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Abstract Recent Ebola outbreaks underscore the importance of continuous prevention and disease control efforts. Authorized vaccines include Merck’s Ervebo (rVSV-ZEBOV) and Johnson & Johnson’s two-dose combination (Ad26.ZEBOV/MVA-BN-Filo). Here, in a five-year follow-up of the PREVAC randomized trial (NCT02876328), we report the results of the immunology ancillary study of the trial. The primary endpoint is to evaluate long-term memory T-cell responses induced by three vaccine regimens: Ad26–MVA, rVSV, and rVSV–booster. Polyfunctional EBOV-specific CD4+ T-cell responses increase after Ad26 priming and are further boosted by MVA, whereas minimal responses are observed in the rVSV groups, declining after one year. In-vitro expansion for eight days show sustained EBOV-specific T-cell responses for up to 60 months post-prime vaccination with both Ad26-MVA and rVSV, with no decline. Cytokine production analysis identify shared biomarkers between the Ad26-MVA and rVSV groups. In secondary endpoint, we observed an elevation of pro-inflammatory cytokines at Day 7 in the rVSV group. Finally, we establish a correlation between EBOV-specific T-cell responses and anti-EBOV IgG responses. Our findings can guide booster vaccination recommendations and help identify populations likely to benefit from revaccination.

Summary: Besides neutralizing antibodies, which are considered an important measure for vaccine immunogenicity, Fc-mediated antibody functions can contribute to antibody-mediated protection. They are strongly influenced by structural antibody properties such as subclass and Fc glycan composition. We here applied a systems serology approach to dissect humoral immune responses induced by MVA-MERS-S, an MVA-vectored vaccine against the Middle East respiratory syndrome coronavirus (MERS-CoV). Building on preceding studies reporting the safety and immunogenicity of MVA-MERS-S, our study highlights the potential of a late boost, administered one year after prime, to enhance both neutralizing and Fc-mediated antibody functionality compared to the primary vaccination series. Distinct characteristics were observed for antibodies specific to the MERS-CoV spike protein S1 and S2 subunits, regarding subclass and glycan compositions as well as Fc functionality. These findings highlight the benefit of a late homologous booster vaccination with MVA-MERS-S and may be of interest for the design of future coronavirus vaccines.

IntroductionMental health conditions are associated with cognition and physical function in older adults. We examined whether worry and ruminative brooding, key symptoms of certain mental health conditions, are related to subjective and/or objective measures of cognitive and physical (cardiovascular) health.MethodsWe used baseline data from 282 participants from the SCD-Well and Age-Well trials (178 female; agemean = 71.1 years). We measured worry and ruminative brooding using the Penn State Worry Questionnaire and the Ruminative Response Scale-brooding subscale. We assessed subjective physical health using the WHOQOL-Bref physical subscale, and objective physical health via blood pressure and modified versions of the Framingham Risk Score and Charlson Comorbidity Index. With subjective and objective cognition, we utilized the Cognitive Difficulties Scale and a global composite (modified Preclinical Alzheimer’s Cognitive Composite, PACC5, with the Wechsler Adult Intelligence Scale-IV, category fluency, Mattis Dementia Rating Scale-2, and either the California Verbal Learning Test or the Rey Auditory Verbal Learning Test). We conducted linear regressions, adjusted for education, age, sex and cohort.ResultsWorry and ruminative brooding were negatively associated with subjective physical health (worry: β = −0.245, 95%CI −0.357 to −0.133, p

Abstract Translational oncology research strives to explore a new aspect: identifying subgroups that exhibit treatment response even during pre-clinical phases. In this study, we focus on PDX models and their implementation in mouse clinical trials (MCT). Our primary objective was to identify subgroups with different treatment responses using Latent Class Mixed Model (LCMM).We used a public dataset and focused on one treatment, encorafenib, and two indications, melanoma and colorectal cancer, for which efficacy depends on a specific mutation BRAF V600E. One LCMM per indication was implemented to classify treatment responses at the PDX level, analyzing the growth kinetics of treated tumors and matched controls within the PDX models. A simulation study was carried out to explore the performance of LCMM in this context. For both applications, LCMM identified classes for which the higher the proportion of mutated BRAF V600E PDX models the greater the treatment effect, which is aligned with encorafenib use recommendations. The simulation study showed that LCMM could identify classes with large differences in treatment effects. LCMM is a suitable tool for MCT to explore treatment response subgroups of PDX. Once these subgroups are defined, characterization of their phenotypes/genotypes could be performed to explore treatment response predictors.

Summary: Background: Interventional trials that evaluate treatment effects using surrogate endpoints have become increasingly common. This paper describes four linked empirical studies and the development of a framework for defining, interpreting and reporting surrogate endpoints in trials. Methods: As part of developing the CONSORT (Consolidated Standards of Reporting Trials) and SPIRIT (Standard Protocol Items: Recommendations for Interventional Trials) extensions for randomised trials reporting surrogate endpoints, we undertook a scoping review, e-Delphi study, consensus meeting, and a web survey to examine current definitions and stakeholder (including clinicians, trial investigators, patients and public partners, journal editors, and health technology experts) interpretations of surrogate endpoints as primary outcome measures in trials. Findings: Current surrogate endpoint definitional frameworks are inconsistent and unclear. Surrogate endpoints are used in trials as a substitute of the treatment effects of an intervention on the target outcome(s) of ultimate interest, events measuring how patients feel, function, or survive. Traditionally the consideration of surrogate endpoints in trials has focused on biomarkers (e.g., HDL cholesterol, blood pressure, tumour response), especially in the medical product regulatory setting. Nevertheless, the concept of surrogacy in trials is potentially broader. Intermediate outcomes that include a measure of function or symptoms (e.g., angina frequency, exercise tolerance) can also be used as substitute for target outcomes (e.g., all-cause mortality)—thereby acting as surrogate endpoints. However, we found a lack of consensus among stakeholders on accepting and interpreting intermediate outcomes in trials as surrogate endpoints or target outcomes. In our assessment, patients and health technology assessment experts appeared more likely to consider intermediate outcomes to be surrogate endpoints than clinicians and regulators. Interpretation: There is an urgent need for better understanding and reporting on the use of surrogate endpoints, especially in the setting of interventional trials. We provide a framework for the definition of surrogate endpoints (biomarkers and intermediate outcomes) and target outcomes in trials to improve future reporting and aid stakeholders' interpretation and use of trial surrogate endpoint evidence. Funding: SPIRIT-SURROGATE/CONSORT-SURROGATE project is Medical Research Council Better Research Better Health (MR/V038400/1) funded.

Summary: Ebola virus disease is a severe hemorrhagic fever with a high fatality rate. We investigate transcriptome profiles at 3 h, 1 day, and 7 days after vaccination with Ad26.ZEBOV and MVA-BN-Filo. 3 h after Ad26.ZEBOV injection, we observe an increase in genes related to antigen presentation, sensing, and T and B cell receptors. The highest response occurs 1 day after Ad26.ZEBOV injection, with an increase of the gene expression of interferon-induced antiviral molecules, monocyte activation, and sensing receptors. This response is regulated by the HESX1, ATF3, ANKRD22, and ETV7 transcription factors. A plasma cell signature is observed on day 7 post-Ad26.ZEBOV vaccination, with an increase of CD138, MZB1, CD38, CD79A, and immunoglobulin genes. We have identified early expressed genes correlated with the magnitude of the antibody response 21 days after the MVA-BN-Filo and 364 days after Ad26.ZEBOV vaccinations. Our results provide early gene signatures that correlate with vaccine-induced Ebola virus glycoprotein-specific antibodies.

The crosstalk between NK cells and their surrounding environment is enabled through activating and inhibitory receptors, which tightly control NK cell activity. The co-inhibitory receptor TIGIT decreases NK cell cytotoxicity and is involved in NK cell exhaustion, but has also been associated with liver regeneration, highlighting that the contribution of human intrahepatic CD56bright NK cells in regulating tissue homeostasis remains incompletely understood. A targeted single-cell mRNA analysis revealed distinct transcriptional differences between matched human peripheral blood and intrahepatic CD56bright NK cells. Multiparameter flow cytometry identified a cluster of intrahepatic NK cells with overlapping high expression of CD56, CD69, CXCR6, TIGIT and CD96. Intrahepatic CD56bright NK cells also expressed significantly higher protein surface levels of TIGIT, and significantly lower levels of DNAM-1 compared to matched peripheral blood CD56bright NK cells. TIGIT+ CD56bright NK cells showed diminished degranulation and TNF-α production following stimulation. Co-incubation of peripheral blood CD56bright NK cells with human hepatoma cells or primary human hepatocyte organoids resulted in migration of NK cells into hepatocyte organoids and upregulation of TIGIT and downregulation of DNAM-1 expression, in line with the phenotype of intrahepatic CD56bright NK cells. Intrahepatic CD56bright NK cells represent a transcriptionally, phenotypically, and functionally distinct population of NK cells that expresses higher levels of TIGIT and lower levels of DNAM-1 than matched peripheral blood CD56bright NK cells. Increased expression of inhibitory receptors by NK cells within the liver environment can contribute to tissue homeostasis and reduction of liver inflammation.

NK cells play a pivotal role in viral immunity, utilizing a large array of activating and inhibitory receptors to identify and eliminate virus-infected cells. Killer-cell immunoglobulin-like receptors (KIRs) represent a highly polymorphic receptor family, regulating NK cell activity and determining the ability to recognize target cells. Human leukocyte antigen (HLA) class I molecules serve as the primary ligand for KIRs. Herein, HLA-C stands out as being the dominant ligand for the majority of KIRs. Accumulating evidence indicated that interactions between HLA-C and its inhibitory KIR2DL receptors (KIR2DL1/L2/L3) can drive HIV-1-mediated immune evasion and thus may contribute to the intrinsic control of HIV-1 infection. Of particular interest in this context is the recent observation that HIV-1 is able to adapt to host HLA-C genotypes through Vpu-mediated downmodulation of HLA-C. However, our understanding of the complex interplay between KIR/HLA immunogenetics, NK cell-mediated immune pressure and HIV-1 immune escape is still limited. Therefore, we investigated the impact of specific KIR/HLA-C combinations on the NK cell receptor repertoire and HIV-1 Vpu protein sequence variations of 122 viremic, untreated HIV-1+ individuals. Compared to 60 HIV-1- controls, HIV-1 infection was associated with significant changes within the NK cell receptor repertoire, including reduced percentages of NK cells expressing NKG2A, CD8, and KIR2DS4. In contrast, the NKG2C+ and KIR3DL2+ NK cell sub-populations from HIV-1+ individuals was enlarged compared to HIV-1- controls. Stratification along KIR/HLA-C genotypes revealed a genotype-dependent expansion of KIR2DL1+ NK cells that was ultimately associated with increased binding affinities between KIR2DL1 and HLA-C allotypes. Lastly, our data hinted to a preferential selection of Vpu sequence variants that were associated with HLA-C downmodulation in individuals with high KIR2DL/HLA-C binding affinities. Altogether, our study provides evidence that HIV-1-associated changes in the KIR repertoire of NK cells are to some extent predetermined by host KIR2DL/HLA-C genotypes. Furthermore, analysis of Vpu sequence polymorphisms indicates that differential KIR2DL/HLA-C binding affinities may serve as an additional mechanism how host genetics impact immune evasion by HIV-1.

Abstract Introduction The Ebola virus disease (EVD) outbreak in 2014–2016 in West Africa was the largest on record and provided an opportunity for large clinical trials and accelerated efforts to develop an effective and safe preventative vaccine. Multiple questions regarding the safety, immunogenicity, and efficacy of EVD vaccines remain unanswered. To address these gaps in the evidence base, the Partnership for Research on Ebola Vaccines (PREVAC) trial was designed. This paper describes the design, methods, and baseline results of the PREVAC trial and discusses challenges that led to different protocol amendments. Methods This is a randomized, double-blind, placebo-controlled phase 2 clinical trial of three vaccine strategies against the Ebola virus in healthy volunteers 1 year of age and above. The three vaccine strategies being studied are the rVSVΔG-ZEBOV-GP vaccine, with and without a booster dose at 56 days, and the Ad26.ZEBOV,MVA-FN-Filo vaccine regimen with Ad26.ZEBOV given as the first dose and the MVA-FN-Filo vaccination given 56 days later. There have been 4 versions of the protocol with those enrolled in Version 4.0 comprising the primary analysis cohort. The primary endpoint is based on the antibody titer against the Ebola virus surface glycoprotein measured 12 months following the final injection. Results From April 2017 to December 2018, a total of 5002 volunteers were screened and 4789 enrolled. Participants were enrolled at 6 sites in four countries (Guinea, Liberia, Sierra Leone, and Mali). Of the 4789 participants, 2560 (53%) were adults and 2229 (47%) were children. Those

Abstract Natural killer (NK) cells are implicated among immune effectors after vaccination against viral pathogens, including Ebola virus. The two-dose heterologous Ebola virus vaccine regimen, adenovirus type 26.ZEBOV followed by modified vaccinia Ankara-BN-Filo (EBOVAC2 consortium, EU Innovative Medicines Initiative), induces NK cell activation and anti-Ebola glycoprotein (GP) antibody-dependent NK cell activation post-dose 1, which is further elevated post-dose 2. Here, in a multicentre, phase 2 clinical trial (EBL2001), we demonstrate durable ex vivo NK cell activation 180 days after dose 2, with responses enriched in CD56bright NK cells. In vitro antibody-dependent responses to immobilised Ebola GP increased after dose 1, and remained elevated compared to pre-vaccination levels in serum collected 180 days later. Peak NK cell responses were observed post-dose 2 and NK cell IFN-γ responses remained significantly elevated at 180 days post-dose 2. Individual variation in NK cell responses were influenced by both anti-Ebola GP antibody concentrations and intrinsic interindividual differences in NK cell functional capacity. In summary, this study demonstrates durable NK cell responses after Ad26.ZEBOV, MVA-BN-Filo Ebola virus vaccination and could inform the immunological evaluation of future iterations of the vaccine regimen and vaccination schedules.

The transcription factor promyelocytic leukemia zinc finger protein (PLZF) is involved in the development of natural killer (NK) cells and innate lymphoid cells, including liver‐resident NK cells in mice. In human NK cells, the role of PLZF in liver residency is still unknown. Expression of PLZF in matched human peripheral blood‐ and liver‐derived NK cells and the association of PLZF expression with surface molecules and transcription factors relevant for tissue residency were investigated using multiparameter flow cytometry and assessing single‐cell messenger RNA (mRNA) levels. Intrahepatic cluster of differentiation (CD)56bright NK cells expressed significantly higher levels of PLZF than peripheral blood CD56bright NK cells, which were predominantly PLZFlo. Expression of PLZF was highest within C‐X‐C motif chemokine receptor 6 (CXCR6)+CD69+ liver‐resident NK cells among intrahepatic CD56bright NK cell populations. Association of PLZF with liver‐residency markers was also reflected at mRNA levels. A small PLZFhiCD56bright NK cell population was identified in peripheral blood that also expressed the liver‐residency markers CXCR6 and CD69 and shared functional characteristics with liver‐resident NK cells. Conclusion: PLZF is implicated as part of a transcriptional network that promotes liver residency of human NK cells. Expression of liver‐homing markers on peripheral blood PLZFhiCD56bright NK cells identifies an intermediate population potentially contributing to the maintenance of liver‐resident NK cells.

Background Reoccurring Ebola outbreaks in West and Central Africa have led to serious illness and death in thousands of adults and children. The objective of this study was to assess safety, tolerability, and immunogenicity of the heterologous 2-dose Ad26.ZEBOV, MVA-BN-Filo vaccination regimen in adolescents and children in Africa. Methods and findings In this multicentre, randomised, observer-blind, placebo-controlled Phase II study, 131 adolescents (12 to 17 years old) and 132 children (4 to 11 years old) were enrolled from Eastern and Western Africa and randomised 5:1 to receive study vaccines or placebo. Vaccine groups received intramuscular injections of Ad26.ZEBOV (5 × 1010 viral particles) and MVA-BN-Filo (1 × 108 infectious units) 28 or 56 days apart; placebo recipients received saline. Primary outcomes were safety and tolerability. Solicited adverse events (AEs) were recorded until 7 days after each vaccination and serious AEs (SAEs) throughout the study. Secondary and exploratory outcomes were humoral immune responses (binding and neutralising Ebola virus [EBOV] glycoprotein [GP]-specific antibodies), up to 1 year after the first dose. Enrolment began on February 26, 2016, and the date of last participant last visit was November 28, 2018. Of the 263 participants enrolled, 217 (109 adolescents, 108 children) received the 2-dose regimen, and 43 (20 adolescents, 23 children) received 2 placebo doses. Median age was 14.0 (range 11 to 17) and 7.0 (range 4 to 11) years for adolescents and children, respectively. Fifty-four percent of the adolescents and 51% of the children were male. All participants were Africans, and, although there was a slight male preponderance overall, the groups were well balanced. No vaccine-related SAEs were reported; solicited AEs were mostly mild/moderate. Twenty-one days post-MVA-BN-Filo vaccination, binding antibody responses against EBOV GP were observed in 100% of vaccinees (106 adolescents, 104 children). Geometric mean concentrations tended to be higher after the 56-day interval (adolescents 13,532 ELISA units [EU]/mL, children 17,388 EU/mL) than the 28-day interval (adolescents 6,993 EU/mL, children 8,007 EU/mL). Humoral responses persisted at least up to Day 365. A limitation of the study is that the follow-up period was limited to 365 days for the majority of the participants, and so it was not possible to determine whether immune responses persisted beyond this time period. Additionally, formal statistical comparisons were not preplanned but were only performed post hoc. Conclusions The heterologous 2-dose vaccination was well tolerated in African adolescents and children with no vaccine-related SAEs. All vaccinees displayed anti-EBOV GP antibodies after the 2-dose regimen, with higher responses in the 56-day interval groups. The frequency of pyrexia after vaccine or placebo was higher in children than in adolescents. These data supported the prophylactic indication against EBOV disease in a paediatric population, as licenced in the EU. Trial registration ClinicalTrials.govNCT02564523. Zacchaeus Anywaine and co-workers study safety and immunogenicity of an Ebola vaccine among children and adolescents across four African countries. Author summary Why was the study done? There have been larger and more extensive Ebola virus disease (EVD) outbreaks in Africa in the past decade with no licenced treatments available. As such, there is an unmet medical need for prophylactic Ebola vaccines. This study was performed to evaluate whether a 2-dose heterologous Ad26.ZEBOV, MVA-BN-Filo Ebola vaccination was safe and immunogenic in healthy African children. What did the researchers do and find? In this randomised, placebo-controlled, Phase II clinical trial, the Ad26.ZEBOV, MVA-BN-Filo Ebola vaccination regimen was administered to African participants in 2 age cohorts (12 to 17 and 4 to 11 years). No vaccine-related serious adverse events were reported, and robust immune responses were induced in both adolescents and children after receiving the active 2-dose regimen. What do these findings mean? These data support the use of the Ad26.ZEBOV, MVA-BN-Filo vaccination regimen in African adolescents and children at risk of Ebola infection. Although vaccination according to a 28-day regimen may lead to protection against EVD in a shorter time frame, vaccination according to a 56-day regimen results in higher EBOV GP binding and neutralising antibody responses. The observation that Ad26 preexisting immunity in the majority of participants does not affect the EBOV GP-specific antibody responses postvaccination augurs well for the use of this vaccine regimen even in regions with a high prevalence of preexisting Ad26 seropositivity.

The mechanisms underlying rapid elimination of herpes simplex virus-2 (HSV-2) in the human genital tract despite low [CD8.sup.+] and [CD4.sup.+] tissue-resident T cell (Trm cell) density are unknown. We analyzed shedding episodes during chronic HSV-2 infection; viral clearance always predominated within 24 hours of detection even when viral load exceeded 1 x 107 HSV DNA copies, and surges in granzyme B and IFN-[gamma] occurred within the early hours after reactivation and correlated with local viral load. We next developed an agent-based mathematical model of an HSV-2 genital ulcer to integrate mechanistic observations of Trm cells in in situ proliferation, trafficking, cytolytic effects, and cytokine alarm signaling from murine studies with viral kinetics, histopathology, and lesion size data from humans. A sufficiently high density of HSV-2-specific Trm cells predicted rapid elimination of infected cells, but our data suggest that such Trm cell densities are relatively uncommon in infected tissues. At lower, more commonly observed Trm cell densities, Trm cells must initiate a rapidly diffusing, polyfunctional cytokine response with activation of bystander T cells in order to eliminate a majority of infected cells and eradicate briskly spreading HSV-2 infection., Introduction [CD8.sup.+] and [CD4.sup.+] tissue-resident T cells (Trm cells) rapidly identify and control recurrent human viral infections within peripheral sites (1-11). Trm cells lodge preferentially at prior sites of pathogen [...]

Background We investigated safety, tolerability, and immunogenicity of the heterologous 2-dose Ebola vaccination regimen in healthy and HIV-infected adults with different intervals between Ebola vaccinations. Methods and findings In this randomised, observer-blind, placebo-controlled Phase II trial, 668 healthy 18- to 70-year-olds and 142 HIV-infected 18- to 50-year-olds were enrolled from 1 site in Kenya and 2 sites each in Burkina Faso, Cote d’Ivoire, and Uganda. Participants received intramuscular Ad26.ZEBOV followed by MVA-BN-Filo at 28-, 56-, or 84-day intervals, or saline. Females represented 31.4% of the healthy adult cohort in contrast to 69.7% of the HIV-infected cohort. A subset of healthy adults received booster vaccination with Ad26.ZEBOV or saline at Day 365. Following vaccinations, adverse events (AEs) were collected until 42 days post last vaccination and serious AEs (SAEs) were recorded from signing of the ICF until the end of the study. The primary endpoint was safety, and the secondary endpoint was immunogenicity. Anti-Ebola virus glycoprotein (EBOV GP) binding and neutralising antibodies were measured at baseline and at predefined time points throughout the study. The first participant was enrolled on 9 November 2015, and the date of last participant’s last visit was 12 February 2019. No vaccine-related SAEs and mainly mild-to-moderate AEs were observed among the participants. The most frequent solicited AEs were injection-site pain (local), and fatigue, headache, and myalgia (systemic), respectively. Twenty-one days post-MVA-BN-Filo vaccination, geometric mean concentrations (GMCs) with 95% confidence intervals (CIs) of EBOV GP binding antibodies in healthy adults in 28-, 56-, and 84-day interval groups were 3,085 EU/mL (2,648 to 3,594), 7,518 EU/mL (6,468 to 8,740), and 7,300 EU/mL (5,116 to 10,417), respectively. In HIV-infected adults in 28- and 56-day interval groups, GMCs were 4,207 EU/mL (3,233 to 5,474) and 5,283 EU/mL (4,094 to 6,817), respectively. Antibody responses were observed until Day 365. Ad26.ZEBOV booster vaccination after 1 year induced an anamnestic response. Study limitations include that some healthy adult participants either did not receive dose 2 or received dose 2 outside of their protocol-defined interval and that the follow-up period was limited to 365 days for most participants. Conclusions Ad26.ZEBOV, MVA-BN-Filo vaccination was well tolerated and immunogenic in healthy and HIV-infected African adults. Increasing the interval between vaccinations from 28 to 56 days improved the magnitude of humoral immune responses. Antibody levels persisted to at least 1 year, and Ad26.ZEBOV booster vaccination demonstrated the presence of vaccination-induced immune memory. These data supported the approval by the European Union for prophylaxis against EBOV disease in adults and children ≥1 year of age. Trial registration ClinicalTrials.govNCT02564523 Houreratou Barry and co-workers report on safety and immunogenicity of an Ebola vaccine in adults across four African countries. Author summary Why was this study done? With Ebola outbreaks increasing, there is an unmet medical need for a prophylactic vaccine to prevent and mitigate Ebola outbreaks. To address the urgent medical need during the 2014 to 2016 outbreak, the clinical development of the 2-dose vaccine regimen comprising of Ad26.ZEBOV and MVA-BN-Filo was accelerated. This Phase II study was part of this accelerated program, evaluating the safety and immunogenicity of the 2-dose vaccine regimen in healthy and HIV-infected African adults, with 28-, 56-, and 84-day intervals between doses. The study was amended to evaluate safety and immunogenicity of a booster vaccination with Ad26.ZEBOV, administered approximately 1 year after the first vaccination, in healthy adults. What did the researchers do and find? We conducted a randomised trial to assess the safety and the immunogenicity of the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen in 3 different vaccination intervals in healthy and HIV-infected adults. The vaccine regimen was well tolerated and induced marked immune responses; the highest humoral responses were observed after vaccination with 56-day and 84-day intervals. Anamnestic responses were observed in all healthy participants receiving Ad26.ZEBOV as booster at 1 year after the first dose. What do these findings mean? Our results demonstrate that the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen is safe and immunogenic in healthy and HIV-infected adults and induces immune memory that can rapidly be reactivated. Our findings support the prophylactic use of the 2-dose Ad26.ZEBOV, MVA-BN-Filo vaccine regimen against Ebola infection in African adult populations. The 2-dose vaccine regimen comprising of Ad26.ZEBOV and MVA-BN-Filo has received marketing authorisation under exceptional circumstances for prophylactic use against EVD in adults and children ≥1 year old within the European Union.

Abstract Objectives To assess the efficacy of several repurposed drugs to prevent hospitalisation or death in patients aged 65 or more with recent symptomatic SARS-CoV-2 infection (COVID-19) and no criteria for hospitalisation. Trial design Phase III, multi-arm (5) and multi-stage (MAMS), randomized, open-label controlled superiority trial. Participants will be randomly allocated 1:1:1:1:1 to the following strategies: Arm 1: Control arm Arms 2 to 5: Experimental treatment arms Planned interim analyses will be conducted at regular intervals. Their results will be reviewed by an Independent Data and Safety Monitoring Board. Experimental arms may be terminated for futility, efficacy or toxicity before the end of the trial. New experimental arms may be added if new evidence suggests that other treatments should be tested. A feasibility and acceptability substudy as well as an immunological substudy will be conducted alongside the trial. Participants Inclusion criteria are: 65-year-old or more; Positive test for SARS-CoV-2 on a nasopharyngeal swab; Symptoms onset within 3 days before diagnosis; No hospitalisation criteria; Signed informed consent; Health insurance. Exclusion criteria are: Inability to make an informed decision to participate (e.g.: dementia, guardianship); Rockwood Clinical Frailty Scale ≥7; Long QT syndrome; QTc interval > 500 ms; Heart rate 5.5 mmol/L or

Abstract Background Enrolling participants in clinical trials can be challenging, especially with respect to prophylactic vaccine trials. The vaccination of study personnel in Ebola vaccine trials during the 2014–2016 epidemic played a crucial role in inspiring trust and facilitating volunteer enrollment. We evaluated the ethical and methodological considerations as they applied to an ongoing phase 2 randomized prophylactic Ebola vaccine trial that enrolled healthy volunteers in Guinea, Liberia, Sierra Leone, and Mali in a non-epidemic context. Methods On the assumption that the personnel on site involved in executing the protocol, as well as community mobilizers (not involved in the on-site procedures), might also volunteer to enter the trial, we considered both ethical and methodological considerations to set clear rules that can be shared a priori with these persons. We reviewed the scientific and gray literature to identify relevant references and then conducted an analysis of the ethical and methodological considerations. Results There are currently no regulations preventing a clinical investigator or site staff from participating in a trial. However, the enrollment of personnel raises the risk of undue influence and challenges the basic ethical principle of voluntary participation. The confidentiality of personal medical information, such as HIV test results, may also be difficult to ensure among personnel. There is a risk of disruption of trial operations due to the potential absence of the personnel for their commitment as trial participants, and there is also a potential for introducing differential behavior of on-site staff as they obtain access to accumulating information during the trial (e.g., the incidence of adverse events). Blinding could be jeopardized, given knowledge of product-specific adverse event profiles and the proximity to unblinded site staff. These aspects were considered more relevant for on-site staff than for community mobilizers, who have limited contact with site staff. Conclusion In a non-epidemic context, ethical and methodological considerations limit the collective benefit of enrolling site staff in a vaccine trial. These considerations do not apply to community mobilizers, whose potential enrollment should be considered as long as they meet the inclusion criteria and they are not exposed to any form of coercion.

NK cells are phenotypically and functionally diverse lymphocytes due to variegated expression of a large array of receptors. NK-cell activity is tightly regulated through integration of receptor-derived inhibitory and activating signals. Thus, the receptor profile of each NK cell ultimately determines its ability to sense aberrant cells and subsequently mediate anti-viral or anti-tumor responses. However, an in-depth understanding of how different receptor repertoires enable distinct immune functions of NK cells is lacking. Therefore, we investigated the phenotypic diversity of primary human NK cells by performing extensive phenotypic characterization of 338 surface molecules using flow cytometry (n = 18). Our results showed that NK cells express at least 146 receptors on their surface. Of those, 136 (>90%) exhibited considerable inter-donor variability. Moreover, comparative analysis of CD56bright and CD56dim NK cells identified 70 molecules with differential expression between the two major NK-cell subsets and allowed discrimination of these subsets via unsupervised hierarchical clustering. These receptors were associated with a broad range of NK-cell functions and multiple molecules were not previously associated with predominant expression on either subset (e.g. CD82 and CD147). Altogether, our study contributes to an improved understanding of the phenotypic diversity of NK cells and its potential functional implications on a cellular and population level. While the identified distinct signatures in the receptor repertoires provide a molecular basis for the differential immune functions exerted by CD56bright and CD56dim NK cells, the observed inter-individual differences in the receptor repertoire of NK cells may contribute to a diverging ability to control certain diseases.

Natural killer cells play an important role in the control of viral infections both by regulating acquired immune responses and as potent innate or antibody-mediated cytotoxic effector cells. NK cells have been implicated in control of Ebola virus infections and our previous studies in European trial participants have demonstrated durable activation, proliferation and antibody-dependent NK cell activation after heterologous two-dose Ebola vaccination with adenovirus type 26.ZEBOV followed by modified vaccinia Ankara-BN-Filo. Regional variation in immunity and environmental exposure to pathogens, in particular human cytomegalovirus, have profound impacts on NK cell functional capacity. We therefore assessed the NK cell phenotype and function in African trial participants with universal exposure to HCMV. We demonstrate a significant redistribution of NK cell subsets after vaccine dose two, involving the enrichment of less differentiated CD56dimCD57− and CD56dimFcεR1γ+ (canonical) cells and the increased proliferation of these subsets. Sera taken after vaccine dose two support robust antibody-dependent NK cell activation in a standard NK cell readout; these responses correlate strongly with the concentration of anti-Ebola glycoprotein specific antibodies. These sera also promote comparable IFN-γ production in autologous NK cells taken at baseline and post-vaccine dose two. However, degranulation responses of post-vaccination NK cells were reduced compared to baseline NK cells and these effects could not be directly attributed to alterations in NK cell phenotype after vaccination. These studies demonstrate consistent changes in NK cell phenotypic composition and robust antibody-dependent NK cell function and reveal novel characteristics of these responses after heterologous two dose Ebola vaccination in African individuals.

Objectives: For Ebola vaccine development, antibody response is a major endpoint although its determinants are not well known. We aimed to review Ebola vaccine studies and to assess factors associated with antibody response variability in humans. Methods: We searched PubMed and Scopus for preventive Ebola vaccine studies in humans or non-human primates (NHP), published up to February 2018. For each vaccination group with Ebola Zaire antibody titre measurements after vaccination, data about antibody response and its potential determinants were extracted. A random-effects meta-regression was conducted including human groups with at least 8 individuals. Results: We reviewed 49 studies (202 vaccination groups including 74 human groups) with various vaccine platforms and antigen inserts. Mean antibody titre was slightly higher in NHP (3.10, 95% confidence interval [293; 327]) than in humans (2.75 [257; 293]). Vaccine platform (p

Predicting vaccine efficacy remains a challenge. We used a systems vaccinology approach to identify early innate immune correlates of antibody induction in humans receiving the Ebola vaccine rVSV-ZEBOV. Blood samples from days 0, 1, 3, 7, and 14 were analyzed for changes in cytokine levels, innate immune cell subsets, and gene expression. Integrative statistical analyses with cross-validation identified a signature of 5 early innate markers correlating with antibody titers on day 28 and beyond. Among those, IP-10 on day 3 and MFI of CXCR6 on NK cells on day 1 were independent correlates. Consistently, we found an early gene expression signature linked to IP-10. This comprehensive characterization of early innate immune responses to the rVSV-ZEBOV vaccine in humans revealed immune signatures linked to IP-10. These results suggest correlates of vaccine-induced antibody induction and provide a rationale to explore strategies for augmenting the effectiveness of vaccines through manipulation of IP-10.

The goal of HIV therapeutic vaccination is to induce HIV-specific immune response able to control HIV replication. We previously reported that vaccination with ex vivo generated Dendritic Cells (DC) loaded with HIV-lipopeptides in HIV-infected patients (n = 19) on antiretroviral therapy (ART) was well-tolerated and immunogenic. Vaccine-elicited HIV-specific T cell responses were associated with improved control of viral replication following antiretroviral interruption (ATI from w24 to w48). We show an inverse relationship between HIV-specific responses (production of IL-2, IL-13, IL-21, IFN-g, CD4 polyfunctionality, i.e., production of at least two cytokines) and the peak of viral load during ATI. Here we have performed an integrative systems vaccinology analysis including: (i) post vaccination (w16) immune responses assessed by cytometry, cytokine secretion, and Interferon-γ ELISPOT assays; (ii) whole blood and cellular gene expression measured during vaccination; and (iii) viral parameters following ATI, with the objective to disentangle the relationships between these markers and to identify vaccine signatures. During vaccination, 69 gene expression modules out of 260 varied significantly including (by order of significance) modules related to inflammation (Chaussabel Modules M3.2, M4.13, M4.6, M5.7, M7.1, M4.2), plasma cells (M4.11) and T cells (M4.1, 4.15). Cellular immune responses were positively correlated to genes belonging to T cell functional modules (M4.1, M4.15) at w16 and negatively correlated to genes belonging to inflammation modules (M7.1, M5.7, M3.2, M4.13, M4.2). More specifically, we show that prolonged increased abundance of inflammatory gene pathways related to toll-like receptor signaling (especially TLR4) are associated with both lower vaccine immune responses and control of viral replication post ATI. Further comparison of DC vaccine gene signatures with previously reported non-HIV vaccine signatures, such as flu and pneumococcal vaccines, revealed common pathways across vaccines. Overall, these results show that too long duration and too high intensity of vaccine inflammatory responses hamper the magnitude of effector responses.

BackgroundComparison of changes in body composition, adipokines and inflammatory markers after initial therapy with a nucleos(t)ide reverse transcriptase inhibitor (N(t)RTI)- sparing or containing regimen are scarce.DesignRandomised Clinical Trial.MethodsThis is the body composition substudy of NEAT 001/ANRS 143, a randomised trial comparing darunavir/ritonavir (DRV/r) plus either raltegravir (RAL) or tenofovir disoproxil fumarate/emtricitabine (TDF/FTC) in 805 ART naïve HIV-infected adults. The primary endpoint was percentage change in limb fat at week 96. Secondary endpoints were associations among these changes and metabolic markers (IL-6, insulin, leptin, adiponectin, FGF-23).Results126 subjects (61 DRV/r + RAL and 65 DRV/r + TDF/FTC) were included. The rate of change in BMI between groups for RAL versus TDF/FTC at week 96 was 1.5% per 48-week period (p = 0.015). The rate of change in limb fat mass, trunk fat mass, total body fat and total lean mass was for RAL versus TDF/FTC at week 96 was 2.5% (p = 0.38), 7.3% ((p = 0.021), 4.9% (p = 0.061) and 1.3% (p = 0.12) respectively. Baseline insulin and leptin levels were correlated with baseline limb fat and trunk fat mass [r = 0.31 (p = 0.0043)/r = 0.28 (p = 0.0011) for limb fat, and r = 0.63 (pConclusionsAfter week 96 a N(t)RTI sparing regimen of DRV/r + RAL produced a numerically greater percentage increase in body composition variables with only change in trunk fat mass and BMI being significant.

HIV-1 infection can be controlled by anti-retroviral drug therapy, but this is a lifetime treatment and the virus remains latent and rapidly rebounds if therapy is stopped. HIV-1-infected individuals under this drug regimen have increased rates of cancers, cardiovascular diseases, and autoimmunity due to compromised immunity. A therapeutic vaccine boosting cellular immunity against HIV-1 is therefore desirable and, possibly combined with other immune modulating agents, could obviate the need for long-term drug therapies. An approach to elicit strong T cell-based immunity is to direct virus protein antigens specifically to dendritic cells (DCs), which are the key cell type for controlling immune responses. For eliciting therapeutic cellular immunity in HIV-1-infected individuals, we developed vaccines comprised of five T cell epitope-rich regions of HIV-1 Gag, Nef, and Pol (HIV5pep) fused to monoclonal antibodies that bind either, the antigen presenting cell activating receptor CD40, or the endocytic dendritic cell immunoreceptor DCIR. The study aimed to demonstrate vaccine safety, establish efficacy for broad T cell responses in both primed and naïve settings, and identify one candidate vaccine for human therapeutic development. The vaccines were administered to Rhesus macaques by intradermal injection with poly-ICLC adjuvant. The animals were either i) naïve or, ii) previously primed with modified vaccinia Ankara vector (MVA) encoding HIV-1 Gag, Pol, and Nef (MVA GagPolNef). In the MVA-primed groups, both DC-targeting vaccinations boosted HIV5pep-specific blood CD4+ T cells producing multiple cytokines, but did not affect the MVA-elicited CD8+ T cell responses. In the naive groups, both DC-targeting vaccines elicited antigen-specific polyfunctional CD4+ and CD8+ T cell responses to multiple epitopes and these responses were unchanged by a subsequent MVA GagPolNef boost. In both settings, the T cell responses elicited via the CD40-targeting vaccine were more robust and were detectable in all the animals, favoring further development of the CD40-targeting vaccine for therapeutic vaccination of HIV-1-infected individuals.

Metabolism is a critical basis for immune cell functionality. It was recently shown that NK cell subsets from peripheral blood modulate their expression of nutrient receptors following cytokine stimulation, demonstrating that NK cells can adjust to changes in metabolic requirements. As nutrient availability in blood and tissues can significantly differ, we examined NK cells isolated from paired blood-liver and blood-spleen samples and compared expression of the nutrient transporters Glut1, CD98 and CD71. CD56bright tissue-resident (CXCR6+) NK cells derived from livers and spleens expressed lower levels of Glut1 but higher levels of the amino acid transporter CD98 following stimulation than CD56bright NK cells from peripheral blood. In line with that, CD56dim NK cells, which constitute the main NK cell population in the peripheral blood, expressed higher levels of Glut1 and lower levels of CD98 and CD71 compared to liver CD56bright NK cells. Our results show that NK cells from peripheral blood differ from liver- and spleen-resident NK cells in the expression profile of nutrient transporters, consistent with a cell-adaptation to the different nutritional environment in these compartments.

The recruitment and retention of Natural Killer (NK) cells in the liver are thought to play an important role during hepatotropic infections and liver cirrhosis. The aims of this study were to determine differences between liver-derived and peripheral blood-derived NK cells in the context of liver inflammation and cirrhosis. We conducted a prospective dual-center cross-sectional study in patients undergoing liver transplantation or tumor-free liver resections, in which both liver tissue and peripheral blood samples were obtained from each consenting study participants. Intrahepatic lymphocytes and PBMCs were stained, fixed and analyzed by flow cytometry. Our results showed that, within cirrhotic liver samples, intrahepatic NK cells were particularly enriched for CD49a+ NK cells when compared to tumor-free liver resection samples. CD49a+ liver-derived NK cells included populations of cells expressing CD25, CD34 and CXCR3. Moreover, CD49a+CD25+ liver-derived NK cells exhibited high proliferative capacity in vitro in response to low doses of IL-2. Our study identified a specific subset of CD49a+CD25+ NK cells in cirrhotic livers bearing functional features of proliferation.

Initial evaluation of a candidate vaccine against HIV includes an assessment of the vaccine's ability to generate immune responses. However, the dynamics of vaccine-induced immune responses are unclear. We hypothesized that the IFN-γ producing cytotoxic CD8+ (CD8+ IFN-γ+) T cell responses could be predicted by early IL-2 producing CD4+ (CD4+ IL-2+) helper T cell responses, and we evaluated this hypothesis using data from a phase I/II prophylactic HIV vaccine trial. The objective was to assess the dynamics and correlations between CD4+ IL-2+ T cell and CD8+ IFN-γ+ T cell responses after vaccination with a recombinant adenoviral serotype 5 (rAd5) HIV vaccine.We analyzed data from the HVTN 068 HIV vaccine trial, which evaluated the immunogenicity of two different strategies for prime and boost vaccination (rAd5-rAd5 vaccine versus DNA-rAd5) in 66 healthy volunteers. Spearman correlations between immunogenicity markers across time-points were calculated. CD8+ IFN-γ+ T cell response in the rAd5-rAd5 arm was modeled as a function of CD4+ IL-2+ T cell response and time using mixed effects regression models.Moderate to high correlations (r = 0.48-0.76) were observed in the rAd5-rAd5 arm between the CD4+ IL-2+ T cell response at week 2 and later CD8+ IFN-γ+ T cell responses (weeks 2-52). Regression models confirmed this relationship with a significant association between the two markers: for a 1.0% increase in CD4+ IL-2+ T cells at week 2 post-prime, a 0.3% increase in CD8+ IFN-γ+ T cell responses across subsequent time points, including post-boost time points, was observed (p

To assess whether higher 25-hydroxyvitamin D (25OHD) levels are associated with subsequent better immune responses to hepatitis B and Streptococcus pneumoniae vaccination in HIV-infected patients.25OHD was measured on stored baseline plasma samples from two randomized vaccine trials in HIV-infected adults: the ANRS HB03 VIHVAC B trial and an immunological sub-study of the ANRS 114-PNEUMOVAC trial. In ANRS HB03 VIHVAC B, participants received three or four doses of recombinant HBV vaccine strategies. Anti-HBs IgG titers were measured four weeks after the last injection. Associations between baseline 25OHD levels and ordered IgG response categories were analyzed in multivariable proportional odds models. In the ANRS 114-PNEUMOVAC sub-study, two strategies of pneumococcal vaccination were tested, cellular immune responses were measured at repeated time points, and IgG responses four weeks after the last vaccine injection. Exploratory statistical analyses were performed on this sub-study data set.Three hundred and thirty-nine ANRS HB03 VIHVAC B and 25 ANRS 114-PNEUMOVAC sub-study participants were included in the analyses. Median age in each of the two studies was 43 years, 68% were male, and 77-92% on antiretroviral treatment. Median 25OHD level was 18 ng/mL (IQR: 12-25) and 24 ng/mL (IQR: 13-32) in the two trial populations, respectively. In the multivariable model, there was no significant association between baseline 25OHD level and vaccine responses in ANRS HB03 VIHVAC B (proportional odds ratio 0.83 per 10 ng/mL 25OHD increase; 95% confidence interval 0.65-1.07, p = 0.14). Exploratory analyses of ANRS 114-PNEUMOVAC showed consistent results.This study does not support a positive association between 25OHD and immune responses to hepatitis B or pneumococcal vaccination in HIV-infected patients.

Improved antigenicity against HIV-1 envelope (Env) protein is needed to elicit vaccine-induced protective immunity in humans. Here we describe the first tests in non-human primates (NHPs) of Env gp140 protein fused to a humanized anti-LOX-1 recombinant antibody for delivering Env directly to LOX-1-bearing antigen presenting cells, especially dendritic cells (DC). LOX-1, or 1ectin-like oxidized low-density lipoprotein (LDL) receptor-1, is expressed on various antigen presenting cells and endothelial cells, and is involved in promoting humoral immune responses. The anti-LOX-1 Env gp140 fusion protein was tested for priming immune responses and boosting responses in animals primed with replication competent NYVAC-KC Env gp140 vaccinia virus. Anti-LOX-1 Env gp140 vaccination elicited robust cellular and humoral responses when used for either priming or boosting immunity. Co-administration with Poly ICLC, a TLR3 agonist, was superior to GLA, a TLR4 agonist. Both CD4+ and CD8+ Env-specific T cell responses were elicited by anti-LOX-1 Env gp140, but in particular the CD4+ T cells were multifunctional and directed to multiple epitopes. Serum IgG and IgA antibody responses induced by anti-LOX-1 Env gp140 against various gp140 domains were cross-reactive across HIV-1 clades; however, the sera neutralized only HIV-1 bearing sequences most similar to the clade C 96ZM651 Env gp140 carried by the anti-LOX-1 vehicle. These data, as well as the safety of this protein vaccine, justify further exploration of this DC-targeting vaccine approach for protective immunity against HIV-1.

The role of regulatory T cells (Tregs) in vaccination has been poorly investigated. We have reported that vaccination with ex vivo-generated dendritic-cells (DC) loaded with HIV-lipopeptides (LIPO-5-DC vaccine) in HIV-infected patients was well tolerated and highly immunogenic. These responses and their relation to viral replication following analytical treatment interruption (ATI) were variable. Here, we investigated whether the presence of HIV-specific Tregs might explain these differences. Co-expression of CD25, CD134, CD39 and FoxP3 was used to delineate both antigen-specific Tregs and effectors T cells (Teffs). Median LIPO-5 specific-CD25+CD134+ polyfunctional T cells increased from 0.1% (IQR 0-0.3) before vaccination (week -4) to 2.1% (IQR 1.1-3.9) at week 16 following 4 immunizations (p=0.001) and were inversely correlated with maximum viral load following ATI (r=-0.77, p=0.001). Vaccinees who displayed lower levels of HIV-specific CD4+CD134+CD25+CD39+FoxP3+ Tregs responded better to the LIPO-5-DC vaccine. After vaccination, the frequency of HIV-specific Tregs decreased (from 69.3 at week -4 to 31.7% at week 16) and inversely correlated with HIV-specific IFN-γ-producing cells (r=-0.64, p=0.002). We show that therapeutic immunization skewed the HIV-specific response from regulatory to effector phenotype which impacts on the magnitude of viral replication following ATI.

We investigated the relative effect of amenorrhea and insulin-like growth factor-I (sIGF-I) levels on cancellous and cortical bone density and size. We investigated 66 adult women with anorexia nervosa. Lumbar spine and proximal femur bone mineral density was measured by DXA. We calculated bone mineral apparent density. Structural geometry of the spine and the hip was determined from DXA images. Weight and BMI, but not height, as well as bone mineral content and density, but not area and geometry parameters, were lower in patients with anorexia nervosa as compared with the control group. Amenorrhea, disease duration, and sIGF-I were significantly associated with lumbar spine and proximal femur BMD. In a multiple regression model, we found that sIGF-I was the only significant independent predictor of proximal femur BMD, while duration of amenorrhea was the only factor associated with lumbar spine BMD. Finally, femoral neck bone mineral apparent density, but not hip geometry variables, was correlated with sIGF-I. In anorexia nervosa, spine BMD was related to hypogonadism, whereas sIGF-I predicted proximal femur BMD. The site-specific effect of sIGF-I could be related to reduced volumetric BMD rather than to modified hip geometry.