Viral Genomics and Vaccination - CNRS URA 3015  


  HEADDr. TANGY Frédéric / frederic.tangy@pasteur.fr
  MEMBERSBOURAI Mehdi PhD Student / CAIGNARD Grégory PhD Student / COMBREDET Chantal Technician IP / FEVRIER Michèle CR1 CNRS / GAU Mireille Secretary IP / GUERBOIS Mathilde PhD Student / KOMAROVA Anastasia Post-doc Researcher / LUCAS-HOURANI Marianne Engineer IP / MESEL-LEMOINE Mariana Post-doc Researcher / MILED Chaouki Post-doc Researcher / MILLET Jean PhD Student / NAJBURG Valérie Technician IP / RUFFIE Claude Technician IP / VABRET Nicolas PhD Student / VIDALAIN Pierre-Olivier CR2 CNR


  Annual Report

Bivalent vaccines derived from measles vaccine

The measles vaccine has been administered to hundreds of millions of children over the last 30 years and has proved both effective and safe, inducing life-long immunity after only one injection. It is produced on a large scale in many countries and is distributed at low cost through the Expanded Program on Immunization of WHO. These characteristics led us to consider its use as a pediatric vector designed to protect children against measles and simultaneously to immunize them against other infections, such as AIDS, malaria or flaviviral diseases. These affordable recombinant vaccines might be attractive for the developing word. With this aim, we cloned the genome of the attenuated Schwarz strain of the measles virus (MV) and made it into a vector by inserting new transcriptional units. We demonstrated the strong and stable expression from this vector of genes encoding proteins from HIV, West Nile or dengue viruses. These vectors are immunogenic in mice transgenic for MV receptor and in macaques. Recombinant vaccines expressing the HIV Env gene with deletions of the hypervariable sequences induced neutralizing antibodies and T-cell responses after a single injection. Pre-existing immunity to the vector did not affect the immunogenic potential of the vectors, since two successive injections led to the induction of anti-HIV antibodies in mice and macaques previously vaccinated with standard vaccine. An immunization/challenge experiment performed on macaques using MV-SHIV89.6 vectors demonstrated strong cellular responses and reduction of viral load during primo-infection after a rectal SHIV89.6challenge. A clinical development program is running in collaboration with GlaxoSmithKline to evaluate in phase I/II clinical trials the safety of MV-HIV vector and its capacity to induce immune responses to HIV in humans. The vaccine candidate has been produced and proved to be immunogenic in mice and macaques. A clinical GMP lot is currently evaluated in a large toxcicity study in primates before Phase I testing. New MV vectors promoting the intense budding of assembled HIV1 Gag virus-like-particles coated with HIV1 Env proteins have been produced and proved to be immunogenic. In a similar program, we have constructed a MV vector expressing a new combined dengue antigen derived from the four serotypes of dengue virus and demonstrated its capacity to induce durable specific neutralizing antibodies to dengue virus. The efficacy of this vaccine is currently evaluated in macaques. Other programs are also developed in the laboartory using the same technology, particularly a MV-malaria vaccination program in collaboration with New York University and GlaxoSmithKline.

Proteomics of virus-host interactions

To identify virus-host protein-protein interactions that determine virulence and pathogenesis, we have developed a large-scale mapping project based on high-throughput technologies, including recombination-based cloning and state-of-the-art yeast two-hybrid. Our screening platform is now fully operational and more than 100 screens a year can be performed. Using this tool, we have identified hundreds of virus-host protein-protein interactions, taking advantage of the unique collection of viruses available at Institut Pasteur. Besides collaborative programs developed inside or outside of this institution, our efforts now focus on two viral families of higher interest for our research group : Paramyxoviridae, including measles, Nipah, mumps, human parainfluenza 3 and Tioman viruses, and arboviruses from flavivirus and alphavirus genera including chikungunya, Sindbis, Semliki Forest and yellow fever viruses. Virus-host protein-protein interactions identified by yeast two-hybrid will be validated in the near future using a high-throughput binding assay recently developed in our laboratory. Among identified interactions, we recently demonstrated that the V protein of measles virus targets STAT1, STAT2 and Jak1 to inhibit type I interferon signalling pathway, thereby blocking the innate antiviral immune response. In addition to this fundamental research program, we engaged in the large-scale screening of chemical compound libraries to identify immunostimulators of the antiviral immune response and specific inhibitors of alphavirus virulence factor nsP2.

Keywords: measles, HIV, Chikungunya, dengue, vaccine, interactome, yeast two-hybrid, antiviral molecules

Gvv.jpg

Fig. 1. Budding of HIV-1 virus-like-particles from Vero cells infected with measles-HIV live recombinant vaccines. In E and F, anti-Env immunogold staining with 10-nanometer colloidal gold particles shows the incorporation of gp160 Env proteins onto the particles

Fig. 2. Yeast two-hybrid mapping of MV-V binding region to STAT1 using a systemic deletion-based mapping procedure. Fragments of the MV-V were generated by PCR using a matrix combination of specific primers, and introduced into Gal4-DB vector by gap-repair in yeast cells expressing STAT1 fused to Gal4-AD. Yeast cells were grown on selective medium lacking histidine and supplemented with 10 mM of 3-amino-triazole (3-AT) to test the interaction-dependent transactivation of HIS3 reporter gene. Vertical and horizontal axes indicate the first and the last amino acid residues of each fragment tested, respectively. (1), (2) and (3) correspond to three iterations of this process. The third iteration led to the identification of a 11 amino acid peptide encompassing position 110 to 120 of MV-V as the minimal STAT1 binding peptide.



  Publications

Ozden S., Lucas-Hourani M., Ceccaldi P.-E., Basak A., Valentine M., Benjannet S., Hamelin J., Jacob Y., Mamchaoui K., Mouly V., Desprès P., Gessain A., Butler-Browne G., Chrétien M., Tangy F., Vidalain P.-O, Seidah N.G. 2008. Inhibition of Chikungunya virus infection in cultured human muscle cells by furin inhibitors: impairment of the maturation of the E2 surface glycoprotein. J Biol Chem., 283(32):21899-908. PMID: 18559340

Brandler S., Lucas-Hourani M., Moris A., Frenkiel M.-P., Combredet C., Février M., Bedouelle H., Schwartz O., Desprès P., & Tangy F. 2007. Pediatric Measles Vaccine Expressing a Dengue Antigen Induces Durable Serotype-specific Neutralizing Antibodies to Dengue Virus. PLoS Neglected Tropical Diseases, 1(3):e96. PMID: 18160988

Caignard G., Guerbois M., Labernardière J.-L., Jacob Y., Jones L.M., Wild F., Tangy F., & Vidalain P.-O. 2007. Measles virus V protein blocks Jak1-mediated phosphorylation of STAT1 to escape IFN-α/β signaling. Virology, 368(2):351-62. PMID: 17686504

Lorin C., Delebecque F., Labrousse V., Da Silva L., Lemonnier F., Brahic M., & Tangy F. 2005. A recombinant live attenuated measles vaccine vector primes effective HLA-A0201-restricted cytotoxic T lymphocytes and broad neutralizing antibodies against HIV-1 conserved epitopes. Vaccine, 23(36):4463-72. PMID: 15993518

Despres P., Combredet C., Frenkiel M.-P., Brahic M. & Tangy F. 2005. Live measles vaccine expressing the secreted form of the West Nile virus envelope glycoprotein protects against West Nile virus encephalitis. J. Infect. Dis., 191(2):207-14. PMID: 15609230





Activity Reports 2009 - Institut Pasteur
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