A pan-influenza monoclonal antibody neutralizes H5 strains and prophylactically protects through intranasal administration.

Avian A(H5N1) influenza virus poses an elevated zoonotic threat to humans, and no pharmacological products are currently registered for fast-acting pre-exposure protection in case of spillover leading to a pandemic. Here, we show that an epitope on the stem domain of H5 hemagglutinin is highly conserved and that the human monoclonal antibody CR9114, targeting that epitope, potently neutralizes all pseudotyped H5 viruses tested, even in the rare case of substitutions in its epitope. Further, intranasal administration of CR9114 fully protects mice against A(H5N1) infection at low dosages, irrespective of pre-existing immunity conferred by the quadrivalent seasonal influenza vaccine. These data provide a proof-of-concept for broad, pre-exposure protection against a potential future pandemic using the intranasal administration route. Studies in humans should assess if autonomous administration of a broadly-neutralizing monoclonal antibody is safe and effective and can thus contribute to pandemic preparedness.

Genome-wide profiling of circulatory microRNAs associated with cognition and dementia.

INTRODUCTION: MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression. Their role in the pathophysiology of dementia and potential as biomarkers remains undetermined. METHODS: We conducted a single- (one-by-one) and multi-marker (joint) analysis to identify well-expressed circulating miRNAs in plasma (total = 591) associated with general cognition and incident dementia, for 1615 participants of the population-based Rotterdam Study. RESULTS: During single-marker analysis, 47 miRNAs were nominally (P ≤ .05) associated with cognition and 18 miRNAs were nominally associated with incident dementia, after adjustment for potential confounders. Three miRNAs were common between cognition and dementia (miR-4539, miR-372-3p, and miR-566), with multi-marker analysis revealing another common miRNA (miR-7106-5p). In silico analysis of these four common miRNAs led to several putative target genes expressed in the brain, highlighting the mitogen-activated protein kinase signaling pathway. DISCUSSION: We provide population-based evidence on the relationship between circulatory miRNAs with cognition and dementia, including four common miRNAs that may elucidate downstream mechanisms. HIGHLIGHTS: MicroRNAs (miRNAs) are involved in the (dys)function of the central nervous system. Four circulating miRNAs in plasma are associated with cognition and incident dementia. Several predicted target genes of these four miRNAs are expressed in the brain. These four miRNAs may be linked to pathways underlying dementia. Although miRNAs are promising biomarkers, experimental validation remains essential.

Biological Aging for Risk Prediction of First-Ever Intracerebral Hemorrhage and Cerebral Infarction in Advanced Age.

BACKGROUND AND OBJECTIVES: successful interventions to prevent cerebrovascular disease and stroke require early identification of persons at risk before clinical manifestation of disease. The literature remains to be sparse on accessible plasma-based biomarkers for monitoring brain health and cerebrovascular disease in advanced age. We assessed the predictive value of biological age (BA) as an early indicator for cerebrovascular disease and risk of first-ever intracerebral hemorrhage (ICH) and cerebral infarction (CI) in advanced age and compared these relationships with chronological age (CA) and commonly used biomarkers including tau and Aß40 and Aß42. METHODS: The study included Individuals who consented for blood draw and follow-up. We computed biological age using structural equation modelling. The criteria for the biomarkers included their representability of the various body systems; their availability in the Rotterdam study and their pre-hypothesized reflection of aging in other populations. The algorithm integrates biomarkers that represent six body systems involved in overall cerebrovascular health including metabolic function, cardiac function, lung function, kidney function, liver function, immunity, and inflammation. Time to event analysis was conducted using Cox-regression models. Prediction analysis was conducted using Harrel's C and Area under the receiver operating characteristic curve. RESULTS: The sample included a total of 1699 individuals at baseline followed up over a median of 11 years. During a period of 15, 780 and 16, 172 person-years, a total of 17 first-ever intracerebral hemorrhage and 83 cerebral infarction cases occurred. In time-to-event analysis, BA showed higher magnitude of associations with ICH compared to CA (HRBA-ICH: 2.30, 95% CI: 1.20, 4.30; HRCA-ICH: 1.40, 95% CI: 0.76, 2.53) and higher precision with CI (HRBA-CI: 1.30, 95% CI: 1.01,1.75; HRCA-CI:1.90, 95% CI: 1.48, 2.66). BA outperformed CA for prediction of ICH (AUC: 0.68 vs 0.53; Harrel's C: 0.72 vs 0.53) and for CI (AUC:0.63 vs 0.62; Harrel's C: 0.68 vs 0.67). CONCLUSIONS: Biological aging (delta biological aging) based on integrated physiology biomarkers provides a novel tool for monitoring and identification of persons at highest risk of cerebrovascular disease in advanced age with varying degrees of precision and magnitude for stroke subtypes. These variations are likely related to differences in pathophysiology of intracerebral hemorrhage and cerebral infarction. Wider validation and applicability require extension of these findings in other comparable samples and in clinical settings.

Plasma tau, neurofilament light chain and amyloid-ß levels and risk of dementia; a population-based cohort study.

CSF biomarkers, including total-tau, neurofilament light chain (NfL) and amyloid-ß, are increasingly being used to define and stage Alzheimer's disease. These biomarkers can be measured more quickly and less invasively in plasma and may provide important information for early diagnosis of Alzheimer's disease. We used stored plasma samples and clinical data obtained from 4444 non-demented participants in the Rotterdam study at baseline (between 2002 and 2005) and during follow-up until January 2016. Plasma concentrations of total-tau, NfL, amyloid-ß40 and amyloid-ß42 were measured using the Simoa NF-light® and N3PA assays. Associations between biomarker plasma levels and incident all-cause and Alzheimer's disease dementia during follow-up were assessed using Cox proportional-hazard regression models adjusted for age, sex, education, cardiovascular risk factors and APOE ε4 status. Moreover, biomarker plasma levels and rates of change over time of participants who developed Alzheimer's disease dementia during follow-up were compared with age and sex-matched dementia-free control subjects. During up to 14 years follow-up, 549 participants developed dementia, including 374 cases with Alzheimer's disease dementia. A log2 higher baseline amyloid-ß42 plasma level was associated with a lower risk of developing all-cause or Alzheimer's disease dementia, adjusted hazard ratio (HR) 0.61 [95% confidence interval (CI), 0.47-0.78; P < 0.0001] and 0.59 (95% CI, 0.43-0.79; P = 0.0006), respectively. Conversely, a log2 higher baseline plasma NfL level was associated with a higher risk of all-cause dementia [adjusted HR 1.59 (95% CI, 1.38-1.83); P < 0.0001] or Alzheimer's disease [adjusted HR 1.50 (95% CI, 1.26-1.78); P < 0.0001]. Combining the lowest quartile group of amyloid-ß42 with the highest of NfL resulted in a stronger association with all-cause dementia [adjusted HR 9.5 (95% CI, 2.3-40.4); P < 0.002] and with Alzheimer's disease [adjusted HR 15.7 (95% CI, 2.1-117.4); P < 0.0001], compared to the highest quartile group of amyloid-ß42 and lowest of NfL. Total-tau and amyloid-ß40 levels were not associated with all-cause or Alzheimer's disease dementia risk. Trajectory analyses of biomarkers revealed that mean NfL plasma levels increased 3.4 times faster in participants who developed Alzheimer's disease compared to those who remained dementia-free (P < 0.0001), plasma values for cases diverged from controls 9.6 years before Alzheimer's disease diagnosis. Amyloid-ß42 levels began to decrease in Alzheimer's disease cases a few years before diagnosis, although the decline did not reach significance compared to dementia-free participants. In conclusion, our study shows that low amyloid-ß42 and high NfL plasma levels are each independently and in combination strongly associated with risk of all-cause and Alzheimer's disease dementia. These data indicate that plasma NfL and amyloid-ß42 levels can be used to assess the risk of developing dementia in a non-demented population. Plasma NfL levels, although not specific, may also be useful in monitoring progression of Alzheimer's disease dementia.

Multi-Omics Analysis Reveals MicroRNAs Associated With Cardiometabolic Traits.

MicroRNAs (miRNAs) are non-coding RNA molecules that regulate gene expression. Extensive research has explored the role of miRNAs in the risk for type 2 diabetes (T2D) and coronary heart disease (CHD) using single-omics data, but much less by leveraging population-based omics data. Here we aimed to conduct a multi-omics analysis to identify miRNAs associated with cardiometabolic risk factors and diseases. First, we used publicly available summary statistics from large-scale genome-wide association studies to find genetic variants in miRNA-related sequences associated with various cardiometabolic traits, including lipid and obesity-related traits, glycemic indices, blood pressure, and disease prevalence of T2D and CHD. Then, we used DNA methylation and miRNA expression data from participants of the Rotterdam Study to further investigate the link between associated miRNAs and cardiometabolic traits. After correcting for multiple testing, 180 genetic variants annotated to 67 independent miRNAs were associated with the studied traits. Alterations in DNA methylation levels of CpG sites annotated to 38 of these miRNAs were associated with the same trait(s). Moreover, we found that plasma expression levels of 8 of the 67 identified miRNAs were also associated with the same trait. Integrating the results of different omics data showed miR-10b-5p, miR-148a-3p, miR-125b-5p, and miR-100-5p to be strongly linked to lipid traits. Collectively, our multi-omics analysis revealed multiple miRNAs that could be considered as potential biomarkers for early diagnosis and progression of cardiometabolic diseases.

Quantification of biological age as a determinant of age-related diseases in the Rotterdam Study: a structural equation modeling approach.

Chronological age alone is not a sufficient measure of the true physiological state of the body. The aims of the present study were to: (1) quantify biological age based on a physiological biomarker composite model; (2) and evaluate its association with death and age-related disease onset in the setting of an elderly population. Using structural equation modeling we computed biological age for 1699 individuals recruited from the first and second waves of the Rotterdam study. The algorithm included nine physiological parameters (c-reactive protein, creatinine, albumin, total cholesterol, cytomegalovirus optical density, urea nitrogen, alkaline phosphatase, forced expiratory volume and systolic blood pressure). We assessed the association between biological age, all-cause mortality, all-cause morbidity and specific age-related diseases over a median follow-up of 11 years. Biological age, compared to chronological age or the traditional biomarkers of age-related diseases, showed a stronger association with all-cause mortality (HR 1.15 vs. 1.13 and 1.10), all-cause morbidity (HR 1.06 vs. 1.05 and 1.03), stroke (HR 1.17 vs. 1.08 and 1.04), cancer (HR 1.07 vs. 1.04 and 1.02) and diabetes mellitus (HR 1.12 vs. 1.01 and 0.98). Individuals who were biologically younger exhibited a healthier life-style as reflected in their lower BMI (P < 0.001) and lower incidence of stroke (P < 0.001), cancer (P < 0.01) and diabetes mellitus (P = 0.02). Collectively, our findings suggest that biological age based on the biomarker composite model of nine physiological parameters is a useful construct to assess individuals 65 years and older at increased risk for specific age-related diseases.

A small-molecule fusion inhibitor of influenza virus is orally active in mice.

Recent characterization of broadly neutralizing antibodies (bnAbs) against influenza virus identified the conserved hemagglutinin (HA) stem as a target for development of universal vaccines and therapeutics. Although several stem bnAbs are being evaluated in clinical trials, antibodies are generally unsuited for oral delivery. Guided by structural knowledge of the interactions and mechanism of anti-stem bnAb CR6261, we selected and optimized small molecules that mimic the bnAb functionality. Our lead compound neutralizes influenza A group 1 viruses by inhibiting HA-mediated fusion in vitro, protects mice against lethal and sublethal influenza challenge after oral administration, and effectively neutralizes virus infection in reconstituted three-dimensional cell culture of fully differentiated human bronchial epithelial cells. Cocrystal structures with H1 and H5 HAs reveal that the lead compound recapitulates the bnAb hotspot interactions.

Maternal antibodies protect offspring from severe influenza infection and do not lead to detectable interference with subsequent offspring immunization.

BACKGROUND: Various studies have shown that infants under the age of 6 months are especially vulnerable for complications due to influenza. Currently there are no vaccines licensed for use in this age group. Vaccination of pregnant women during the last trimester, recommended by the WHO as protective measure for this vulnerable female population, may provide protection of newborns at this early age. Although it has been observed that maternal vaccination can passively transfer protection, maternal antibodies could possibly also interfere with subsequent active vaccination of the offspring. METHODS: Using a mouse model, we evaluated in depth the ability of maternal influenza vaccination to protect offspring and the effect of maternal immunization on the subsequent influenza vaccination of the offspring. By varying the regimen of maternal immunization we explored the impact of different levels of maternal antibodies on the longevity of these antibodies in their progeny. We subsequently assessed to what extent maternal antibodies can mediate direct protection against influenza in their offspring, and whether these antibodies interfere with protection induced by active vaccination of the offspring. RESULTS: The number of immunizations of pregnant mice correlates to the level and longevity of maternal antibodies in the offspring. When these antibodies are present at time of influenza challenge they protect offspring against lethal influenza challenge, even in the absence of detectable HAI titers. Moreover, no detectable interference of passively-transferred maternal antibodies on the subsequent vaccination of the offspring was observed. CONCLUSION: In the absence of a licensed influenza vaccine for young children, vaccination of pregnant women is a promising measure to provide protection of young infants against severe influenza infection.

Evaluation of a virosomal H5N1 vaccine formulated with Matrix M™ adjuvant in a phase I clinical trial.

The avian influenza H5 virus epizootic continues to cause zoonosis with human fatalities, highlighting the continued need for pandemic preparedness against this subtype. This study evaluated the tolerability and immunogenicity of a Matrix M™ adjuvanted virosomal H5N1 vaccine in a phase I clinical trial. Sixty healthy adults were vaccinated intramuscularly with two doses of influenza H5N1 (NIBRG-14) virosomal vaccine alone (30 µg haemagglutinin (HA)) or 1.5, 7.5 or 30 µg HA formulated with 50 µg Matrix M™ adjuvant. The antibody response was analysed by haemagglutination inhibition (HI), microneutralisation (MN) and single radial haemolysis (SRH) assays. The vaccine was well tolerated in all groups but injection site pain was more frequently observed in the Matrix M™ adjuvanted groups. The vaccine elicited homologous and heterologous H5N1-specific antibody responses and the Matrix M™ adjuvanted formulations met all the EU regulatory criteria. In conclusion, Matrix M™ adjuvant was well tolerated and augmented the antibody response allowing considerable dose sparing down to 1.5 µg HA.

Safety and immunogenicity of whole-virus, alum-adjuvanted whole-virus, virosomal, and whole-virus intradermal influenza A/H9N2 vaccine formulations.

Avian influenza H9N2 viruses are considered as a pandemic threat. We assessed the safety and immunogenicity of fourteen H9N2 vaccine formulations. A randomized, phase I trial was done in 353 adults, aged 18-82 years. Subjects received two doses of A/Hong Kong/1073/99 (H9N2) whole-virus, alum-adjuvanted whole-virus, virosomal, or intradermal whole-virus vaccine at four doses (1.7, 5, 15 or 45 microg haemagglutinin). Sera were obtained before and three weeks after each vaccination (days 0, 21, and 42) for haemagglutination-inhibition (HAI) and neutralization assays. All formulations were well tolerated. Pre-vaccination sera from subjects aged below or above 40 years had baseline antibody to H9N2 in 1% and 16% of samples. Compared to intramuscular whole-virus vaccine, alum-adjuvanted vaccine was more immunogenic, intradermal vaccine was comparable, and virosomal vaccine less immunogenic. Among subjects under 40 years, two doses (45, 15, and 5 microg) of alum-adjuvanted vaccine achieved seroprotective HAI titres in 50%, 41%, and 39% respectively, and neutralization seroconversions in 83%, 82%, and 78% of recipients. Among subjects over 40 years, one dose (45, 15, and 5 microg) of alum-adjuvanted vaccine achieved seroprotective HAI titres in 50%, 25% and 0% respectively, and neutralization seroconversions in 88%, 63% and 63% of recipients. Among immunologically naive subjects under 40 years, two doses of vaccine are required and alum-adjuvanted vaccines were most immunogenic. Among immunologically primed subjects over 40 years, one dose of whole-virus or alum-adjuvanted vaccine induced immune responses; the second dose provided less additional benefit. However, no vaccine formulation satisfied all European regulatory criteria for pandemic vaccines.

Evaluation of a prime-boost vaccine schedule with distinct adenovirus vectors against malaria in rhesus monkeys.

A vaccine that elicits both specific antibodies and IFN-gamma-producing T cells is required to protect against pre-erythrocytic malaria. Among the most promising approaches to induce such complex immunity are heterologous prime-boost vaccination regimens, in particular ones containing live viral vector. We have demonstrated previously that adenovectors serotype 35 (Ads35) encoding the circumsporozoite (CS) antigen or liver-stage antigen-1 (LSA-1) are highly effective in improving the T-cell responses induced by immunizations with protein-based vaccines in a heterologous prime-boost schedule. Here we evaluated the potential of a heterologous prime-boost vaccination that combines the Ad35.CS vector with the serologically distinct adenovector Ad5.CS, in rhesus macaques, after establishing the potency in mice. We show that the heterologous Ad35.CS/Ad5.CS prime-boost regimen elicits both antibody responses and robust IFN-gamma-producing CD8(+) T-cell responses against the CS antigen. Analysis of the quality of the antibody responses in rhesus macaques, using indirect immunofluorescence assay (IFA) with Plasmodium falciparum-coated slides, demonstrated that this heterologous prime-boost regimen elicits a high titer of antibodies that are able to bind to P. falciparum sporozoites. Level of the IFA response was superior to the response measured with sera of an adult human population living in endemic malaria region. In conclusion, the combination of Ad35.CS, a vaccine based on a rare serotype adenovirus, with Ad5.CS or possibly another adenovector of a distinct serotype, induces a complex immune response that is required for protection against malaria, and is thus a highly promising approach for pediatric vaccination.

Immunogenicity and safety of low dose virosomal adjuvanted influenza vaccine administered intradermally compared to intramuscular full dose administration.

BACKGROUND: Despite the established benefit of intramuscular (i.m.) influenza vaccination, new adjuvants and delivery methods for comparable or improved immunogenicity are being explored. Intradermal (i.d.) antigen administration is hypothesized to initiate an efficient immune response at reduced antigen doses similar to that observed after i.m. full dose vaccination. METHODS: In a randomized, partially blinded phase II study 224, healthy adults aged >or=18 to

Increased immunogenicity of recombinant Ad35-based malaria vaccine through formulation with aluminium phosphate adjuvant.

Previously, we have shown the potency of recombinant Adenovirus serotype 35 viral vaccines (rAd35) to induce strong immune response against the circumsporozoite protein (CS) of the plasmodium parasite. To further optimize immunogenicity of Ad35-based malaria vaccines we formulated rAd35.CS vaccine with aluminium phosphate adjuvant (AlPO(4)). In contrast to the conventional protein based vaccines no absorption to aluminium adjuvant was observed and rAd35 viral in vitro infectivity in mammalian cells was preserved. Immunization with Ad35.CS formulated with AlPO(4) resulted in significantly higher CS specific T and B cell responses in mice upon either single or prime-boost vaccination regimens as compared to rAd35.CS alone. With these results we report for the first time the feasibility of using an AlPO(4) adjuvant to increase the potency of a live adenovirus serotype 35-based vaccine.