2373 booster to the control vaccine for each prime cohort was met. SII-NVX-CoV2373 booster showed significantly higher immune responses than BBV152 homologous booster. On day 181, seroresponse rates were ≥ 70% in all the groups for both nAbs and anti-S IgG. Solicited adverse events reported were transient and mostly mild in severity in all the groups. No causally related SAE was reported. SII-NVX-CoV2373 as a heterologous booster induced non-inferior immune responses as compared to homologous boosters in adults primed with ChAdOx1 nCoV-19 and BBV152. SII-NVX-CoV2373 showed a numerically higher boosting effect than homologous boosters. The vaccine was also safe and well tolerated.

Immunogenicity and safety of NVX-CoV2373 as a booster: A phase 3 randomized clinical trial in adults.

BACKGROUND: To combat the SARS-CoV-2 pandemic, multiple vaccines using different manufacturing platforms have been developed, including NVX-CoV2373 (an adjuvanted recombinant protein vaccine). As SARS-CoV-2 variants have emerged, some of which evade vaccine-induced immunity, introduction of vaccine booster doses has become critical. Employing different vaccine types for primary series vaccination and boosting could expand vaccine coverage and access. This study assessed whether NVX-CoV2373 would induce robust responses when used as a booster. METHODS: The 2019nCoV-307 study was a phase 3, randomized, observer-blinded trial evaluating immunogenicity and safety of NVX-CoV2373 in previously vaccinated adults aged 18-49 years in the United States (NCT05463068). Participants were randomized 1:1:1 to receive one intramuscular injection of NVX-CoV2373 from one of three different manufacturing lots. Immunogenicity was assessed by immunoglobulin G (IgG) and neutralizing antibodies (NAb). These responses were compared for the three lots, and for participants with primary series with or without a prior booster dose of the mRNA-1273, BNT162b2, Ad26.COV2.S, or NVX-CoV2373 COVID-19 vaccines. RESULTS: A total of 911 participants were randomized between July 11 and 13, 2022, with 905 being assessed for safety and 848 for immunogenicity. Immunogenicity of NVX-CoV2373 met prespecified equivalence criteria between lots, and the booster dose was well-tolerated. NVX-CoV2373 induced robust IgG and NAb responses when used as a first or later booster dose, regardless of primary series vaccine type. Seroconversion rates were also similar across previous vaccine types. Induced antibodies were strongly reactive, even to the immune-evasive Omicron BA.1 and BA.5 variants. CONCLUSIONS: NVX-CoV2373 showed consistent immunogenicity between lots, with no new safety signals identified. Use of NVX-CoV2373 as a booster dose (first or later) is supported.

A Severe Acute Respiratory Syndrome Coronavirus 2 Anti-Spike Immunoglobulin G Assay: A Robust Method for Evaluation of Vaccine Immunogenicity Using an Established Correlate of Protection.

As the COVID-19 pandemic continues, variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continue to emerge. Immunogenicity evaluation of vaccines and identification of correlates of protection for vaccine effectiveness is critical to aid the development of vaccines against emerging variants. Anti-recombinant spike (rS) protein immunoglobulin G (IgG) quantitation in the systemic circulation (serum/plasma) is shown to correlate with vaccine efficacy. Thus, an enzyme-linked immunosorbent assay (ELISA)-based binding assay to detect SARS-CoV-2 (ancestral and variant strains) anti-rS IgG in human serum samples was developed and validated. This assay successfully met acceptance criteria for inter/intra-assay precision, specificity, selectivity, linearity, lower/upper limits of quantitation, matrix effects, and assay robustness. The analyte in serum was stable for up to 8 freeze/thaw cycles and 2 years in -80 °C storage. Similar results were observed for the Beta, Delta, and Omicron BA.1/BA.5/XBB.1.5 variant-adapted assays. Anti-rS IgG assay results correlated significantly with neutralization and receptor binding inhibition assays. In addition, usage of international reference standards allows data extrapolation to WHO international units (BAU/mL), facilitating comparison of results with other IgG assays. This anti-rS IgG assay is a robust, high-throughput method to evaluate binding IgG responses to S protein in serum, enabling rapid development of effective vaccines against emerging COVID-19 variants.

Safety and Immunogenicity of SARS-CoV-2 Recombinant Spike Protein Vaccine in Children and Adolescents in India: A Phase 2-3 Randomized Clinical Trial.

Importance: The recombinant COVID-19 vaccine NVX-CoV2373 has demonstrated efficacy of approximately 90% in adults; however, its safety and efficacy in children is unknown. Objective: To assess the noninferiority of SII-NVX-CoV2373 in children and adolescents compared to adults and to evaluate its safety in comparison with placebo. Design, Setting, and Participants: This phase 2-3 observer-blind randomized clinical trial was conducted in 2 cohorts, children (aged 2 to 11 years) and adolescents (aged 12 to 17 years) between August 2021 and August 2022. Participants were randomized 3:1 to SII-NVX-CoV2373 or placebo and monitored for 179 days. The participants, study team, and laboratory staff were blinded. This was a multicenter study conducted across 10 tertiary care hospitals in India. Exclusion criteria included previous COVID-19 infection or vaccination, immunocompromised condition, and immunosuppressive medications. Interventions: Two doses of 0.5-mL SII-NVX-CoV2373 or placebo were administered intramuscularly on days 1 and 22. Main Outcomes and Measures: Primary outcomes were geometric mean titer ratio of both anti-spike (anti-S) IgG and neutralizing antibodies (NAbs) between both pediatric age groups to that of adults on day 36. Noninferiority was concluded if the lower bound of 95% CI of this ratio was greater than 0.67 for each age group. Both the antibodies were assessed for the index strain and for selected variants at various time points. Solicited adverse events (AEs) were recorded for 7 days after each vaccination, unsolicited AEs were recorded for 35 days, and serious AEs and AEs of special interest were recorded for 179 days. Results: A total of 460 children in each age cohort were randomized to receive vaccine or placebo. The mean (SD) age was 6.7 (2.7) years in the child cohort and 14.3 (1.6) years in the adolescent cohort; 231 participants (50.2%) in the child cohort and 218 in the adolescent cohort (47.4%) were female. Both anti-S IgG and NAb titers were markedly higher in the SII-NVX-CoV2373 group than in the placebo group on both day 36 and day 180. The geometric mean titer ratios compared to those in adults were 1.20 (95% CI, 1.08-1.34) and 1.52 (95% CI, 1.38-1.67) for anti-S IgG in adolescents and children, respectively; while for NAbs, they were 1.33 (95% CI, 1.17-1.50) and 1.93 (95% CI, 1.70-2.18) in adolescents and children, respectively, indicating noninferiority. SII-NVX-CoV2373 also showed immune responses against variants studied. Injection site reactions, fever, headache, malaise, and fatigue were common solicited AEs. There were no AEs of special interest and no causally related serious AEs. Conclusions and Relevance: SII-NVX-CoV2373 was safe and well tolerated in children and adolescents in this study. The vaccine was highly immunogenic and may be used in pediatric vaccination against COVID-19. Trial Registration: Clinical Trials Registry of India Identifier: CTRI/2021/02/031554.

Immunogenicity and safety of a fourth homologous dose of NVX-CoV2373.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has significantly reduced the efficacy of some approved vaccines. A fourth dose of NVX-CoV2373 (5 µg SARS-CoV-2 recombinant spike [rS] protein + 50 µg Matrix-M™ adjuvant; Novavax, Gaithersburg, MD) was evaluated to determine induction of cross-reactive antibodies to variants of concern. A phase II randomized study (NCT04368988) recruited participants in Australia and the United States to assess a primary series of NVX-CoV2373 followed by two booster doses (third and fourth doses at 6-month intervals) in adults 18-84 years of age. The primary series was administered when the SARS-CoV-2 ancestral strain was prevalent and the third and fourth doses while the Alpha and Delta variants were prevalent in AUS and US. Local/systemic reactogenicity was assessed the day of vaccination and for 6 days thereafter. Unsolicited adverse events (AEs) were reported. Immunogenicity was measured before, and 14 days after, fourth dose administration, using anti-spike serum immunoglobulin G (IgG) and neutralization assays against ancestral SARS-CoV-2 strain and Omicron sublineages. Among 1283 enrolled participants, 258 were randomized to receive the two-dose primary series, of whom 104 received a third dose, and 45 received a fourth dose of NVX-CoV2373. The incidence of local/systemic reactogenicity events increased after the first three doses of NVX-CoV2373 and leveled off after dose 4. Unsolicited AEs were reported in 9 % of participants after dose 4 (none of which were severe or serious). Anti-rS IgG levels and neutralization antibody titers increased following booster doses to a level approximately four-fold higher than that observed after the primary series, with a progressively narrowed gap in response between the ancestral strain and Omicron BA.5. A fourth dose of NVX-CoV2373 enhanced immunogenicity for ancestral and variant SARS-CoV-2 strains without increasing reactogenicity, indicating that updates to the vaccine composition may not be currently warranted.

Immunogenicity of NVX-CoV2373 in PREVENT-19: A Phase 3, Randomized, Placebo-Controlled Trial in Adults in the United States and Mexico.

Background: NVX-CoV2373, an adjuvanted, recombinant SARS-CoV-2 spike (rS) protein vaccine, consistently demonstrated protective efficacy against COVID-19 in clinical trials and has received regulatory authorizations or approvals worldwide. Methods: PREVENT-19 (NCT04611802) is a phase 3, randomized, observer-blinded, placebo-controlled trial evaluating safety, immunogenicity, and efficacy of NVX-CoV2373 in ≈30 000 participants ≥18 years in the United States and Mexico. Vaccine humoral immune response (ie, serum immunoglobulin [IgG] antibodies, hACE2 receptor binding inhibition antibodies, and neutralizing antibodies to SARS-CoV-2) (ancestral strain) was assessed in 1200 participants randomly selected and equally divided between participants 18-64 and ≥65 years. Results: In the per protocol analysis, NVX-CoV2373 induced vigorous serum antibody responses among the 1063 analyzed participants who were SARS-CoV-2 seronegative at baseline, received both doses of study treatment, and had serology results available 2 weeks after dose 2. Geometric mean (GM) responses in both younger and older adults were higher among recipients of vaccine versus placebo for IgG (64 259 vs 121 and 37 750 vs 133 ELISA units, respectively), hACE2 receptor binding inhibition GM titers (GMTs) (222 vs 5 and 136 vs 5, respectively), and neutralizing antibody GMTs (1303 vs 11 and 900 vs 11, respectively). Humoral responses were 30-40% lower in participants ≥65 years or HIV-positive; however, seroconversion rates were high and comparable between the age cohorts, regardless of HIV serostatus. Conclusions: NVX-CoV2373 elicited robust humoral immune responses against ancestral SARS-CoV-2 virus 2 weeks following the second vaccination in adult PREVENT-19 participants, consistent with previously reported high vaccine efficacy.

Safety, Immunogenicity, and Efficacy of the NVX-CoV2373 COVID-19 Vaccine in Adolescents: A Randomized Clinical Trial.

Importance: Greater than 20% of cases and 0.4% of deaths from COVID-19 occur in children. Following demonstration of the safety and efficacy of the adjuvanted, recombinant spike protein vaccine NVX-CoV2373 in adults, the PREVENT-19 trial immediately expanded to adolescents. Objective: To evaluate the safety, immunogenicity, and efficacy of NVX-CoV2373 in adolescents. Design, Setting, and Participants: The NVX-CoV2373 vaccine was evaluated in adolescents aged 12 to 17 years in an expansion of PREVENT-19, a phase 3, randomized, observer-blinded, placebo-controlled multicenter clinical trial in the US. Participants were enrolled from April 26 to June 5, 2021, and the study is ongoing. A blinded crossover was implemented after 2 months of safety follow-up to offer active vaccine to all participants. Key exclusion criteria included known previous laboratory-confirmed SARS-CoV-2 infection or known immunosuppression. Of 2304 participants assessed for eligibility, 57 were excluded and 2247 were randomized. Interventions: Participants were randomized 2:1 to 2 intramuscular injections of NVX-CoV2373 or placebo, 21 days apart. Main Outcomes and Measures: Serologic noninferiority of neutralizing antibody responses compared with those in young adults (aged 18-25 years) in PREVENT-19, protective efficacy against laboratory-confirmed COVID-19, and assessment of reactogenicity and safety. Results: Among 2232 participants (1487 NVX-CoV2373 and 745 placebo recipients), the mean (SD) age was 13.8 (1.4) years, 1172 (52.5%) were male, 1660 (74.4%) were White individuals, and 359 (16.1%) had had a previous SARS-CoV-2 infection at baseline. After vaccination, the ratio of neutralizing antibody geometric mean titers in adolescents compared with those in young adults was 1.5 (95% CI, 1.3-1.7). Twenty mild COVID-19 cases occurred after a median of 64 (IQR, 57-69) days of follow-up, including 6 among NVX-CoV2373 recipients (incidence, 2.90 [95% CI, 1.31-6.46] cases per 100 person-years) and 14 among placebo recipients (incidence, 14.20 [95% CI, 8.42-23.93] cases per 100 person-years), yielding a vaccine efficacy of 79.5% (95% CI, 46.8%-92.1%). Vaccine efficacy for the Delta variant (the only viral variant identified by sequencing [n = 11]) was 82.0% (95% CI, 32.4%-95.2%). Reactogenicity was largely mild to moderate and transient, with a trend toward greater frequency after the second dose of NVX-CoV2373. Serious adverse events were rare and balanced between treatments. No adverse events led to study discontinuation. Conclusions and Relevance: The findings of this randomized clinical trial indicate that NVX-CoV2373 is safe, immunogenic, and efficacious in preventing COVID-19, including the predominant Delta variant, in adolescents. Trial Registration: ClinicalTrials.gov Identifier: NCT04611802.

Severe Acute Respiratory Syndrome Coronavirus 2 Receptor (Human Angiotensin-Converting Enzyme 2) Binding Inhibition Assay: A Rapid, High-Throughput Assay Useful for Vaccine Immunogenicity Evaluation.

Emerging variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) show immune evasion of vaccine-derived immunity, highlighting the need for better clinical immunogenicity biomarkers. To address this need, an enzyme-linked immunosorbent assay-based, human angiotensin-converting enzyme 2 (hACE2) binding inhibition assay was developed to measure antibodies against the ancestral strain of SARS-CoV-2 and was validated for precision, specificity, linearity, and other parameters. This assay measures the inhibition of SARS-CoV-2 spike (S) protein binding to the receptor, hACE2, by serum from vaccine clinical trials. Inter- and intra-assay precision, specificity, linearity, lower limit of quantitation, and assay robustness parameters successfully met the acceptance criteria. Heme and lipid matrix effects showed minimal interference on the assay. Samples were stable for testing in the assay even with 8 freeze/thaws and up to 24 months in -80 °C storage. The assay was also adapted for variants (Delta and Omicron BA.1/BA.5), with similar validation results. The hACE2 assay showed significant correlation with anti-recombinant S immunoglobulin G levels and neutralizing antibody titers. This assay provides a rapid, high-throughput option to evaluate vaccine immunogenicity. Along with other clinical biomarkers, it can provide valuable insights into immune evasion and correlates of protection and enable vaccine development against emerging COVID-19 variants.

Safety, Immunogenicity and Efficacy of NVX-CoV2373 in Adolescents in PREVENT-19: A Randomized, Phase 3 Trial.

BACKGROUND: Over 20% of cases and 0.4% of deaths from Covid-19 occur in children. Following demonstration of safety and efficacy of the adjuvanted, recombinant spike protein vaccine NVX-CoV2373 in adults, the PREVENT-19 trial enrolled adolescents. METHODS: Safety, immunogenicity, and efficacy of NVX-CoV2373 were evaluated in adolescents aged 12 to <18 years in an expansion of PREVENT-19, a phase 3, randomized, observer-blinded, placebo-controlled trial in the United States. Participants were randomized 2:1 to two doses of NVX-CoV2373 or placebo 21 days apart, and followed for a median of 2 months after second vaccination. Primary end points were serologic non-inferiority of neutralizing antibody (NA) responses compared with young adults (18 to <26 years) in PREVENT-19, protective efficacy against laboratory-confirmed Covid-19, and assessment of reactogenicity/safety. RESULTS: Among 2,247 participants randomized between April-June 2021, 1,491 were allocated to NVX-CoV2373 and 756 to placebo. Post-vaccination, the ratio of NA geometric mean titers in adolescents compared to young adults was 1.5 (95% confidence interval [CI] 1.3 to 1.7). Twenty Covid-19 cases (all mild) occurred: 6 among NVX-CoV2373 and 14 among placebo recipients (vaccine efficacy [VE]: 79.5%, 95% CI, 46.8 to 92.1). All sequenced viral genomes (11/20) were identified as Delta variant (Delta variant VE: 82.0% [95% CI: 32.4 to 95.2]). Reactogenicity was largely mild-to-moderate, transient, and more frequent in NVX-CoV2373 recipients and after the second dose. Serious adverse events were rare and evenly distributed between treatments. CONCLUSIONS: NVX-CoV2373 was safe, immunogenic, and efficacious in the prevention of Covid-19 and those cases caused by the Delta variant in adolescents. (Funded by the Office of the Assistant Secretary for Preparedness and Response, Biomedical Advanced Research and Development Authority and the National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health; PREVENT-19 ClinicalTrials.gov number, NCT04611802 ).

NVX-CoV2373 vaccination induces functional SARS-CoV-2-specific CD4+ and CD8+ T cell responses.

NVX-CoV2373 is an adjuvanted recombinant full-length SARS-CoV-2 spike trimer protein vaccine demonstrated to be protective against COVID-19 in efficacy trials. Here we demonstrate that vaccinated individuals made CD4+ T cell responses after 1 and 2 doses of NVX-CoV2373, and a subset of individuals made CD8+ T cell responses. Characterization of the vaccine-elicited CD8+ T cells demonstrated IFN-γ production. Characterization of the vaccine-elicited CD4+ T cells revealed both circulating T follicular helper (cTfh) cells and Th1 cells (IFN-γ+, TNF-α+, and IL-2+) were detectable within 7 days of the primary immunization. Spike-specific CD4+ T cells were correlated with the magnitude of the later SARS-CoV-2-neutralizing antibody titers, indicating that robust generation of CD4+ T cells, capable of supporting humoral immune responses, may be a key characteristic of NVX-CoV2373 that utilizes Matrix-M adjuvant.

Safety and immunogenicity following a homologous booster dose of a SARS-CoV-2 recombinant spike protein vaccine (NVX-CoV2373): a secondary analysis of a randomised, placebo-controlled, phase 2 trial.

BACKGROUND: Emerging SARS-CoV-2 variants and evidence of waning vaccine efficacy present substantial obstacles towards controlling the COVID-19 pandemic. Booster doses of SARS-CoV-2 vaccines might address these concerns by amplifying and broadening the immune responses seen with initial vaccination regimens. We aimed to assess the immunogenicity and safety of a homologous booster dose of a SARS-CoV-2 recombinant spike protein vaccine (NVX-CoV2373). METHODS: This secondary analysis of a phase 2, randomised study assessed a single booster dose of a SARS-CoV-2 recombinant spike protein vaccine with Matrix-M adjuvant (NVX-CoV2373) in healthy adults aged 18-84 years, recruited from 17 clinical centres in the USA and Australia. Eligible participants had a BMI of 17-35 kg/m2 and, for women, were heterosexually inactive or using contraception. Participants who had a history of SARS-CoV or SARS-CoV-2, confirmed diagnosis of COVID-19, serious chronic medical conditions, or were pregnant or breastfeeding were excluded. Approximately 6 months following their primary two-dose vaccination series (administered day 0 and day 21), participants who received placebo for their primary vaccination series received a placebo booster (group A) and participants who received NVX-CoV2373 for their primary vaccination series (group B) were randomly assigned (1:1) again, via centralised interactive response technology system, to receive either placebo (group B1) or a single booster dose of NVX-CoV2373 (5 µg SARS-CoV-2 rS with 50 µg Matrix-M adjuvant; group B2) via intramuscular injection; randomisation was stratified by age and study site. Vaccinations were administered by designated site personnel who were masked to treatment assignment, and participants and other site staff were also masked. Administration personnel also assessed the outcome. The primary endpoints are safety (unsolicited adverse events) and reactogenicity (solicited local and systemic) events and immunogenicity (serum IgG antibody concentrations for the SARS-CoV-2 rS protein antigen) assessed 14 days after the primary vaccination series (day 35) and 28 days following booster (day 217). Safety was analysed in all participants in groups A, B1, and B2, according to the treatment received; immunogenicity was analysed in the per-protocol population (ie, participants in groups A, B1, and B2) who received all assigned doses and who did not test SARS-CoV-2-positive or received an authorised vaccine, analysed according to treatment assignment). This trial is registered with ClinicalTrials.gov, NCT04368988. FINDINGS: 1610 participants were screened from Aug 24, 2020, to Sept 25, 2020. 1282 participants were enrolled, of whom 173 were assigned again to placebo (group A), 106 were re-randomised to NVX-CoV2373-placebo (group B1), and 104 were re-randomised to NVX-CoV2373-NVX-CoV2373 (group B2); after accounting for exclusions and incorrect administration, 172 participants in group A, 102 in group B1, and 105 in group B2 were analysed for safety. Following the active booster, the proportion of participants with available data reporting local (80 [82%] of 97 participants had any adverse event; 13 [13%] had a grade ≥3 event) and systemic (75 [77%] of 98 participants had any adverse event; 15 [15%] had a grade ≥3 event) reactions was higher than after primary vaccination (175 [70%] of 250 participants had any local adverse event, 13 [5%] had a grade ≥3 event; 132 [53%] of 250 had any systemic adverse event, 14 [6%] had a grade ≥3 event). Local and systemic events were transient in nature (median duration 1·0-2·5 days). In the per-protocol immunogenicity population at day 217 (167 participants in group A, 101 participants in group B1, 101 participants in group B2), IgG geometric mean titres (GMT) had increased by 4·7-fold and MN50 GMT by 4·1-fold for the ancestral SARS-CoV-2 strain compared with the day 35 titres. INTERPRETATION: Administration of a booster dose of NVX-CoV2373 resulted in an incremental increase in reactogenicity. For both the prototype strain and all variants evaluated, immune responses following the booster were similar to or higher than those associated with high levels of efficacy in phase 3 studies of the vaccine. These data support the use of NVX-CoV2373 in booster programmes. FUNDING: Novavax and the Coalition for Epidemic Preparedness Innovations.

Immunogenicity and safety of a SARS-CoV-2 recombinant spike protein nanoparticle vaccine in people living with and without HIV-1 infection: a randomised, controlled, phase 2A/2B trial.

BACKGROUND: There is a paucity of data on COVID-19 vaccines in people living with HIV-1, who could be at increased risk of severe illness and death from COVID-19. We evaluated the safety and immunogenicity of a Matrix-M adjuvanted recombinant spike protein nanoparticle COVID-19 vaccine (NVX-CoV2373; Novavax) in HIV-negative people and people living with HIV-1. METHODS: In this randomised, observer-blinded, multicentre, placebo-controlled phase 2A/B trial in South Africa, participants aged 18-84 years, with and without underlying HIV-1, were enrolled from 16 sites and randomly assigned (1:1) to receive two intramuscular injections of NVX-CoV2373 or placebo, 21 days apart. People living with HIV-1 were on stable antiretroviral therapy and had an HIV-1 viral load of less than 1000 copies per mL. Vaccine dosage was 5 µg SARS-CoV-2 recombinant spike protein with 50 µg Matrix-M adjuvant, whereas 0·9% saline was used as placebo injection (volume 0·5 mL each). All study staff and participants remained masked to study group assignment. We previously reported an interim analysis on the efficacy and safety of the NVX-CoV2373 vaccine (coprimary endpoints). In this Article, we present an expanded safety analysis for the full cohort of participants and report on the secondary objective of vaccine immunogenicity in the full cohort of people living with HIV-1 and in HIV-negative individuals overall and stratified by baseline SARS-CoV-2 serostatus. This trial is registered with ClinicalTrials.gov, NCT04533399, and the Pan-African Clinical Trials Registry, PACTR202009726132275. FINDINGS: Participants were enrolled between Aug 17 and Nov 25, 2020. The safety analysis set included 4164 HIV-negative participants (2089 in the intervention group and 2075 in the placebo group) and 244 people living with HIV-1 (122 in the intervention group and 122 in the placebo group). 1422 (34·1%) of 4164 HIV-negative people and 83 (34·0%) of 244 people living with HIV-1 were categorised as baseline SARS-CoV-2-positive (ie, anti-spike IgG reactive at enrolment or had a reactive SARS-CoV-2 nucleic acid amplification test by 14 days after the second study vaccination). In the NVX-CoV2373 group, solicited local and systemic adverse events were more common in HIV-negative participants (427 [30·6%] local and 401 [28·7%] systemic) than in people living with HIV-1 (20 [25·3%] local and 20 [25·3%] systemic) among those who were baseline SARS-CoV-2-seronegative (naive). Of the serious adverse events that occurred among HIV-negative people (of whom, two [0·1%] were baseline SARS-CoV-2-negative and four [0·6%] were baseline SARS-CoV-2-positive) and people living with HIV-1 (for whom there were no serious adverse events) in the NVX-CoV2373 group, none were assessed as related to the vaccine. Among participants who were baseline SARS-CoV-2-negative in the NVX-CoV2373 group, the anti-spike IgG geometric mean titres (GMTs) and seroconversion rates (SCRs) were lower in people living with HIV-1 (n=62) than in HIV-negative people (n=1234) following the first vaccination (GMT: 508·6 vs 1195·3 ELISA units [EU]/mL; SCR: 51·6% vs 81·3%); and similarly so 14 days after the second vaccination for GMTs (14 420·5 vs 31 631·8 EU/mL), whereas the SCR was similar at this point (100·0% vs 99·3%). In the NVX-CoV2373 group, anti-spike IgG GMTs 14 days after the second vaccination were substantially higher in those who were baseline SARS-CoV-2-positive than in those who were baseline SARS-CoV-2-seronegative for HIV-negative participants (100 666·1 vs 31 631·8 EU/mL) and for people living with HIV-1 (98 399·5 vs 14 420·5 EU/mL). This was also the case for angiotensin-converting enzyme 2 receptor-binding antibody and neutralising antibody titres. INTERPRETATION: The safety of the NVX-CoV2373 vaccine in people living with HIV-1 was similar to that in HIV-negative participants. However, people living with HIV-1 not previously exposed to SARS-CoV-2 had attenuated humoral immune responses to NVX-CoV2373 compared with their HIV-negative vaccine counterparts, but not so if they were baseline SARS-CoV-2-positive. FUNDING: Novavax and the Bill & Melinda Gates Foundation; investigational vaccine manufacturing support was provided by the Coalition for Epidemic Preparedness Innovations.

Comparison of the safety and immunogenicity of a novel Matrix-M-adjuvanted nanoparticle influenza vaccine with a quadrivalent seasonal influenza vaccine in older adults: a phase 3 randomised controlled trial.

BACKGROUND: Improved seasonal influenza vaccines for older adults that can induce broadly cross-reactive antibodies and enhanced T-cell responses, particularly against A H3N2 viruses, while avoiding egg-adaptive antigenic changes, are needed. We aimed to show that the Matrix-M-adjuvanted quadrivalent nanoparticle influenza vaccine (qNIV) was immunologically non-inferior to a licensed, standard-dose quadrivalent inactivated influenza vaccine (IIV4) in older adults. METHODS: This was a phase 3 randomised, observer-blinded, active-comparator controlled trial done across 19 US community-based clinical research sites during the 2019-20 influenza season. Participants were clinically stable and community-dwelling, aged at least 65 years, and were randomised in a 1:1 ratio using an interactive web response system to receive a single intramuscular dose of qNIV or IIV4. The primary objective was to describe safety and show that qNIV was immunologically non-inferior to IIV4. The primary outcomes were adverse events by treatment group and comparative haemagglutination-inhibiting antibody responses (assayed with egg-propagated virus) on day 28, summarised in terms of the ratio of geometric mean titres (GMTRqNIV/IIV4) and seroconversion rate (SCR) difference between participants receiving qNIV or IIV4 for all four vaccine homologous influenza strains. The immunogenicity outcome was measured in the per-protocol population. Non-inferiority was shown if the lower bound of the two-sided 95% CI on the GMTRqNIV/IIV4 was at least 0·67 and the lower bound of the two-sided 95% CI on the SCR difference -was at least -10%. The study is registered with clinicaltrials.gov, NCT04120194, and is active and not recruiting. FINDINGS: 2742 adults were assessed for eligibility and 2654 were enrolled and randomised between Oct 14, 2019, and Oct 25, 2019; 1333 participants were randomised to the qNIV group and 1319 to the IIV4 group (two participants withdrew consent before being assigned to a group). qNIV showed immunological non-inferiority to IIV4: GMTRqNIV/IIV4 for the four vaccine homologous influenza strains was A/Brisbane 1·09 (95% CI 1·03 to 1·15), A/Kansas 1·19 (1·11 to 1·27), B/Maryland 1·03 (0·99 to 1·07), and B/Phuket 1·23 (1·16 to 1·29); and SCR difference was A/Brisbane 5·0 (95% CI 1·9 to 8·1), A/Kansas 7·3 (3·6 to 11·1), B/Maryland 0·5 (-1·9 to 2·9), and B/Phuket 8·5 (5·0 to 11·9). 659 (49·4%) of 1333 of participants in the qNIV group and 551 (41·8%) of 1319 participants in the IIV4 group had at least one treatment-emergent adverse event. More solicited adverse events were reported by participants in the qNIV group (551 [41·3%] of 1333) than in the IIV4 group (420 [31·8%] of 1319), and were comprised primarily of mild to moderate transient injection site pain (341 [25·6%] in the qNIV group vs 212 [16·1%] in the IIV4 group). INTERPRETATION: qNIV was well tolerated and produced qualitatively and quantitatively enhanced humoral and cellular immune response in older adults compared with IIV4. qNIV might enhance the effectiveness of seasonal influenza vaccination, and future studies to show clinical efficacy are planned. FUNDING: Novavax.