Graduated from Kyoto University, Graduate School of Agriculture, Japan M.S degree on Molecular Biology and Microbiology.
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1 Dr.Moto Esaki Senior Chief Reseacher Ceva Japan K.K Graduated from Kyoto University, Graduate School of Agriculture, Japan M.S degree on Molecular Biology and Microbiology : Joined Biology department of Zen Corporation (now Ceva Japan) 2000 : Transferred to Ceva Biomune and engaged in development and licensing of Vectormune vaccine. Later became Manager for Recombinant Vaccine Research 2011 : Rejoin Ceva Japan involve in research on new generation vaccines. Topic : Innovative Vector Vaccine, A new tool to prevent disease.
2 Innovative Vector Vaccine A New Tool to Prevent Diseases Moto Esaki, Kristi Moore Dorsey and Atsushi Yasuda CEVA JAPAN & CEVA Biomune Campus, USA
3 1. What is a Vector Vaccine? 2. Why use Vector Vaccines? 3. Construction and development of Vector Vaccines 4. Active immunity induced by Vector Vaccines 3
4 Vector Vaccine Backgrounds Accumulation of knowledge in immunology and vaccinology Identification of protective antigens Advances in molecular biology Manipulation of bacteria, virus etc. Expression of extraneous genes in modified organisms Unique vaccines that may be safer and more efficacious than existing vaccines? Started research in 1980s 4
5 Vaccines Using Recombinant Technology DNA Vaccines naked plasmid Subunit Vaccines protective antigen protein; E.coli, baculovirus or yeast Live Gene-deleted Vaccines Live Vector Vaccines antigen Deleted gene 5 5
6 What is a Vector Vaccine? Virus or bacteria genetically modified to express exogenous antigen genes Two major components Donor Vector Exogenous antigen genes 6
7 What is a Vector Vaccine? Donor will provide a gene encoding protective antigens Fusion gene of Newcastle disease virus 7
8 What is a Vector Vaccine? Vector Attenuated virus or bacteria used for insertion of a Donor gene HVT FC126 DNA of HVT 8
9 Viral Vectors Large DNA viruses Li Licensed d poultry lt vectors t Herpesvirus Turkey herpesvirus (HVT) Laryngotracheitis virus Poxviruses fowlpox virus Adenoviruses RNA viruses reverse genetics g Newcastle disease virus Avian pneumovirus 9
10 What at is s a Vector ecto Vaccine? acc e Donor The organism g that contributes a gene g encoding protective antigens Vector Attenuated virus or bacteria used for i insertion ti off gene NDV fusion gene Donor Vector (parent)) (p Insertion site Vector Vaccine NDV fusion gene Produce key protective antigen protein = NDV HVT Vectormune HVT NDV 10
11 VECTORMUNE Line Vaccine Technology Making Life Simpler 1. VECTORMUNE FP LT + AE 2. VECTORMUNE FP LT 3. VECTORMUNE FP MG + AE 4. VECTORMUNE FP MG 5. VECTORMUNE FP N the first licensed vector vaccine in VECTORMUNE HVT NDV 7. VECTORMUNE HVT NDV + SB-1 8. VECTORMUNE HVT NDV + Rispens 9. VECTORMUNE HVT NDV + SB-1 + Rispens 10. VECTORMUNE HVT IBD 11. VECTORMUNE HVT IBD + SB VECTORMUNE HVT IBD + Rispens 13. VECTORMUNE HVT IBD + SB-1 + Rispens 14. VECTORMUNE HVT LT (US license obtained Sept 12, 2011) 15. VECTORMUNE HVT AI (US license obtained April 2, 2012) 11
12 1. What is a Vector Vaccine? 2. Why use Vector Vaccines? 3. Construction and development of Vector Vaccines 4. Active immunity induced by Vector Vaccines 12
13 Safety Why Use Vector Vaccines? Reduced vaccine reactions No reversion to virulence Administration Easy or mass administration (in ovo, SQ to day of age, etc.) May reduce or eliminate boosting Efficacy Evade maternal antibodies HVT vector Onset and duration of immunity Broader protection spectrum Multivalent vaccines 13
14 Why Use HVT Vector Vaccines? Improved safety Use at hatchery Evades maternal antibody Efficacy: 2 diseases in 1 vaccine Long duration of immunity 14
15 Safety Vectormune HVT vaccines HVT vaccine has been used safely since 1970s Vectormune HVT vaccines behave the same as HVT No vaccine reactions No reversion to virulence No transmission (horizontal or vertical) No safety concern! 15
16 Automation at Hatchery Vectormune HVT vaccines One time vaccination at hatchery (in ovo or SQ) Active replication in various tissues Evades maternal antibodies 16
17 Cell Associated Herpesvirus Marek s disease (HVT) Cell to cell spread of herpesvirus MDV in nucleus R.L. Witter and K.A. Schat D.C. Johnson and J.D. Baines 17
18 MDV specific maternal antibody interferes with early vaccination with cell free (freeze dried) MD vaccines does NOT interfere with early vaccination with cell associated (frozen) MD vaccines If have early, heavy MD challenge then use ONLY cell associated Same principle p for HVT vector vaccines Shipped and stored in liquid nitrogen Evades Maternal Antibody 18
19 Evading NDV Maternal Antibody Traditional Live NDV Vaccine VECTORMUNE HVT NDV NDV 19
20 HVT Vector Replicates Commercial layers challenged at 19 weeks of age after single vaccination with HVT NDV at day of age % ND protection 100% Control 20 SQ Vaccinated 0 0% (Rosenberger, 2000) 20
21 1. What is a Vector Vaccine? 2. Why use Vector Vaccines? 3. Construction and development of Vector Vaccines 4. Active immunity induced by Vector Vaccines 21
22 Most important characteristics of Vector Vaccines for protection Vector Vaccine Characteristics Vector insertion site affect virus replication Donor gene insert (& source) immunogenicity Promoter affect protein expression Promoter Control amount of antigen protein Antigen gene 22
23 Where is the NDV gene inserted? HVT genome Unique Long Unique Short CEVA TRL UL45/46 IRL IRS TRS Promoter NDV fusion gene Use of different insertion sites may affect virus replication, hence lower efficacy Ceva s insertion site 45/46 does not affect virus replication 23
24 How do we choose genes? Literature Published key protective antigens F and HN of NDV Research Unknown key protective antigens LT and MG Trial and Error - test Multiple genes Multiple promoter - gene combinations 24
25 For example Vectormune FP-LT 25
26 Genes on Vectormune FP-LT LTV genome U L IR S U S TR S gb gene UL-32 gene Group # Group LT challenge Protected/Total % Protected 1 rfpv/gb 6/12 50% 2 rfpv/ul-32 3/11 27% 3 rfpv/gb/ul-32 10/11 91% 4 Challenge Control 0/12 0% 26
27 Different HVT NDV combinations rhvt/p1-f rhvt/p2-f-1 rhvt/p2-f-2 rhvt/p3-f rhvt/p4-f-1 rhvt/p4-f-2 rhvt/p1fp2hn rhvt/p1fp3hn Selected for best Vector Vaccine Qualities 27
28 How Did We Construct Vectormune HVT NDV Plasmid Insertion site NDV F gene HVT Cell culture Homology plasmid HVT HVT DNA (Insertion site) 28
29 How Did We Construct Vectormune HVT NDV HVT Plasmid Antigen gene from pathogen HVT DNA Vector Homology plasmid Viral DNA (Insertion Recombinant site) HVT DNA Homology plasmid Cell Homologous recombination Cell culture HVT/NDV Selection NDV F gene 29
30 VECTORMUNE HVT NDV Expression of Antigen HVT UL45 Pec promoter NDV Fusion gene HVT UL kb 94kb kd 6.6 kb 4.4 kb Fgene 75 kd 2.3 kb 2.0 kb 50 kd 60 kd F protein Presence of F gene in Vectormune Southern Blot Expression of F protein by Vectormune Western Blot 30
31 1. What is a Vector Vaccine? 2. Why use Vector Vaccines? 3. Construction and development of Vector Vaccines 4. Active immunity induced by Vector Vaccines 31
32 Active Immune Response Humoral Observable ELISA titers (HVT NDV) S/P Value Age in Weeks Idexx NDV ELISA kit 32
33 Active Immune Response Humoral Observable HI titers (HVT AI) 10 I titers (lo og 2 ) 5 H 0 Mock-inoculated VTM HVT AI At 5 weeks old after SQ vaccination at day of age in SPF chickens Homologous AIV (H5 subtype) HA antigen was used 33
34 Active Immune Response Local - NDV specific antibody (IgG) in tears 34
35 Active Immune Response NDV specific cell mediate response 35
36 Vectormune HVT Vaccines NDV (Fusion) IBDV (Variant E VP2) Laryngotracheitis (gb) Avian Influenza (HA-H5) Vaccine Efficacious against Marek s disease Marek s disease ND Chimalhuacan, Herts 33, Texas GB, Thailand reference strain, Malaysian 2010 strain etc Classic and Variant IBD Marek s disease Infectious Laryngotracheitis Marek s disease Avian Influenza H5N1 Clade 1, H5N1 Clade 2.2 (Hungarian, Mongolian, and Egyptian variant ), H5N1 Clade 2.1 (Indonesia), H5N2 HPAI 36
37 Conclusions We started research in 1980s with hopes to develop vaccines with unique features using molecular biology We licensed the world s first vector vaccine in 1994 Since then, we have licensed 7 vector vaccines. We have learned a great deal about the advantages and limitations of vector vaccines. Vector vaccines offer various advantages over conventional vaccines including improved safety, easier vaccine administration, and improved efficacy 37
38 Conclusions Vectormune HVT NDV Fully safe No vaccine reactions Insertion of Fusion gene into the HVT genome in between 45/46 does not slow HVT replication Combination of our promoter and Fusion gene produce heavy expression of the Fusion protein, which leads to active immune response against NDV in vaccinated chickens. Protects against various NDVs (pathotypes and genotypes) from around the world (Chimalhuacan, Herts 33, Texas GB, Thailand reference strain, Malaysian 2010 strain etc.) Evades maternal antibodies against NDV and MDV 38
39 Research Group Dedicated for the Future of Animal Health 39
40 Thank You for Your Attention! 1. Boggan, G.D., A. Godoy, E. Brimer, K. Moore Dorsey, Y.Gardin. Comparison of Common Tools Following VECTORMUNE HVT IBD Vaccination. (Abstract 9373). AAAP/AVMA Scientific Program of the 147th Annual Meeting of American Veterinary Medical Association. July 31- August 3, Atlanta, GA. 2. Calnek, B.W. and Smith, M.W. (1972) Vaccination against Marek s Disease with cell-free turkey Herpesvirus: Interference by maternal antibody. Avian Diseases, 16, pp De Vriese, J., Y. Gardin, V. Palya, K. Moore Dorsey, B. Lambrecht, S. Van Borm, T. van den Berg. Efficacy of an rhvt-ai Vector Vaccine in Broilers with Passive Immunity to HVT, MDV and AIV, Against Challenge with H5N1 HPAIV. Poster Seventh International Symposium of Avian Influenza Athens, GA. 4. Esaki, M., K. M. Moore, T. Sato, S. Saitoh, M. Kubomura, A. Yasuda, and J. Leonard. Efficacy of Turkey Herpesvirus Vectored Infectious Bursal Disease Vaccine. (Abstract p 136). AAAP/AVMA Scientific Program of the 143rd Annual Meeting of American Veterinary Medical Association. July 15-19, Honolulu, HI. 5. Esaki, M., K. M. Moore, T. Sato, S. Saitoh, S. Saeki, A. Fujisawa, A. Yasuda, and J. Leonard. Construction and Evaluation of Turkey Herpesvirus Vectored Newcastle Disease Vaccine. (Abstract t p 156). AAAP/AVMA Scientific Program of the 143rd Annual Meeting of American Veterinary Medical Association. July 15-19, Honolulu, HI. 6. Esaki, M., T. Sato, L. Jensen, S. Saitoh, S. Saeki, A. Fujisawa, K. Moore Dorsey. Detection of VECTORMUNE HVT Vaccines in Feather Tips by Real-Time PCR. (Abstract 7657). AAAP/AVMA Scientific Program of the 146rd Annual Meeting of American Veterinary Medical Association. July 11-15, Seattle, WA. 7. Jensen, L., M. Esaki, P. Flegg, K. Moore Dorsey, S. Rosenberger and J.K. Rosenberger. Efficacy Studies of a HVT Vector Expressing Laryngotracheitis Virus Genes. (Abstract p 80). AAAP/AVMA Scientific Program of the 145th Annual Meeting of American Veterinary Medical Association. July 19-23, New Orleans, LA. 8. Kapczynski, D.R., M. Esaki, M.W. Jackwood, and K. Moore Dorsey. Vaccination of SPF Chickens with a Recombinant HVT Expressing the HA from H5N1 Highly Pathogenic Avian Influenza Protects Against Lethal Challenge. 59th Western Poultry Disease Conference Vancouver, BC, Canada. 9. Godoy, A., M. Esaki, K. Varga, P. Flegg, K. Moore Dorsey, S. Rosenberger and J.K. Rosenberger. Compatibility of Turkey Herpesvirus Recombinant Vaccines. (Abstract p 76). AAAP/AVMA Scientific Program of the 145th Annual Meeting of American Veterinary Medical Association. July 19-23, New Orleans, LA. 10. Godoy, A., P. Flegg, I. Oldoni, M. García, and K. Moore Dorsey. Efficacy of a Recombinant Fowl Pox Vectored Laryngotracheitis Vaccine and Sequence Comparison to Recent Field Isolates. (Abstract p 108). AAAP/AVMA Scientific Program of the 145th Annual Meeting of American Veterinary Medical Association. July 19-23, New Orleans, LA. 11. Godoy, A., M. Garcia, K. Moore, M. Esaki, and J. Leonard. Mycoplasma gallisepticum Detection and Genotyping in Vaccinated Layer Flocks. (Abstract p 150). AAAP/AVMA Scientific Program of the 143rd Annual Meeting of American Veterinary Medical Association. July 15-19, Honolulu, HI. 12. Moore, K.M., Y. Gardin and J.D. Leonard. Avian Influenza Vaccines. IABS International Scientific Workshop New Cells for New Vaccines II: Focus on Respiratory Virus Diseases. 2007, Wilmington, DE. 13. Moore, K.M., J. R. Davis, Y. Tsuzaki, D. R. Hout, M. Esaki, T. Okuda, and J. D. Leonard. Update: Efficacy and safety of a recombinant fowl poxvirus containing laryngotracheitis genes. (Abstract p 21). Proceeding of the 51 st Western Poultry Disease Conference. May 1-4, Puerto Vallarta, Mexico. 14. Moore, K.M., J.R. Davis, Y. Tsuzaki, D.R. Hout, M. Esaki, T. Okuda, and J.D. Leonard. Efficacy and safety of a recombinant fowl poxvirus containing laryngotracheitis genes. (Abstract p 40). AAAP/AVMA Scientific Program of the 138 th Annual Meeting of American Veterinary Medical Association. July 14-18, Boston, MA. 15. Palya, V., Z. Penzes, T. Horvath, V. Kardi, K. Moore Dorsey, Y. Gardin. Comparative efficacy of several vaccination programmes including or not recombinant HVT-ND vaccine against challenge with Mexican Chimahuacan NDV strain th Western Poultry Disease Conference & Xxxiii Annual Aneca Convention. 16. Tsuzaki, Y., J. R. Davis,, D. R. Hout, S. Saitoh, M. Esaki, T. Okuda, A. Fujisawa, K. M. Moore, and J. D. Leonard. Analysis ofmycoplasma gallisepticum genes expressed by a fowl poxvirus vector (Abstract p 40). AAAP/AVMA Scientific Program of the 138 th Annual Meeting of American Veterinary Medical Association. July 14-18, Boston, MA. 40
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