Recombinant hemagglutinin of swine H1N1 influenza virus expression in the insect cells: Formulation in Montanide ISA71 adjuvant and the potency studies (original) (raw)
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Iranian Journal of Allergy, Asthma and Immunology, 2020
The H1N1 influenza virus is known as a serious pandemic threat across the globe. Vaccination is one of the most effective methods of protection against this virus and the way to reduce the seasonal pandemic risk. The commercial vaccine does not adequately respond to pandemic strains. This study examines the potential function of formulated H1N1 hemagglutinin with MF59 adjuvant against A/PR/8/34 (H1N1). To this end, a recombinant hemagglutinin (rHA) gene of influenza A virus was designed and expressed in SF9 cell by the Baculovirus expression system. Four groups of mice were immunized by rHA in combination with MF59, Alum adjuvant, and virus split only. The immunized mice subsequently used for the humoral immune assay and the results compared with untreated mice (negative group). Besides, both treated and control mice groups were challenged with mouse-adapted influenza virus A/PR/8/34(H1N1) through the intranasal drop. Bodyweight, survival, temperature variation, and the medical con...
2013
Recombinant hemagglutinin (rHA) and neurominidase (rNA) developed in our investigation are amino acid sequence consensus variants of H1N1 2009 subtype influenza virus strain, also including immunogenic epitopes typical for other influenza virus subtypes (H3N1 and H5N1). Substitutions were made: typical for Russian virus isolates (in HA -S220T, NA -D248N) and in active centers of molecules -R118L, R293L, R368L; C92S, C417S to increase recombinant proteins stability in E. coli. The aim of the present work was to study immunogenicity of the obtained rHA and rNA. Material/Methods: Fragments aa 83-469 of NA and aa 61-287 of HA were chosen because they include the main B-cell epitopes and are the minimal structures for correct folding of target proteins. The designed nucleotide sequences were synthesized and purified and the expression of rNA and rNA were analyzed. For immunization and virus challenge we used influenza viruses A/California/04/2009 (H1N1), A/PR/8/34 (H1N1), A/Perth/16/2009 (H3N2), A/Chicken/Kurgan/05/2005 R.G. (H5N1), and B/Florida/04/2006. Specific IgG levels were determined by ELISA in 96-well ELISA plates. Significant differences of survival in mouse groups were analyzed by Mantel-Cox (logrank) and Gehan-Breslow-Wilcoxon tests.
Virus Research, 2011
Recent outbreaks of highly pathogenic avian influenza (HPAI) H5N1 viruses in poultry and their subsequent transmission to humans have highlighted an urgent need to develop preventive vaccines in the event of a pandemic. In this paper we constructed recombinant adenovirus (rAd)-vectored influenza vaccines expressing different forms of H5 hemagglutinin (HA) from the A/Vietnam/1194/04 (VN/1194/04) virus, a wild-type HA, a sequence codon-optimized HA and a transmembrane (TM) domain-truncated HA. Compared to the rAd vectors expressing the wild-type HA (rAd-04wtHA) and the TM-truncated form of HA (rAd-04optHA-dTM), the rAd vectored vaccine with the sequence codon-optimized HA (rAd-04optHA) showed a tendency to induce much higher hemagglutinin inhibition (HI) antibody titers in mice immunized with a prime-boost vaccine. Furthermore, administration of the rAd-04optHA vaccine to mice could elicit cross-reactive immune responses against the antigenically distinct HK/482/97 virus. Additionally, we constructed another vector containing the codon-optimized HA of the A/Hong Kong/482/97 (HK/482/97) virus. Administration of a bivalent immunization formulation including the rAd-04optHA and rAd-97optHA vaccines to mice induced a stronger immune response against HK/482/97 virus than the monovalent formulation. Taken together, these findings may have some implications for the development of rAd-vectored vaccines in the event of the pandemic spread of HPAI.
Rapid Development of an Efficacious Swine Vaccine for Novel H1N1
PLoS Currents, 2009
Recombinant hemagglutinin (HA) from a novel H1N1 influenza strain was produced using an alphavirus replicon expression system. The recombinant HA vaccine was produced more rapidly than traditional vaccines, and was evaluated as a swine vaccine candidate at different doses in a challenge model utilizing the homologous influenza A/California/04/2009 (H1N1) strain. Vaccinated animals showed significantly higher specific antibody response, reduced lung lesions and viral shedding, and higher average daily gain when compared to non-vaccinated control animals. These data demonstrate that the swine vaccine candidate was efficacious at all of the evaluated doses. triple-reassortant swine influenza subtype H1 in humans from 2005-February 2009 [3]. They found 11 sporadic cases, and all 11 patients recovered after showing clinical influenza symptoms. Nine of these 11 patients had known exposure to pigs, most of which were ill, either at agricultural fairs or at hog farms. These results mirror studies showing increased antibody titers to swine influenza viruses among hog farm workers and family members [4][5]. This demonstrates the role that human and swine interaction can play in the creation of novel influenza viruses, and thus the need for efficacious swine influenza vaccines. Currently, novel H1N1 influenza has been detected in swine in several different countries
Journal of Virology, 2005
In 2003, an outbreak of highly pathogenic avian influenza occurred in the Netherlands. The avian H7N7 virus causing the outbreak was also detected in 88 humans suffering from conjunctivitis or mild respiratory symptoms and one person who died of pneumonia and acute respiratory distress syndrome. Here we describe a mouse model for lethal infection with A/Netherlands/219/03 isolated from the fatal case. Because of the zoonotic and pathogenic potential of the H7N7 virus, a candidate vaccine carrying the avian hemagglutinin and neuraminidase proteins produced in the context of the high-throughput vaccine strain A/PR/8/34 was generated by reverse genetics and tested in the mouse model. The hemagglutinin gene of the recombinant vaccine strain was derived from a low-pathogenicity virus obtained prior to the outbreak from a wild mallard. The efficacy of a classical nonadjuvanted subunit vaccine and an immune stimulatory complex-adjuvanted vaccine was compared. Mice receiving the nonadjuvanted vaccine revealed low antibody titers, lack of clinical protection, high virus titers in the lungs, and presence of virus in the spleen, liver, kidneys, and brain. In contrast, mice receiving two doses of the immune stimulatory complex-adjuvanted vaccine revealed high antibody titers, clinical protection, ϳ1,000-fold reduction of virus titers in the lungs, and rare detection of the virus in other organs. This is the first report of an H7 vaccine candidate tested in a mammalian model. The data presented suggest that vaccine candidates based on low-pathogenicity avian influenza A viruses, which can be prepared ahead of pandemic threats, can be efficacious if an effective adjuvant is used. @erasmusmc.nl. † E.D.W. and V.J.M. have contributed equally to the results of this study.
Development of recombinant protein-based influenza vaccine
Journal of Chromatography A, 2006
The influenza virus surface glycoprotein antigen neuraminidase (NA) is a crucial viral enzyme with many potential medical applications; therefore, the development of efficient upstream and downstream processing strategy for the expression and purification of NA is of high importance. In the present work the NA gene from the H1N1 influenza virus strain A/Beijing/262/95 was cloned from viral RNA and expressed in expresSF+ insect cells using the baculovirus expression vector system (BVES). A limited affinity-ligand library was synthesized and evaluated for its ability to bind and purify the recombinant H1N1 neuraminidase. Affinity-ligand design was based on mimicking the interactions of the lock-and-key (LAK) motif (Phe-Gly-Gln), a common structural moiety found in the subunit interface of glutathione S-transferase I (GST I), and plays an important structural role in subunit-subunit recognition. Solid-phase combinatorial chemistry was used to synthesize 13 variants of the lock-and-key lead ligand (Phe-Trz-X, where X was selected ␣-amino acid) using the 1,3,5-triazine moiety (Trz) as the scaffold for assembly. One immobilized ligand, bearing phenylalanine and isoleucine linked on the chlorotriazine ring (Phe-Trz-Ile), displayed high affinity for NA. Absorption equilibrium and molecular modeling studies were carried out to provide a detailed picture of Phe-Trz-Ile interaction with NA. This LAK-mimetic affinity adsorbent was exploited in the development of a facile purification protocol for NA, which led to 335-fold purification in a single-step. The present purification procedure is the most efficient reported so far for recombinant NA. (N.E. Labrou).
PLoS ONE, 2013
Despite the advantages of DNA vaccines, overcoming their lower efficacy relative to that of conventional vaccines remains a challenge. Here, we constructed a human endogenous retrovirus (HERV) envelope-coated, nonreplicable, baculovirus-based HA vaccine against swine influenza A/California/04/2009(H1N1) hemagglutin (HA) (AcHERV-sH1N1-HA) as an alternative to conventional vaccines and evaluated its efficacy in two strains of mice, BALB/c and C57BL/6. A commercially available, killed virus vaccine was used as a positive control. Mice were intramuscularly administered AcHERV-sH1N1-HA or the commercial vaccine and subsequently given two booster injections. Compared with the commercial vaccine, AcHERV-sH1N1-HA induced significantly higher levels of cellular immune responses in both BALB/c and C57BL/6 mice. Unlike cellular immune responses, humoral immune responses depended on the strain of mice. Following immunization with AcHERV-sH1N1-HA, C57BL/6 mice showed HAspecific IgG titers 10-to 100-fold lower than those of BALB/c mice. In line with the different levels of humoral immune responses, the survival of immunized mice after intranasal challenge with sH1N1 virus (A/California/04/2009) depended on the strain. After challenge with 10-times the median lethal dose (MLD 50) of sH1N1 virus, 100% of BALB/c mice immunized with the commercial vaccine or AcHERV-sH1N1-HA survived. In contrast, C57BL/6 mice immunized with AcHERV-sH1N1-HA or the commercial vaccine showed 60% and 70% survival respectively, after challenge with sH1N1 virus. In all mice, virus titers and results of histological analyses of lung tissues were consistent with the survival data. Our results indicate the importance of humoral immune response as a major defense system against influenza viral infection. Moreover, the complete survival of BALB/c mice immunized with AcHERV-sH1N1-HA after challenge with sH1N1 virus suggests the potential of baculoviral vector-based vaccines to achieve an efficacy comparable to that of killed virus vaccines.
Recombinant Influenza A Virus Vaccines for the Pathogenic Human A/Hong Kong/97 (H5N1) Viruses
The Journal of Infectious Diseases, 1999
Recombinant reassortment technology was used to prepare H5N1 influenza vaccine strains containing a modified hemagglutinin (HA) gene and neuraminidase gene from the A/Hong Kong/156/97 and A/Hong Kong/483/97 isolates and the internal genes from the attenuated cold-adapted A/Ann Arbor/6/60 influenza virus strain. The HA cleavage site (HA1/HA2) of each H5N1 isolate was modified to resemble that of "low-pathogenic" avian strains. Five of 6 basic amino acids at the cleavage site were deleted, and a threonine was added upstream of the remaining arginine. The H5 HA cleavage site modification resulted in the expected trypsin-dependent phenotype without altering the antigenic character of the H5 HA molecule. The temperature-sensitive and cold-adapted phenotype of the attenuated parent virus was maintained in the recombinant strains, and they grew to 10 8.5-9.4 EID 50 /mL in eggs. Both H5N1 vaccine virus strains were safe and immunogenic in ferrets and protected chickens against wild-type H5N1 virus challenge.