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Otfried Kistner,
Brian A Crowe,
Walter Wodal,
Astrid Kerschbaum,
Helga Savidis-Dacho,
Nicolas Sabarth,
Falko G Falkner,
Ines Mayerhofer,
Wolfgang Mundt,
Manfred Reiter, Leopold Grillberger,
Christa Tauer,
Michael Graninger,
Alois Sachslehner,
Michael Schwendinger,
Peter Brühl,
Thomas R Kreil,
Hartmut J Ehrlich,
P Noel Barrett
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ABSTRACT: The recent emergence and rapid spread of a novel swine-derived H1N1 influenza virus has resulted in the first influenza pandemic of this century. Monovalent vaccines have undergone preclinical and clinical development prior to initiation of mass immunization campaigns. We have carried out a series of immunogenicity and protection studies following active immunization of mice, which indicate that a whole virus, nonadjuvanted vaccine is immunogenic at low doses and protects against live virus challenge. The immunogenicity in this model was comparable to that of a whole virus H5N1 vaccine, which had previously been demonstrated to induce high levels of seroprotection in clinical studies. The efficacy of the H1N1 pandemic vaccine in protecting against live virus challenge was also seen to be equivalent to that of the H5N1 vaccine. The protective efficacy of the H1N1 vaccine was also confirmed using a severe combined immunodeficient (SCID) mouse model. It was demonstrated that mouse and guinea pig immune sera elicited following active H1N1 vaccination resulted in 100% protection of SCID mice following passive transfer of immune sera and lethal challenge. The immune responses to a whole virus pandemic H1N1 and a split seasonal H1N1 vaccine were also compared in this study. It was demonstrated that the whole virus vaccine induced a balanced Th-1 and Th-2 response in mice, whereas the split vaccine induced mainly a Th-2 response and only minimal levels of Th-1 responses. These data supported the initiation of clinical studies with the same low doses of whole virus vaccine that had previously been demonstrated to be immunogenic in clinical studies with a whole virus H5N1 vaccine.
PLoS ONE 01/2010; 5(2):e9349. · 4.09 Impact Factor
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ABSTRACT: Mammalian cells are the expression system of choice for therapeutic proteins, especially those requiring complex post-translational modifications. Traditionally, these cells are grown in medium supplemented with serum and other animal- or human-derived components to support viability and productivity. Such proteins are also typically added as excipients and stabilizers in the final drug formulation. However, the transmission of hepatitis B in the 1970s and of hepatitis C and HIV in the 1980s through plasma-derived factor VIII concentrates had catastrophic consequences for hemophilia patients. Thus, due to regulatory concerns about the inherent potential for transmission of infectious agents as well as the heterogeneity and lack of reliability of the serum supply, a trend has emerged to eliminate the use of plasma-derived additives in the production and formulation of recombinant protein therapeutics. This practice began with products used in the treatment of hemophilia and is progressively expanding throughout the entire industry. The plasma-free method of producing recombinant therapeutics is accomplished by the use of both cell culture media and final product formulations that do not contain animal- or human-derived additives. A number of recombinant therapeutic proteins for the treatment of several different diseases have been produced by plasma-free processes, with the objective of improving safety by eliminating blood-borne pathogens or by reducing immunogenicity. This review describes the factors that drove the development of plasma-free protein therapeutics and provides examples of advances in manufacturing that have made possible the removal of human and animal-derived products from all steps of recombinant protein production.
Biotechnology Journal 03/2009; 4(2):186-201.
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Otfried Kistner,
M Keith Howard,
Martin Spruth,
Walter Wodal,
Peter Brühl,
Marijan Gerencer,
Brian A Crowe,
Helga Savidis-Dacho,
Ian Livey,
Manfred Reiter,
Ines Mayerhofer,
Christa Tauer, Leopold Grillberger,
Wolfgang Mundt,
Falko G Falkner,
P Noel Barrett
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ABSTRACT: The rapid spread and the transmission to humans of avian influenza virus (H5N1) have induced world-wide 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 also 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.
Vaccine 09/2007; 25(32):6028-36. · 3.77 Impact Factor
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ABSTRACT: In the last years evidence has been provided for the importance of B cells in the pathogenesis of rheumatoid arthritis (RA). Several studies have supported the concept that humoral immunity, manifested by the production of autoantibodies, such as rheumatoid factors (RFs), plays a significant role in the course of the disease. Specific targeting of autoantibody-producing B cells, such as RF-producing B cells, should therefore be a promising new approach in the treatment of RA. We used a mouse model to induce human RF responses and asked the question whether oral treatment with the antigen (human IgG) recognized by RFs could induce immune tolerance to RF responses. Balb/c mice were orally treated with polyvalent human IgG before and after immunization with insoluble immune complexes (ICs) that triggered the induction of RFs. Serum titers of RFs were significantly reduced after both primary and booster immunization when human IgG was given as a single oral dose or continuously in drinking water. Continuous treatment with human IgG even prevented booster effects on RFs when treatment started after primary immunization. Treatment with IgG fragments provided evidence that the observed effect of human IgG was mediated by the Fc part and not the Fab part of IgG. Furthermore, transfer of spleen cells obtained from mice after oral treatment with human IgG suppressed RF responses in recipient mice. These data give promising indications that oral human IgG might represent an alternative approach for immunosuppressive B-cell targeted therapies in RA.
International Immunopharmacology 04/2007; 7(3):351-9. · 2.38 Impact Factor
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Martin Spruth,
Otfried Kistner,
Helga Savidis-Dacho,
Elisabeth Hitter,
Brian Crowe,
Marijan Gerencer,
Peter Brühl, Leopold Grillberger,
Manfred Reiter,
Christa Tauer,
Wolfgang Mundt,
P Noel Barrett
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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.
Vaccine 02/2006; 24(5):652-61. · 3.77 Impact Factor
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ABSTRACT: A mutant derivative of human prothrombin in which active site aspartate at position 419 is replaced by an asparagine (D419N-prothrombin)
has been designed, expressed in recombinant Chinese hamster ovary cells, and purified to homogeneity. D419N-prothrombin was
converted to the related molecules D419N-meizothrombin and D419N-thrombin by limited proteolysis by Echis carinatus and Oxyuranus scutellatus venom protease, respectively, and affinity-purified using an immobilized modified C-terminal hirudin-derived peptide. Neither
D419N-thrombin nor D419N-meizothrombin exhibited thrombin activity. Titration resulted in no detection of the active site,
but binding to the most specific thrombin inhibitor, hirudin, was conserved in both proteins. In vitro examinations showed that D419N-thrombin and D419N-meizothrombin bind to immobilized hirudin, neutralize hirudin in human
blood plasma as well as in the purified system, and reactivate the thrombin-hirudin complex. Animal model studies confirmed
that D419N-thrombin and D419N-meizothrombin act as hirudin antagonist in blood circulation without detectable effects on the
coagulation system. Thus, both D419N-thrombin and D419N-meizothrombin combine for the first time hirudin-neutralizing properties
with the advantages of recombinant production of human coagulation factors.
Journal of Biological Chemistry 09/1996; 271(39):23737-23742. · 4.77 Impact Factor
