A measles virus-based vaccine induces robust chikungunya virus-specific CD4⁺ T-cell responses in a phase II clinical trial

Chikungunya virus (CHIKV) is an alphavirus transmitted by mosquitos that causes a debilitating disease characterized by fever and long-lasting polyarthralgia. To date, no vaccine has been licensed, but multiple vaccine candidates are under evaluation in clinical trials. One of these vaccines is based on a measles virus vector encoding for the CHIKV structural genes C, E3, E2, 6K, and E1 (MV-CHIK), which proved safe in phase I and II clinical trials and elicited CHIKV-specific antibody responses in adult measles seropositive vaccine recipients. Here, we predicted T-cell epitopes in the CHIKV structural genes and investigated whether MV-CHIK vaccination induced CHIKV-specific CD4+ and/or CD8+ T-cell responses. Immune-dominant regions containing multiple epitopes in silico predicted to bind to HLA class II molecules were found for four of the five structural proteins, while no such regions were predicted for HLA class I. Experimentally, CHIKV-specific CD4+ T-cells were detected in six out of twelve participants after a single MV-CHIK vaccination and more robust responses were found 4 weeks after two vaccinations (ten out of twelve participants). T-cells were mainly directed against the three large structural proteins C, E2 and E1. Next, we sorted and expanded CHIKV-specific T cell clones (TCC) and identified human CHIKV T-cell epitopes by deconvolution. Interestingly, eight out of nine CD4+ TCC recognized an epitope in accordance with the in silico prediction. CHIKV-specific CD8+ T-cells induced by MV-CHIK vaccination were inconsistently detected. Our data show that the MV-CHIK vector vaccine induced a functional transgene-specific CD4+ T cell response which, together with the evidence of neutralizing antibodies as correlate of protection for CHIKV, makes MV-CHIK a promising vaccine candidate in the prevention of chikungunya.</p

Захист суспільної моралі в інформаційному суспільстві

До питання ролі держави та її інституцій в захисті суспільної моралі, в удосконаленні чинного законодавства з питань захисту суспільної моралі.К вопросу роли государства и его институций в защите общественной морали, в усовершенствовании действующего законодательства по вопросам защиты общественной морали.As to the role of the state and its institutions in the protection of social morality, in improvement of current legislation on protection of social morality

Primary Human Influenza B Virus Infection Induces Cross-Lineage Hemagglutinin Stalk-Specific Antibodies Mediating Antibody-Dependent Cellular Cytoxicity.

Influenza A virus (IAV) and influenza B virus (IBV) cause substantial morbidity and mortality during annual epidemics. Two distinct lineages of IBV are distinguished, based on variation in hemagglutinin (HA): B/Victoria/2/87-like (B/Vic) and B/Yamagata/16/88-like (B/Yam). Here, we show that, in humans, primary IBV infection with either lineage induces HA-specific antibody-dependent cellular cytotoxicity (ADCC)-mediating antibodies. IBV infection induced antibodies specific to the HA head and stalk, but only HA stalk-specific antibodies mediated ADCC efficiently and displayed cross-reactivity with IBV of both lineages. This corresponds to recent findings that 2 points of contact between the effector and target cell (ie, HA and sialic acid, respectively, and the fragment crystallizable [Fc] domain and Fcγ receptor IIIα, respectively) are required for efficient ADCC activity and that antibodies specific for the receptor-binding site located in the head domain of HA therefore fail to mediate ADCC. Potentially, ADCC-mediating antibodies directed to the HA stalk of IBV contribute to cross-protective immunity to IBV of both lineages

Design and use of conditional MHC class I ligands

Major histocompatibility complex (MHC) class I molecules associate with a variety of peptide ligands during biosynthesis and present these ligands on the cell surface for recognition by cytotoxic T cells. We have designed conditional MHC ligands that form stable complexes with MHC molecules but degrade on command, by exposure to a defined photostimulus. 'Empty MHC molecules' generated in this manner can be loaded with arrays of peptide ligands to determine MHC binding properties and to monitor antigen-specific T-cell responses in a high-throughput manner. We document the value of this approach by identifying cytotoxic T-cell epitopes within the H5N1 influenza A/Vietnam/1194/04 genom

The number of glycosylation sites present on HA determines the virus infection rates in DC-SIGN expressing cells.

<p>MDCK (A) and Vero (B) cells, transfected with the DC-SIGN gene (black bars) or not (white bars) were treated with neuraminidase from vibrio cholerae and GolgiStop for 30 minutes to remove sialic acids from the cell surface. These cells were subsequently inoculated with A/Netherlands/26/07, A/Netherlands/26/07-Δ125, A/Netherlands/602/09, A/Netherlands/602/09-Δ276 or A/Netherlands/602/09-VN54 N125 N160. The percentage of infected cells relative to the positive control (untreated cells still possessing sialic acid) was assessed after detecting infected cells using a FITC-labeled antibody to the viral nucleoprotein and flow-cytometry.</p

Replication kinetics of viruses GFP-H1 and L194AY195F-GFP-H1 in MDCK and DC-SIGN-expressing MDCK cells.

<p>After transfection of 293T cells with reverse genetics plasmids, culture supernatants of influenza viruses GFP-H1 (A, C, E, G) and L194AY195F-GFP-H1 (B, D, F, H) virus passaged in MDCK (A–D) and MDCK-DC-SIGN (E–H) cells were obtained and used to inoculate MDCK (solid symbols) or MDCK DC-SIGN cells (open symbols) at a moi of 0.01. At the indicated time points post inoculation culture supernatant were tested for the infectious virus titers to determine the replication kinetics. Virus L194AY195F-GFP-H1 could not be rescued in MDCK cells.</p

Expression of DC-SIGN supports replication of influenza A viruses in the absence of sialic acids.

<p>MDCK (A and C) and Vero cells (B and D) transfected with the DC-SIGN gene (black bars) or not (white bars), were treated with neuraminidase from vibrio cholerae and GolgiStop for 30 minutes to remove sialic acids from the cell surface. These cells were subsequently inoculated with five different A/H1N1 viruses (A and B) and four A/H3N2 viruses (C and D). The percentage infected cells relative to the untreated control cells, still possessing sialic acid, was assessed after detecting infected cells using a FITC-labelled antibody to the viral nucleoprotein and flow-cytometry. To confirm that the entry was mediated via DC-SIGN, Vero and Vero DC-SIGN were treated with neuraminidase from vibrio cholerae for 30 minutes to remove sialic acids from the cell surface and incubated with or without antibodies to DC-SIGN or an isotype control antibody as indicated (E and F). These cells were subsequently inoculated with influenza viruses. NL/312/03 and USSR/90/77. The percentage of infected cells compared to the positive control (untreated cells, still possessing sialic acid) was assessed as described above.</p

Schematic representation of the passage history and infection experiments with influenza viruses GFP-H1 and L194AY195F-GFP-H1.

<p>Schematic representation of the passage history and infection experiments with influenza viruses GFP-H1 and L194AY195F-GFP-H1.</p

Number of putative N linked glycosylation sites present in HA1 and in HA2 of the viruses used in this study.

<p>The software NetNGlyc (<a href="http://www.cbs.dtu.dk/services/NetNGlyc/" target="_blank">http://www.cbs.dtu.dk/services/NetNGlyc/</a>)was used to predict the number of glycosylation sites that will be utilized.</p

GFP expression after infection with L194AY195F-GFP and GFP-H1 in MDCK and DC-SIGN-expressing MDCK cells.

<p>MDCK and MDCK DC-SIGN cells were inoculated with influenza viruses GFP-H1 (A) and L194AY195F-GFP-H1 (B) at a MOI of 0.01 (solid lines). Both viruses were passaged in MDCK-DC-SIGN cells two or three times as indicated. Twenty-four hours post inoculation the cells were tested for GFP expression by flow cytometry. Uninfected cells were included as negative controls (dotted lines). Infection experiments with GFP-H1 virus passaged in MDCK cells essentially gave the same results as the virus passaged in MDCK DC-SIGN cells (data not shown).</p