No abstract

A two-dose vaccine regimen of either 7.5 microg or 15 microg of hemagglutinin antigen without adjuvant induced neutralizing antibodies against diverse H5N1 virus strains in a high percentage of subjects, suggesting that this may be a useful H5N1 vaccine. (ClinicalTrials.gov number, NCT00349141.)

The development of cell culture systems for virus propagation has led to major advances in virus vaccine development. Primary and diploid cell culture systems are now being replaced by the use of continuous cell lines (CCLs). These substrates are gaining increasing acceptance from regulatory authorities as improved screening technologies remove fears regarding their potential oncogenic properties. The Vero cell line is the most widely accepted CCL by regulatory authorities and has been used for over 30 years for the production of polio and rabies virus vaccines. The recent licensure of a Vero cell-derived live virus vaccine (ACAM2000, smallpox vaccine) has coincided with an explosion in the development of a range of new viral vaccines, ranging from live-attenuated pediatric vaccines against rotavirus infections to inactivated whole-virus vaccines against H5N1 pandemic influenza. These developments have illustrated the value of this cell culture platform in the rapid development of vaccines against a range of virus diseases.

No abstract

A double-inactivated, candidate whole virus vaccine against severe acute respiratory syndrome associated coronavirus (SARS-CoV) was developed and manufactured at large scale using fermenter cultures of serum protein free Vero cells. A two step inactivation procedure involving sequential formaldehyde and U.V. inactivation was utilised in order to ensure an extremely high safety margin with respect to residual infectivity. The immunogenicity of this double-inactivated vaccine was characterised in the mouse model. Mice that were immunised twice with the candidate SARS-CoV vaccine developed high antibody titres against the SARS-CoV spike protein and high levels of neutralising antibodies. The use of the adjuvant Al(OH)3 had only a minor effect on the immunogenicity of the vaccine. In addition, cell mediated immunity as measured by interferon-gamma and interleukin-4 stimulation, was elicited by vaccination. Moreover, the vaccine confers protective immunity as demonstrated by prevention of SARS-CoV replication in the respiratory tract of mice after intranasal challenge with SARS-CoV. Protection of mice was correlated to antibody titre against the SARS-CoV S protein and neutralising antibody titre.

The rapid spread and the transmission to humans of avian influenza virus (H5N1) has induced worldwide fears of a new pandemic and raised concerns over the ability of standard influenza vaccine production methods to rapidly supply sufficient amounts of an effective vaccine. We report here on a robust and flexible strategy which uses wild-type virus grown in a continuous cell culture (Vero) system to produce an inactivated whole virus vaccine. Candidate vaccines based on clade 1 and clade 2 influenza H5N1 strains were developed and demonstrated to be highly immunogenic in animal models. The vaccines induce cross-neutralising antibodies, highly cross-reactive T-cell responses and are protective in a mouse challenge model not only against the homologous virus but against other H5N1 strains, including those from another clade. These data indicate that cell culture-grown, whole virus vaccines, based on the wild-type virus, allow the rapid high yield production of a candidate pandemic vaccine.

SUMMARYAn analysis of the nucleoprotein (NP) of 29 different influenza A viruses by phosphopeptide fingerprinting revealed three prototype patterns. The first, which was a complex pattern consisting of six to seven phosphopeptides, another which was relatively simple consisted of two or three phosphopeptides, and a third one which was complex but was missing the main phosphopeptide shared by the two other patterns. Phosphoserine was the only labelled phosphamino acid detected. A tentative deduction of two of the phosphate attachment sites (serine residues at positions 3 and 473) could be made by comparison of the known amino acid sequences of the NPs of 25 strains. No correlation was found between species specificity or subtype or year of isolation of the strains. During the infectious cycle the fingerprint underwent significant changes, indicating subtle phosphorylation and dephosphorylation of the NP at various stages during viral multiplication. Most of the phosphopeptides were metabolically stable; however one major phosphopeptide, which was not found in the NP of mature virions, exhibited a high turnover (presumably serine at position 3). The phosphopeptide fingerprint could be significantly influenced in vivo by the specific stimulation of cellular protein kinase C by the phorbol ester 12-O-tetradecanolyphorbol 13-acetate or by its inhibition with the isoquinoline sulphonamide H7.

Our investigation verifies the safety margins of plasma derivatives against a potential transmission of WNV and that the model virus concept is valid for predicting the behavior of closely related viruses.