Safety, immunogenicity, and protection provided by unadjuvanted and adjuvanted formulations of a recombinant plant-derived virus-like particle vaccine candidate for COVID-19 in nonhuman primates.

Although antivirals are important tools to control severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, effective vaccines are essential to control the current coronavirus disease 2019 (COVID-19) pandemic. Plant-derived virus-like particle (VLP) vaccine candidates have previously demonstrated immunogenicity and efficacy against influenza. Here, we report the immunogenicity and protection induced in rhesus macaques by intramuscular injections of a VLP bearing a SARS-CoV-2 spike protein (CoVLP) vaccine candidate formulated with or without Adjuvant System 03 (AS03) or cytidine-phospho-guanosine (CpG) 1018. Although a single dose of the unadjuvanted CoVLP vaccine candidate stimulated humoral and cell-mediated immune responses, booster immunization (at 28 days after priming) and adjuvant administration significantly improved both responses, with higher immunogenicity and protection provided by the AS03-adjuvanted CoVLP. Fifteen micrograms of CoVLP adjuvanted with AS03 induced a polyfunctional interleukin-2 (IL-2)-driven response and IL-4 expression in CD4 T cells. Animals were challenged by multiple routes (i.e., intratracheal, intranasal, and ocular) with a total viral dose of 106 plaque-forming units of SARS-CoV-2. Lower viral replication in nasal swabs and bronchoalveolar lavage fluid (BALF) as well as fewer SARS-CoV-2-infected cells and immune cell infiltrates in the lungs concomitant with reduced levels of proinflammatory cytokines and chemotactic factors in the BALF were observed in animals immunized with the CoVLP adjuvanted with AS03. No clinical, pathologic, or virologic evidence of vaccine-associated enhanced disease was observed in vaccinated animals. The CoVLP adjuvanted with AS03 was therefore selected for vaccine development and clinical trials.

Evaluating the efficacy of extracted squalene from seed oil in the form of microemulsion for the treatment of COVID-19: A clinical study.

This study investigates the effect of the nanostructure of squalene in the form of microemulsion on COVID-19 patients. In this blinded clinical trial, a comparison was made between the efficacy of squalene treatment and controls. A total of 30 COVID-19 patients admitted to the emergency department, and the infection ward was equally allocated to case (n = 15) and control (n = 15) groups according to their age and underlying diseases. The baseline characteristics of subjects, including age, gender, time of treatment onset, underlying condition, white blood cells count, and lymphocyte count were similar (p < 0.05). Baseline laboratory tests and computed tomography (CT) scans were performed for the study groups. The treatment group received 5 mg of intravenous squalene twice a day and standard treatment for 6 days, while controls received only standard treatment. After 6 days of treatment, clinical and CT scan changes were evaluated and compared in intervention and control groups. The need for oxygen therapy (p = 0.020), 2 days of no fever (p = 0.025), cough alleviation (p = 0.010), and lung high-resolution computed tomography improvement (p = 0.033) were significantly different between cases and controls within 7 days of admission. No adverse effects were observed in the treatment group. Our data suggest that squalene could be considered as a potential treatment for COVID-19, and further studies are required to confirm the results.

Reassessment of the risk of narcolepsy in children in England 8 years after receipt of the AS03-adjuvanted H1N1 pandemic vaccine: A case-coverage study.

BACKGROUND: Early studies of narcolepsy after AS03-adjuvanted pandemic A/H1N12009 vaccine (Pandemrix) could not define the duration of elevated risk post-vaccination nor the risk in children aged under 5 years who may not present until much older. METHODS/FINDINGS: Clinical information and sleep test results, extracted from hospital notes at 3 large pediatric sleep centers in England between September 2017 and June 2018 for narcolepsy cases aged 4-19 years with symptom onset since January 2009, were reviewed by an expert panel to confirm the diagnosis. Vaccination histories were independently obtained from general practitioners (GPs). The odds of vaccination in narcolepsy cases compared with the age-matched English population was calculated after adjustment for clinical conditions that were indications for vaccination. GP questionnaires were returned for 242 of the 244 children with confirmed narcolepsy. Of these 5 were under 5 years, 118 were 5-11 years, and 119 were 12-19 years old at diagnosis; 39 were vaccinated with Pandemrix before onset. The odds ratio (OR) for onset at any time after vaccination was 1.94 (95% confidence interval [CI] 1.30-2.89), The elevated risk period was restricted to onsets within 12 months of vaccination (OR 6.65 [3.44-12.85]) and was highest within the first 6 months. After one year, ORs were not significantly different from 1 up to 8 years after vaccination. The ORs were similar in under five-year-olds and older ages. The estimated attributable risk was 1 in 34,500 doses. Our study is limited by including cases from only 3 sleep centers, who may differ from cases diagnosed in nonparticipating centers, and by imprecision in defining the centers' catchment population. The potential for biased recall of onset shortly after vaccination in cases aware of the association cannot be excluded. CONCLUSIONS: In this study, we found that vaccine-attributable cases have onset of narcolepsy within 12 months of Pandemrix vaccination. The attributable risk is higher than previously estimated in England because of identification of vaccine-attributable cases with late diagnoses. Absence of a compensatory drop in risk 1-8 years after vaccination suggests that Pandemrix does not trigger onsets in those in whom narcolepsy would have occurred later.

Safety of AS03-adjuvanted influenza vaccines: A review of the evidence.

Clinical and post-licensure data have demonstrated that AS03-adjuvanted inactivated split virion vaccines, many with reduced antigen content, are effective against influenza infection. The objective of this review is to provide a comprehensive assessment of the safety of trivalent seasonal, monovalent pre-pandemic and pandemic AS03-adjuvanted influenza vaccines, based on non-clinical, clinical and post-licensure data in various populations. Non-clinical studies on local tolerance, toxicology and safety pharmacology did not raise any safety concerns with AS03 administered alone or combined with various influenza antigens. Data from clinical trials with over 55,000 vaccinated subjects showed that AS03-adjuvanted influenza vaccines were generally well tolerated and displayed an acceptable safety profile, although the power to detect rare events was limited. Approximately 90 million doses of A/H1N1pdm09 pandemic influenza vaccines (Pandemrix and Arepanrix H1N1) were administered worldwide, which contributed post-licensure data to the collective safety data for AS03-adjuvanted influenza vaccines. An association between Pandemrix and narcolepsy was observed during the A/H1N1pdm09 pandemic, for which a role of a CD4 T cell mimicry sequence in the haemagglutinin protein of A/H1N1pdm09 cannot be excluded. Provided that future AS03-adjuvanted influenza vaccines do not contain this putative mimicry sequence, this extensive safety experience supports the further development and use of AS03-adjuvanted inactivated split virion candidate vaccines against seasonal and pandemic influenza infections.

Clinical and radiological manifestations of lipoid pneumonia according to etiology: Squalene, omega-3-acid ethyl esters, and idiopathic.

OBJECTIVES: Clinical manifestations of lipoid pneumonia (LP) vary depending on the causative agents or underlying causes. The aim of the present study was to investigate the clinical and radiological features of LP, classified according to etiologic agents. METHODS: The clinico-radiological characteristics of LP patients were retrospectively compared among groups: exogenous versus idiopathic and squalene versus omega-3-acid ethyl esters (O-3-AEE) versus idiopathic. Idiopathic group was defined as LP with no proven or reported etiological evidence. RESULTS: Twenty-two patients met the diagnostic criteria for LP: squalene (9 [41%]), O-3-AEE (6 [27%]), olive oil (1 [5%]), and idiopathic (7 [32%]). Compared with the exogenous group, the idiopathic group showed a higher recurrence rate; higher frequencies of bronchial anthracofibrosis (BAF) and bronchoalveolar lavage (BAL) lymphocytosis; and a higher rate of crazy-paving pattern and lower rate of consolidation on computed tomography scan. In three-group tests, compared with the O-3-AEE group, the squalene group exhibited a significantly higher percentage of neutrophils and a higher rate of right middle lobe (RML) involvement. CONCLUSIONS: In comparison with the exogenous group, the idiopathic group demonstrated BAL lymphocytosis, higher rates of recurrence and BAF, and a higher rate of crazy-paving pattern. Compared with the O-3-AEE group, the squlaene group showed a higher percentage of BAL neutrophils and predominant RML involvement.

Preclinical Pharmacology and Pharmacokinetics of Inhaled Hexadecyl-Treprostinil (C16TR), a Pulmonary Vasodilator Prodrug.

This article describes the preclinical pharmacology and pharmacokinetics (PK) of hexadecyl-treprostinil (C16TR), a prodrug of treprostinil (TRE), formulated in a lipid nanoparticle (LNP) for inhalation as a pulmonary vasodilator. C16TR showed no activity (>10 µM) in receptor binding and enzyme inhibition assays, including binding to prostaglandin E2 receptor 2, prostaglandin D2 receptor 1, prostaglandin I2 receptor, and prostaglandin E2 receptor 4; TRE potently bound to each of these prostanoid receptors. C16TR had no effect (up to 200 nM) on platelet aggregation induced by ADP in rat blood. In hypoxia-challenged rats, inhaled C16TR-LNP produced dose-dependent (0.06-6 µg/kg), sustained pulmonary vasodilation over 3 hours; inhaled TRE (6 µg/kg) was active at earlier times but lost its effect by 3 hours. Single- and multiple-dose PK studies of inhaled C16TR-LNP in rats showed proportionate dose-dependent increases in TRE Cmax and area under the curve (AUC) for both plasma and lung; similar results were observed for dog plasma levels in single-dose PK studies. In both species, inhaled C16TR-LNP yielded prolonged plasma TRE levels and a lower plasma TRE Cmax compared with inhaled TRE. Inhaled C16TR-LNP was well tolerated in rats and dogs; TRE-related side effects included cough, respiratory tract irritation, and emesis and were seen only after high inhaled doses of C16TR-LNP in dogs. In guinea pigs, inhaled TRE (30 µg/ml) consistently produced cough, but C16TR-LNP (30 µg/ml) elicited no effect. These results demonstrate that C16TR-LNP provides long-acting pulmonary vasodilation, is well tolerated in animal studies, and may necessitate less frequent dosing than inhaled TRE with possibly fewer side effects.

Evaluation of a primary course of H9N2 vaccine with or without AS03 adjuvant in adults: A phase I/II randomized trial.

BACKGROUND: Avian influenza A H9N2 strains have pandemic potential. METHODS: In this randomized, observer-blind study (ClinicalTrials.gov: NCT01659086), 420 healthy adults, 18-64years of age, received 1 of 10 H9N2 inactivated split-virus vaccination regimens (30 participants per group), or saline placebo (120 participants). H9N2 groups received 2 doses (days 0, 21) of 15µg hemagglutinin (HA) without adjuvant, or 1.9µgHA+AS03A, 1.9µgHA+AS03B, 3.75µgHA+AS03A, or 3.75µgHA+AS03B; followed by the same H9N2 formulation or placebo (day 182). AS03 is an adjuvant system containing α-tocopherol (AS03A: 11.86mg; AS03B: 5.93mg) and squalene in an oil-in-water emulsion. Immunogenicity (hemagglutination inhibition [HI] and microneutralization assays) and safety were assessed up to day 546. RESULTS: All adjuvanted formulations exceeded regulatory immunogenicity criteria at days 21 and 42 (HI assay), with seroprotection and seroconversion rates of ≥94.9% and ≥89.8% at day 21, and 100% and ≥98.1% at day 42. Immunogenicity criteria were also met for unadjuvanted vaccine, with lower geometric mean titers. In groups administered a third vaccine dose (day 182), an anamnestic immune response was elicited with robust increases in HI and microneutralization titers. Injection site pain was reported more frequently with adjuvanted vaccines. No vaccine-related serious adverse events were observed. CONCLUSIONS: All H9N2 vaccine formulations were immunogenic with a clinically acceptable safety profile; adjuvanted formulations were 4-8 times dose-sparing (3.75-1.9vs 15µgHA). TRIAL REGISTRATION: Registered on ClinicalTrials.gov: NCT01659086.

Adjuvanted (AS03) A/H1N1 2009 Pandemic Influenza Vaccines and Solid Organ Transplant Rejection: Systematic Signal Evaluation and Lessons Learnt.

INTRODUCTION: We investigated a signal of solid organ transplant (SOT) rejection after immunisation with (AS03) A/H1N1 2009 pandemic influenza vaccines. METHODS: Potential immunological mechanisms were reviewed and quantitative analyses were conducted. The feasibility of pharmacoepidemiological studies was explored. RESULTS: Overall results, including data from a pharmacoepidemiological study, support the safety of adjuvanted (AS03) pandemic influenza vaccination in SOT recipients. The regulatory commitment to evaluate the signal through a stepwise investigation was closed in 2014. CONCLUSION: Lessons learned highlight the importance of investigating plausible biological mechanisms between vaccines and potentially associated adverse outcomes, and the importance of selecting appropriate study settings and designs for safety signal investigations.

A systematic review and meta-analysis on the safety of newly adjuvanted vaccines among children.

INTRODUCTION: New adjuvants such as the AS- or the MF59-adjuvants improve vaccine efficacy and facilitate dose-sparing. Their use in influenza and malaria vaccines has resulted in a large body of evidence on their clinical safety in children. METHODS: We carried out a systematic search for safety data from published clinical trials on newly adjuvanted vaccines in children ≤10 years of age. Serious adverse events (SAEs), solicited AEs, unsolicited AEs and AEs of special interest were evaluated for four new adjuvants: the immuno-stimulants containing adjuvant systems AS01 and AS02, and the squalene containing oil-in-water emulsions AS03 and MF59. Relative risks (RR) were calculated, comparing children receiving newly adjuvanted vaccines to children receiving other vaccines with a variety of antigens, both adjuvanted and unadjuvanted. RESULTS: Twenty-nine trials were included in the meta-analysis, encompassing 25,056 children who received at least one dose of the newly adjuvanted vaccines. SAEs did not occur more frequently in adjuvanted groups (RR 0.85, 95%CI 0.75-0.96). Our meta-analyses showed higher reactogenicity following administration of newly adjuvanted vaccines, however, no consistent pattern of solicited AEs was observed across adjuvant systems. Pain was the most prevalent AE, but often mild and of short duration. No increased risks were found for unsolicited AEs, febrile convulsions, potential immune mediated diseases and new onset of chronic diseases. CONCLUSIONS: Our meta-analysis did not show any safety concerns in clinical trials of the newly adjuvanted vaccines in children ≤10 years of age. An unexplained increase of meningitis in one Phase III AS01-adjuvanted malaria trial and the link between narcolepsy and the AS03-adjuvanted pandemic vaccine illustrate that continued safety monitoring is warranted.

Alp Rose stem cells, olive oil squalene and a natural alkyl polyglucoside emulsifier: Are they appropriate ingredients of skin moisturizers - in vivo efficacy on normal and sodium lauryl sulfate - irritated skin?.

Background/Aim: Since skin moisturization may be achieved by both actives and chosen carrier, plant stem cells, squalene and natural alkyl polyglucoside emulsifier may be potential components of contemporary cosmetic products. The aim of the study was in vivo evaluation of the skin irritation potential and the efficacy of Alpine Rose stem cells incorporated into li-posomes and olive oil squalene as ingredients of moisturizing creams, with respect to the novel emulsifier used for creams' stabilization. Methods: With the employment of noninvasive skin biophysical measurements, skin hydration (EC), transepi-dermal water loss (TEWL), erythema index (EI) and viscoelas-ticity were measured on 76 healthy volunteers. In the first phase, skin irritation after a 24-hour occlusion and the long-term efficacy of creams (a 21-day study) on healthy skin were evaluated. Phase II of the study focused on the cream efficacy assessment after a 6-day treatment of sodium lauryl sulfate-irritated skin. Results: After a 24-hour occlusion, there were no significant changes in the EI for any tested sample. In the second phase of the study, the EI was not significantly altered for the cream containing squalene, while the application of all active samples resulted in a significant reduction of TEWL. In both phases of the study an EC increase was recorded, espe-cially for the squalene-containing cream. Conclusion: Due to the lack of skin irritation and skin barrier impairment along with the marked hydration effect, it could be said that the in-vestigated actives incorporated into alkyl polyglucoside emulsi-fier-stabilized creams may be safely applied as ingredients for "tailor-made" cosmetic moisturizers intended for normal and dry skin care, whereas olive oil squalene could be used for the treatment of irritated or sensitive skin as well. [Projekat Ministarstva nauke Republike Srbije, br. TR34031]

Assessment of Prime-boost Vaccination Using an AS03B-adjuvanted Influenza A (H5N1) Vaccine: A Randomized Trial in Children of Three to Less Than Eighteen Years of Age.

BACKGROUND: Heterologous prime-boost vaccination is a pandemic response strategy utilizing subtype-matched vaccine at pandemic onset followed by strain-matched vaccine once available. Persistence of immune response and safety of influenza A (H5N1) vaccine adjuvanted with adjuvant system containing α-tocopherol and squalene in an oil-in-water emulsion (AS03B) were evaluated. METHODS: An open phase 3 active-controlled study (www.clinicaltrials.gov NCT01379937) assessed immunogenicity and reactogenicity of a heterologous booster dose of A/turkey/Turkey/1/2005-H5N1-AS03B in children 3 to <18 years of age, given 6 months after 2-dose priming with A/Indonesia/05/2005-H5N1-AS03B (H5N1(2) -H5N1 group) compared with a single dose of A/turkey/Turkey/1/2005-H5N1-AS03B in unprimed subjects (hepatitis A vaccine (HAV)-H5N1 group). Hemagglutinin inhibition responses and microneutralization antibodies were assessed to 6 months after booster vaccination. RESULTS: Hemagglutinin inhibition antibody responses against A/turkey/Turkey/1/2005-H5N1 were superior in the H5N1(2)-H5N1 versus the hepatitis A vaccine-H5N1 group overall and in each age strata (3 to <10 and 10 to <18 years). Anamnestic immune responses were demonstrated against vaccine-homologous/heterologous strains in the H5N1(2)-H5N1 group. Injection site pain and fever increased with consecutive doses for children <6 years (H5N1(2)-H5N1). Immune responses to vaccine-homologous/heterologous strains persisted to 6 months after booster vaccination in the H5N1(2)-H5N1 group. CONCLUSIONS: Heterologous H5N1-AS03B-adjuvanted booster vaccination in children/adolescents was immunogenic for vaccine-homologous and heterologous strains following 2-dose priming, with immune persistence for at least 6 months. Prime-boost strategies using H5N1-AS03 could be effectively employed in this age group.

Perinatal survival and health after maternal influenza A(H1N1)pdm09 vaccination: A cohort study of pregnancies stratified by trimester of vaccination.

Large cohort studies demonstrated the safety of vaccination with the AS03 adjuvanted pandemic influenza vaccine, but data on first trimester vaccination safety are limited. We conducted a nationwide register-based retrospective cohort study in Finland, included singleton pregnancies present on 01 November 2009 and followed them from 01 November 2009 until delivery. Pregnancies with abortive outcome, pregnancies that started before 01 February 2009 and pregnancies of women, who received the AS03 adjuvanted pandemic influenza vaccine prior to the onset of pregnancy, were excluded. Our main outcome measures were hazard ratios comparing the risk of stillbirth, early neonatal death, moderately preterm birth, very preterm birth, moderately low birth weight, very low birth weight, and being small for gestational age between pregnancies exposed and unexposed to maternal influenza A(H1N1)pdm09 vaccination. The study population comprised 43,604 pregnancies; 34,241 (78.5%) women were vaccinated at some stage during pregnancy. The rates of stillbirth, early neonatal death, moderately preterm birth, and moderately low birth weight were similar between pregnant women exposed and unexposed to influenza A(H1N1)pdm09 vaccination. After adjusting for known risk factors, the relative rates were 0.90 (95% confidence interval 0.55-1.45) for very preterm birth, 0.84 (0.61-1.16) for very low birth weight, and 1.17 (0.98-1.40) for being small for gestational age. Also, in the subanalysis of 7839 women vaccinated during the first trimester, the rates did not indicate that maternal vaccination during the first trimester had any adverse impact on perinatal survival and health. The risk of adverse pregnancy outcomes was not associated with the exposure to the AS03 adjuvanted pandemic influenza vaccine. This study adds reassuring evidence on the safety of AS03 adjuvanted influenza vaccines when given in the first trimester and supports the recommendation of influenza vaccination to all pregnant women through all stages of pregnancy.

Safety and persistence of the humoral and cellular immune responses induced by 2 doses of an AS03-adjuvanted A(H1N1)pdm09 pandemic influenza vaccine administered to infants, children and adolescents: Two open, uncontrolled studies.

In children, 2 AS03-adjuvanted A(H1N1)pdm09 vaccine doses given 21 days apart were previously shown to induce a high humoral immune response and to have an acceptable safety profile up to 42 days following the first vaccination. Here, we analyzed the persistence data from 2 open-label studies, which assessed the safety, and humoral and cell-mediated immune responses induced by 2 doses of this vaccine. The first study was a phase II, randomized trial conducted in 104 children aged 6-35 months vaccinated with the A(H1N1)pdm09 vaccine containing 1.9 µg haemagglutinin antigen (HA) and AS03B (5.93 mg tocopherol) and the second study, a phase III, non-randomized trial conducted in 210 children and adolescents aged 3-17 years vaccinated with the A(H1N1)pdm09 vaccine containing 3.75 µg HA and AS03A (11.86 mg tocopherol). Approximately one year after the first dose, all children with available data were seropositive for haemagglutinin inhibition and neutralising antibody titres, but a decline in geometric mean antibody titres was noted. The vaccine induced a cell-mediated immune response in terms of antigen-specific CD4(+) T-cells, which persisted up to one year post-vaccination. The vaccine did not raise any safety concern, though these trials were not designed to detect rare events. In conclusion, 2 doses of the AS03-adjuvanted A(H1N1)pdm09 vaccine at 2 different dosages had a clinically acceptable safety profile, and induced high and persistent humoral and cell-mediated immune responses in children aged 6-35 months and 3-17 years. These studies have been registered at www.clinicaltrials.gov NCT00971321 and NCT00964158.

Paresthesia and sensory disturbances associated with 2009 pandemic vaccine receipt: Clinical features and risk factors.

BACKGROUND: Paresthesia was the third-most-common adverse event following immunization (AEFI) with 2009 monovalent AS03-adjuvanted A(H1N1)pdm09 vaccine in Quebec, Canada and was also frequently reported in Europe. This study assessed clinical features and risk factors associated with this unexpected AEFI. METHODS: Reports to the passive surveillance system were summarized. A case-control study was conducted to assess risk factors and additional investigations were undertaken among cases with symptoms persisting ≥12 months. RESULTS: There were 328 reports of paresthesia affecting the vaccinated arm (58%), but also face (45%), lower limbs (40%) and back/thorax (23%) with numbness but also muscle weakness (61%), motor impairment (61%), generalized myalgia (37%), visual (14%) and/or speech effects (15%). Reporting rate was highest in women of reproductive age, peaking at 30-39 years-old (28/100,000 doses administered) and exceeding that of men of the same age (7/100,000 doses) by 4-fold. Median time to onset was 2h. Symptoms subsided within one week in 37% but lasted ≥6 months in 26%. No consistent or objective neurological findings were identified. Risk was increased with allergy history, respiratory illness the day of vaccination, depressive symptoms and family history of pulmonary disease, but decreased with physical activity the day of vaccination, and regular weekly alcohol consumption. CONCLUSION: Paresthesia following 2009 pandemic vaccine receipt lasted several weeks and included other motor-sensory disturbances in an important subset of patients. Although it does not correspond with known neurological disease, and causality remains uncertain, further investigation is warranted to understand the nature and frequency of paresthesia as a possible AEFI with influenza vaccines.

Immunogenicity and Safety of an EB66 Cell-Culture-Derived Influenza A/Indonesia/5/2005(H5N1) AS03-Adjuvanted Vaccine: A Phase 1 Randomized Trial.

BACKGROUND: Cell-culture-derived (CC) influenza vaccine production methods could provide benefits over classical embryonated-egg technology, including a higher production capacity and the faster creation of a supply that meets demand. METHODS: A CC-inactivated split-virus influenza A/Indonesia/5/2005(H5N1) vaccine derived from the EB66 cell line (hereafter, "CC-H5N1") was investigated in a phase 1 randomized, blinded study. Healthy adults (n = 521) received 2 vaccine doses (days 0 and 21) of either investigational CC-H5N1 vaccine (1.9 µg or 3.75 µg of hemagglutinin antigen [HA] with the AS03 adjuvant system or 15 µg of plain HA), embryonated-egg-derived vaccines (3.75 µg of HA with AS03 or 15 µg of plain HA), or placebo. Assessment of the adjuvant effect and immunogenicity was performed using Center for Biologics Evaluation and Research acceptability criteria 21 days after dose 2. Safety was assessed until month 12. RESULTS: AS03-adjuvanted CC-H5N1 elicited a homologous hemagglutination inhibition antibody response that satisfied immunogenicity criteria 21 days after dose 2 and persisted at month 12. Adjuvant effect and immune response against a drift-variant strain were demonstrated. No vaccine-related serious adverse events were reported. The immunogenicity and safety of the CC-H5N1 formulation containing 3.75 µg of HA and AS03 appeared to be similar to those for the licensed egg-derived AS03-adjuvanted control vaccine. CONCLUSIONS: The feasibility of the EB66 cell line to produce an immunogenic influenza vaccine with acceptable safety profile was demonstrated. Antigen sparing was achieved through combination with AS03 adjuvant. This CC-H5N1 might contribute to the rapid access of vaccine in the event of an influenza A(H5N1) pandemic. CLINICAL TRIALS REGISTRATION: NCT01236040.

A first-generation physiologically based pharmacokinetic (PBPK) model of alpha-tocopherol in human influenza vaccine adjuvant.

Alpha (α)-tocopherol is a component of a new generation of squalene-containing oil-in-water (SQ/W) emulsion adjuvants that have been licensed for use in certain influenza vaccines. Since regulatory pharmacokinetic studies are not routinely required for influenza vaccines, the in vivo fate of this vaccine constituent is largely unknown. In this study, we constructed a physiologically based pharmacokinetic (PBPK) model for emulsified α-tocopherol in human adults and infants. An independent sheep PBPK model was also developed to inform the local preferential lymphatic transfer and for the purpose of model evaluation. The PBPK model predicts that α-tocopherol will be removed from the injection site within 24h and rapidly transfer predominantly into draining lymph nodes. A much lower concentration of α-tocopherol was estimated to peak in plasma within 8h. Any systemically absorbed α-tocopherol was predicted to accumulate slowly in adipose tissue, but not in other tissues. Model evaluation and uncertainty analyses indicated acceptable fit, with the fraction of dose taken up into the lymphatics as most influential on plasma concentration. In summary, this study estimates the in vivo fate of α-tocopherol in adjuvanted influenza vaccine, may be relevant in explaining its immunodynamics in humans, and informs current regulatory risk-benefit analyses.

AS03B-adjuvanted H5N1 influenza vaccine in children 6 months through 17 years of age: a phase 2/3 randomized, placebo-controlled, observer-blinded trial.

BACKGROUND: This phase 2/3, randomized, placebo-controlled, observer-blinded study assessed the immunogenicity, reactogenicity, and safety of an inactivated, split-virion H5N1 influenza vaccine (A/Indonesia/5/2005) in children aged 6 months through 17 years. METHODS: Children received 2 influenza vaccine doses 21 days apart, each containing 1.9 µg of hemagglutinin and AS03B adjuvant (5.93 mg of α-tocopherol). The randomization ratio was 8:3 for vaccine to placebo, with equal allocation between 3 age strata (6-35 months, 3-8 years, and 9-17 years). Immunogenicity against the vaccine strain was assessed 21 days after the first and second vaccine doses for all vaccinees, at day 182 for half, and at day 385 for the remaining half. Reactogenicity after each dose and safety up to 1 year after vaccination were evaluated. RESULTS: Within each age stratum, the lower limit of the 98.3% confidence interval for the day 42 seroprotection rate was ≥70%, thus fulfilling the US and European licensure criteria. The immune responses elicited by vaccine persisted well above baseline levels for 1 year. The vaccine was more reactogenic than placebo, but no major safety concerns were identified. CONCLUSIONS: AS03B-adjuvanted H5N1 influenza vaccine was immunogenic and showed an acceptable safety profile in all age groups studied. Clinical Trials Registration: NCT01310413.

Safety of AS03-adjuvanted inactivated split virion A(H1N1)pdm09 and H5N1 influenza virus vaccines administered to adults: pooled analysis of 28 clinical trials.

Clinical trials have shown that AS03-adjuvanted H5N1 and A(H1N1)pdm09 vaccines are highly immunogenic, although with an increased reactogenicity profile relative to non-adjuvanted vaccines in terms of the incidence of common injection site and systemic adverse events (AEs). We evaluated pooled safety data from 22,521 adults who had received an AS03-adjuvanted H5N1 or A(H1N1)pdm09 influenza or control vaccine with the purpose to identify medically-attended AEs (MAEs), including subsets of serious AEs (SAEs), potentially immune-mediated diseases (pIMDs), and AEs of special interest (AESI), and to explore a potential association of these AEs with the administration of an AS03-adjuvanted influenza vaccine. For participants who had received an AS03-adjuvanted vaccine, the relative risks (RRs) for experiencing a MAE or a SAE compared to control group (participants who had received a non-adjuvanted vaccine or saline placebo) were 1.0 (95% confidence interval [CI]: 0.9; 1.1) and 1.1 (95% CI: 0.9; 1.4), respectively. The overall RRs for experiencing an AESI or a pIMD (AS03-adjuvanted vaccine/control) were 1.2 (95% CI: 0.9; 1.6) and 1.7 (95% CI: 0.8; 3.8), respectively. Thirty-8 participants in the AS03-adjuvanted vaccine group had a pIMD reported after vaccine administration, yielding an incidence rate (IR) of 351.9 (95% CI: 249.1; 483.1) per 100,000 person-years. The estimated IRs in the AS03-adjuvanted vaccine group were greater than the literature reported rates for: facial paresis/VIIth nerve paralysis, celiac disease, thrombocytopenia and ulcerative colitis. These results do not support an association between AS03-adjuvanted H5N1 and A(H1N1)pdm09 vaccines and the AEs collected in the trials included in the analysis.

A phase 1, open-label safety and immunogenicity study of an AS03-adjuvanted trivalent inactivated influenza vaccine in children aged 6 to 35 months.

BACKGROUND:  There is a need for better vaccines and vaccine strategies to reduce the burden of influenza in very young children.   METHODS:  This phase 1, open-label study assessed the reactogenicity, safety, and immunogenicity of an inactivated trivalent influenza vaccine (TIV) containing low doses of hemagglutinin antigen (7.5 µg each strain), adjuvanted with a tocopherol-based oil-in-water emulsion Adjuvant System (AS03). Influenza vaccine-naïve children aged 6-35 months were sequentially enrolled to receive TIV-AS03D (1.48 mg tocopherol) or TIV-AS03C (2.97 mg tocopherol), then a 6-month booster of conventional TIV. The primary endpoint was the incidence of fever (axillary temperature >38 °C) for 7 days post-vaccination. Immune responses were assessed by hemagglutination-inhibition (HI) assay. RESULTS:  Forty children were sequentially enrolled into the TIV-AS03D or the TIV-AS03C group. Fever >38.0 °C was reported in 5/20 (25.0%) and 7/20 (35.0%) children after the first and second doses of TIV-AS03D, respectively, and in 7/20 (35.0%) children after 1 dose of TIV-AS03C; the latter fulfilled the holding rule for safety, and the second dose of TIV-AS03C was cancelled. HI immune responses exceeded adult European licensure criteria for the immunogenicity, and all children had HI antibody titers ≥ 1:40 after 1 dose of TIV booster against booster strains. CONCLUSIONS:  One dose of primary vaccine containing a low dose of antigen and AS03 may be a possible influenza vaccination strategy for young children. The relatively high frequency of fever warrants further investigation, although the generalizability of the findings are uncertain given that many of the children had antibody evidence suggesting recent infection with A(H1N1)pdm09.

Long-term outcome of the humoral and cellular immune response of an H5N1 adjuvanted influenza vaccine in elderly persons: 2-year follow-up of a randomised open-label study.

BACKGROUND: Older individuals often have a reduced immune response to influenza vaccination, which might be improved by administering a higher vaccine dose. We compared the immune response to two single doses of the AS03A-adjuvanted H5N1 pandemic vaccine (3.75 µg hemagglutinin of A/Vietnam/1194/2004) with that of two double vaccine doses (7.5 µg hemagglutinin) in adults aged ≥61 years. Here we report the 2-year persistence of the humoral and cellular immune response. METHODS: In this phase II, open-label study, healthy participants aged 61 to 88 years (median 68 years) were randomised (3:1:3:1) to receive two single doses of the AS03A-adjuvanted vaccine (1xH5N1-AS) or the non-adjuvanted vaccine (1xH5N1), or two double doses of the AS03A-adjuvanted vaccine (2xH5N1-AS) or the non-adjuvanted vaccine (2xH5N1), 21 days apart. Serum haemagglutination inhibition antibodies and cellular immune responses against A/Vietnam/1194/2004 were measured in all groups at months 12 and 24; neutralising antibodies were assessed in a subset of the adjuvanted groups. Serious adverse events and adverse events of specific interest were recorded. RESULTS: At month 24, haemagglutination inhibition antibody seroprotection rates were 37.2% (95% CI 27.0% to 48.3%) for 1xH5N1-AS, 30.9% (95% CI 21.1% to 42.1%) for 2xH5N1-AS, 16.2% (95% CI 6.2% to 32.0%) for 1xH5N1, and 8.3% (95% CI 1.0% to 27.0%) for 2xH5N1. Haemagglutination inhibition antibody geometric mean titres were 17.6 (95% CI 13.7 to 22.5) for 1xH5N1-AS, 18.4 (95% CI 14.2 to 23.8) for 2xH5N1-AS, 12.3 (95% CI 8.9 to 16.9) for 1xH5N1 and 9.8 (95% CI 6.7 to 14.4) for 2xH5N1. The median frequency of antigen-specific CD4+ T cells per 106 T cells (25th quartile; 75th quartile) was 852 (482; 1477) for 1xH5N1-AS, 1147 (662; 1698) for 2xH5N1-AS, 556 (343; 749) for 1x-H5N1 and 673 (465; 1497) for 2xH5N1. Neutralising antibody geometric mean titres were 391.0 (95% CI 295.5 to 517.5) in the 1xH5N1-AS group and 382.8 (95% CI 317.4 to 461.6) in the 2xH5N1-AS group. CONCLUSIONS: Antibody levels declined substantially in all groups. Seroprotection rates, geometric mean titres for haemagglutination inhibition antibodies, and CD4+ T-cell responses tended to be higher in the AS03A-adjuvanted groups. There was no clear benefit, in terms of long-term persistence of the immune response, of doubling the dose of the adjuvanted vaccine. No safety concern was observed up to 24 months post-primary vaccination. TRIAL REGISTRATION: NCT00397215 (7 November 2006).