WO1995012682A2 - Vecteur viral a antigenes du virus de la maladie des muqueuses (bvdv) - Google Patents

Vecteur viral a antigenes du virus de la maladie des muqueuses (bvdv) Download PDF

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WO1995012682A2
WO1995012682A2 PCT/US1994/012198 US9412198W WO9512682A2 WO 1995012682 A2 WO1995012682 A2 WO 1995012682A2 US 9412198 W US9412198 W US 9412198W WO 9512682 A2 WO9512682 A2 WO 9512682A2
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virus
gene
plasmid
thymidine kinase
combination
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PCT/US1994/012198
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WO1995012682A3 (fr
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Elizabeth J. Haanes
Richard C. Wardley
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The Upjohn Company
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Priority to AU10423/95A priority Critical patent/AU688819B2/en
Priority to KR1019960702343A priority patent/KR960705944A/ko
Priority to JP7513263A priority patent/JPH09504435A/ja
Priority to NZ276234A priority patent/NZ276234A/en
Priority to EP95901037A priority patent/EP0725831A1/fr
Publication of WO1995012682A2 publication Critical patent/WO1995012682A2/fr
Publication of WO1995012682A3 publication Critical patent/WO1995012682A3/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
    • C12N15/86—Viral vectors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
    • C12N2710/00011—Details
    • C12N2710/16011—Herpesviridae
    • C12N2710/16611—Simplexvirus, e.g. human herpesvirus 1, 2
    • C12N2710/16641—Use of virus, viral particle or viral elements as a vector
    • C12N2710/16643—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
    • C12N2770/00011—Details
    • C12N2770/24011—Flaviviridae
    • C12N2770/24311—Pestivirus, e.g. bovine viral diarrhea virus
    • C12N2770/24322—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes

Definitions

  • This invention relates to the field of Bovine Viral Diarrhea Virus (BVDV), and vaccines for the treatment thereof.
  • Bovine viral diarrhea virus is a Pestivirus belonging to the family of the Flaviviridae. It causes a number of different conditions in sheep, goats, and especially cattle. The symptoms depend upon the age, physiological and virological state of the animal. In young susceptible calves and young adults it causes a disease which is characterized by high morbidity and low mortality. The symptoms can include fever, depression, occulo-nasal discharges, diarrhea and occasionally oral ulcerations. Apart from these primary effects the virus also causes immunosuppression. Although primary BVDV infections are normally relatively mild, the virus may potentiate or enhance the pathogenicity of other co-infecting microorganisms.
  • BVDV causes similar symptoms to those described above for younger susceptible calves.
  • the virus in pregnant a * n * i ⁇ nalp the virus has the ability to cross the placenta and infect the fetus. The outcome of this infection depends upon the age of the fetus and whether it is at a stage where its immune system is fully competent.
  • the possible outcome of infections include fetal reabsorption, abortion, mummification, congenital defects, birth defects, calves born which are persistently infected with BVDV and completely normal calves. Calves born which are persistently infected with BVDV, represent the most important segment of this BVDV pathogenesis complex.
  • Persistently infected animals shed large amounts of virus into their environment which can infect susceptible animals. Furthermore, even though persistently infected animals are immunotolerant to the virus which infected them in utero, they do develop disease when infected with other closely related BVDV biotypes. These infections are characterized by low morbidity (because relatively speaking there will not be many pregnant animals infected at the right time during pregnancy to produce BVDV persistently infected normal calves), but high mortality. This disease syndrome is known as mucosal disease and often manifests itself as a peracute condition with calves dying of a profuse watery diarrhea which contains large amounts of fresh blood.
  • inactivated vaccine preparations are not as effective as attenuated live vaccines.
  • Inactivated antigen from inactivated vaccine preparation undergoes exogenous processing. After injection into the animal the antigen becomes part of the animal's soluble protein mileau. The antigen enters antigen presenting cells through pinocytotic mechanisms and this usually produces antibodies.
  • Antigen from live virus which replicates inside cells undergoes endogenous processing and this mechanism produces the preferred cell mediated immune responses.
  • Cell mediated immune responses can recognize cells infected with viruses and have the potential of interrupting the virus life cycle at a much earlier stage. Cell mediated responses are thus thought to be extremely important in the immunological defense to many viral infections.
  • An animal owner may be vaccinating animals to protect against a disease but because of the properties of the vaccine the owner provides an opportunity for other diseases to afflict the animals. This forces the owner to use inactivated BVDV vaccines, which because of the way in which the immune system operates, are not particularly effective.
  • inactivated vaccines are safe but not particularly effective while the attenuated live vaccines are more effective but under certain conditions may not be very safe.
  • Bovine herpesvirus type l (BHV-l) is another major pathogen of cattle which produces respiratory disease.
  • BHV-1 also replicates at a mucosal surface.
  • gp53 a major glycoprotein of the BVDV virus and against which the host produces substantial immune responses
  • BHV/BVDVgp53 this recombinant virus
  • a replicating nonpathogenic virus for preventing disease caused by Bovine Viral Diarrhea Virus (BVDV), where said replicating nonpathogenic virus comprises: a gene or gene combination taken from a BVDV virus, and said replicating nonpathogenic virus functionally expresses said gene or gene combination.
  • BVDV Bovine Viral Diarrhea Virus
  • Embodiments of this invention include the following: A virus where said replicating nonpathogenic virus is attenuated, is selected from attenuated Bovine Herpes Virus type 1 (BHV-1), attenuated adenoviruses, attenuated bovine mammillitis virus, attenuated bovine papillomavirus, or attenuated pseudorabies virus.
  • a virus where the attenuation is created by making the thymidine kinase (tk) gene nonfunctional.
  • a virus where said gene that codes for gp53 is inserted into the inactivated thymidine kinase (tk) gene site.
  • a virus where the functionally expressing gene or gene combination, used to create the virus comprises a recombined plasmid with intact viral DNA, said plasmid comprising: a) a BHV-1 genomic DNA fragment containing the thymidine kinase (tk) gene and having a deletion to the thymidine kinase (tk) gene, b) a promoter/polyadenylation signal inserted in the thymidine kinase (tk) gene deletion, c) a signal peptide gene sequence preceding a gp53 gene or gene combination all of which is inserted between the promoter and the polyadenylation signal.
  • a virus where said plasmid is made from a plasmid having the characteristics of plasmid pHAS4.
  • a virus where said signal peptide gene sequence is taken from any well characterized signal peptide sequences such as any of the thirty-nine examples of well characterized signal peptide sequences found in Perlman, D., et al., J. Mol . Biol. Vol. 167 pp. 391-409 (1983), incorporated by reference.
  • a virus where said signal peptide gene sequence is taken from Psuedorabies Virus gill gene (PRV) and/or Bovine Growth Hormone (BGH).
  • PRV Psuedorabies Virus gill gene
  • BGH Bovine Growth Hormone
  • a virus where a plasmid is selected from the following plasmids, a) pBHVtkex-l::BGH p53; b) pBHVtkex-l::gHI/p53; c) pBHVtkex-3::BGH/p53; or d) pBHVtkex-3::gIII/p53.
  • a virus that produces the product of a functionally expressing gene or gene combination is selected from one of the following viruses, Tll-3, Tll-6, or Tll-8.
  • a virus where the functionally expressing gene or gene combination, used to create the virus comprises a recombined plasmid with intact viral DNA said plasmid comprising: a) a BHV-1 genomic DNA fragment containing the thymidine kinase (tk) gene and having a deletion to the thymidine kinase (tk) gene, b) a promoter/polyadenylation signal inserted in the thymidine kinase (tk) gene deletion, c) a gp53 gene or gene combination inserted between the promoter and the polyadenylation signal.
  • a virus where the plasmid is pBHVtkex-3::p53.
  • a vaccine for preventing disease caused by Bovine Viral Diarrhea Virus (BDVD) comprising a pharmaceutically effective amount of the viruses described herein and a carrier.
  • a vaccine as described above for preventing disease caused by Bovine Viral Diarrhea Virus (BDVD) comprising a pharmaceutically effective amount of a virus described above and a carrier, said carrier comprising any physiological buffered medium, i.e. about pH 7.0 to 7.4 containing from about 2.5 to 15% serum which does not contain antibodies to BHV.
  • Bovine Viral Diarrhea Virus comprising administering to an animal a pharmaceutically effective amount of a virus or vaccine described herein.
  • a process of preparing a virus described herein comprising: a) isolation of a functionally expressing gene or gene combination that causes BVDV, b) inserting said gene or gene combination into a replicating nonpathogenic virus, c) selecting a live-virus that functionally expresses the product of said gene or gene combination.
  • a method of preparing a virus described herein where the functionally expressing gene or gene combination, used to create the virus, is produced by a process comprising the recombination of a plasmid with intact viral DNA said plasmid comprising: a) a BHV-1 genomic DNA fragment containing the thymidine kinase (tk) gene and having a deletion to the thymidine kinase (tk) gene, b) inserting into the thymidine kinase (tk) gene deletion of said plasmid a promoter/polyadenylation signal, c) inserting a gp53 gene or gene combination between the promoter and the polyadenylation signal, d) transfecting cells with said plasmid to produce a recombinate virus containing said functional gene or gene combination inserted into a live virus that does not cause immunosuppression in the usual host and expressing said functional gene or gene combination.
  • a method of preparing a virus described herein where the functionally expressing gene or gene combination, used to create the virus, is produced by a process comprising the recombination of a plasmid with intact viral DNA said plasmid comprising: a) a BHV-1 genomic DNA fragment containing the thymidine kinase (tk) gene and having a deletion to the thymidine kinase (tk) gene, b) inserting into the thymidine kinase (tk) gene deletion of said plasmid a promoter/polyadenylation signal, c) inserting a gp53 gene or gene combination preceded by a signal peptide gene sequence between the promoter and the polyadenylation signal, d) transfecting cells with said plasmid to produce a recombinate virus containing said functional gene or gene combination inserted into a live virus that does not cause immunosuppression in the usual host and expressing said functional gene or gene combination.
  • FIG. 1 Construction of the shuttle vectors for inserting foreign genes into BHV-1.
  • Figure 2. Strategy for appending signal peptide sequences to the BVDV gp53 gene.
  • Figure 3. Maps of the five shuttle plasmids for inserting gp53 into BHV-1 a.
  • EXAMPLE 1. pBHVtkex-3::p53.
  • b. EXAMPLE 2: pBHVtkex-l::BGH/p53 c.
  • EXAMPLE 3 pBHVtkex-l::gIII/p53 d.
  • EXAMPLE 4 pBHVtkex-3::BGH/p53 e.
  • Figure 4. Predicted transcript maps of the BHV-l/gp53 recombinant viruses.
  • Figure 5. Northern blots showing transcription of gp53 messenger RNAs in the BHV-1 recombin
  • This invention combines the effectiveness of the attenuated live vaccines with the safety of the inactivated vaccines.
  • BHV bovine herpesvirus -1
  • Bovine herpesvirus is another major pathogen of cattle which produces respiratoiy disease.
  • BHV also replicates at a mucosal surface.
  • replication is mainly at the gut mucosal interface with less replication at the respiratory interface.
  • BHV it is the respiratory interface which dominates.
  • the common mucosal immune system ensures that immune responses produced at one surface will be effective at other surfaces.
  • the recombinant virus of this invention, BHV/BVDVgp53 will, when administered to cows, sheep or goats, preferably via the intranasal route, replicate in the respiratory mucosae and produce an immune response.
  • the thymidine kinase gene was deleted from the BHV virus using a process known to attenuate the virus.
  • the BHV a live attenuated virus, will replicate and produce a cell mediated response.
  • the BVDV gp53 gene will be expressed and, because the virus is inside the cell, the correct processing for a cell mediated response to the BVDV gp53 part of the recombinant virus will also occur. Most importantly, this response will occur without the possible side effects of immunosuppression, as only part of the BVDV virus is present.
  • the invention combines the efficacy of an attenuated live virus vaccine for BVDV, with the safety of an inactivated preparation.
  • compositions and Administrations - A pharmaceutically effective amount of the vaccine of the present invention can be employed along with a pharmaceutically acceptable carrier or diluent as a vaccine against BHV-1 and BVDV in animals, such as bovine, sheep and goats.
  • Examples of pharmaceutically acceptable carriers or diluents useful in the present invention include any physiological buffered medium, i.e., about pH 7.0 to 7.4, containing from about 2.5 to 15% serum which does not contain antibodies to BHV, i.e., is seronegative for BHV. Serum which does not contain gamma globulin is preferred to serum which contains gamma globulin.
  • Examples of serum to be employed in the present invention include fetal calf serum, lamb serum, horse serum, swine serum, and goat serum. Serum protein such as porcine albumin or bovine serum albumin (hereinafter "BSA") in an amount of from about 0.5 to 3.0% can be employed as a substitute for the serum.
  • BSA bovine serum albumin
  • the virus may be diluted in any of the conventional stabilizing solutions containing phosphate buffer, glutamate, casitone, and sucrose or sorbose, or containing phosphate buffer, lactose, dextran and glutamate. It is preferred that the vaccine viruses of the present invention be stored at a titer of at least IO 5 to IO 6 PFU/ml at -70°C to -90°C or in a lyophilized state at 2°C to 7°C.
  • the lyophilized virus may be reconstituted for use with sterile distilled water or using an aqueous diluent containing preservatives such as gentamicin and amphotericin B or penicillin and streptomycin.
  • the useful dosage to be administered will vary depending upon the age, weight and species of the animal vaccinated and the mode of administration.
  • a suitable dosage can be, for example, about IO 4,5 to IO 7 PFU/animal, preferably about IO 4 - 5 to IO 5 5 PFU.
  • the vaccines of the present invention can be administered intranasally, intravaginally or intramuscularly. Intranasally is the preferred mode of administration.
  • Utility of the Invention - This invention is intended to provide the user with an effective vaccine for prevention of BVDV caused disease, where the vaccine can be safely and efficaciously administered intramuscularly, intranasally, or intravaginally. Intranasally may be the preferred route of administration.
  • the vaccines of this invention are created with the intention of treating disease, preferably through prevention.
  • prevent or prevention applicant means to keep the host from developing symptoms of the disease or to mitigate the effects of the disease, that is to avert the typical diseased state.
  • Prevention implies decisive action to stop, impede or delay the onset of disease.
  • Prevention can include the following concepts: to hinder, frustrate, to obstruct; to intercept, possibly prohibit, impede or preclude.
  • Preclude would suggest the onset of the disease state either does not occur or the disease pathogen is largely ineffectual in causing the disease state.
  • Prevent or prevention can indicate the disease state is forstalled, meaning that anticipatory action to prevent or hinder the disease has occurred but the conditions creating the disease have not been eliminated.
  • Another object of the invention is to provide a BVDV vaccine which can be administered safely to calves and to pregnant cows in all stages of pregnancy.
  • the vaccine uses gp53, a major glycoprotein of BVDV, and one against which the host should produces a substantial immune response.
  • gp53 is highly immunogenic.
  • Donis, R.O. and Dubovi, E.J. Glycoproteins of Bovine Viral Diarrhoea-Mucosal Disease Virus in Infected Bovine Cells, Journal of General Virology, Vol. 68, pp. 1607-1616 (1987). It is well known that agents that produce substantial immune responses can make effective vaccines.
  • Magar, R., et al. Bovine Viral Diarrhea Virus Proteins: Heterogeneity of
  • the vaccines of this invention contain genes that express large quantities of gp53, this is shown in figure 5. Because of the expression of large quantities of gp53 the vaccines of this invention will confer substantial benefits upon the treated potential host.
  • Preferred Compounds - Any BHV-1 virus attenuated with a tk deletion and carrying the gp53 gene, the gp53 gene being preceded by a signal peptide, that expresses abundant amounts of gp53, should be a preferred suitable vaccine candidate. It appears the signal peptide sequence may be taken from any suitable source. We chose to examine two different signal peptides to ensure the best localization of the gp53 protein in vivo. We chose two candidates we call “Tll-6", embodied in Example 2, and “Tll-3", embodied in Example 3 for vaccine trials. The former virus was deposited to the ATCC under the designation UC VR-58. The latter, "Tll-3" plasmid was also deposited.
  • Tll-8 The virus we labeled "Tll-8" might contain truncated forms of the tk transcript and this might suggest, but does not necessarly mean, that it would be less attractive as a vaccine candidate.
  • a large number of existing cell lines are persistently infected with non-cytopathic BVDV from passage in media containing fetal bovine serum taken from infected calves. For this invention, it is imperative that viruses used as live, attenuated vaccines are free of contaminating BVDV.
  • BHV-1 a vector for BVDV genes
  • Other examples from cattle, sheep and goats would include cow, goat and sheep pox viruses, adenoviruses, bovine mammillitis virus, bovine papillomavirus, and pseudorabies virus.
  • a non ⁇ pathogenic virus refers to any virus which has the ability to replicate in one of its host species but does not produce any signs of disease in that species.
  • non ⁇ pathogenic viruses might arise from pathogenic parent viruses by natural mutation, might be mutagenized by, for instance, chemicals or light to produce a non ⁇ pathogenic virus, or could be rendered non-pathogenic through the use of recombinant DNA technologies. See, 1) Mapping Neutralization Domains of Viruses, E.Wimmer, E.A. Emini, and D.C. Diamondand 2) Immunogenicity of Vaccine Products and Neutralizing Antibodies, E Norrby. Both articles are in Edited by Notkins and Oldstone Published by Springer-Verlag New York Inc. 1986.
  • a 424bp deletion was introduced into the tk gene by digesting pHAS4 with Bglll and Xhol, filling in the ends with the Klenow Fragment of DNA polymerase I (Klenow) and religating the resulting blunt ended fragments. This manipulation restored the Bglll recognition site, but not the Xhol site (Fig. 1). The resulting plasmid was named pHAS4 ⁇ BX. This deletion was chosen because it does not impede on the previously identified transcription initiation sites for the UL24 homolog which overlaps the 5' end of the tk gene. L. J. Bello, et al., Virology, 189:407-414 (1992); J. G.
  • CMV Human cytomegalovirus
  • bovine growth hormone polyadenylation sequence R. J. Brideau, et al., J Gen Virol, 74:471-477 (1993 ).
  • CMV Human cytomegalovirus
  • the cassette, in vector p3CL-DHFR, is bounded by unique EcoRI and Bglll sites and contains, between the promoter and the polyadenylation signal, unique Hindlll and Sail restriction sites for cloning of foreign genes.
  • the p3CL-DHFR vector was digested with EcoRI, then filled in and ligated to a BamHI linker (New England Biolabs, Beverly, Massachusetts). This manipulation regenerated the EcoRI site.
  • the construct was then digested with BamHI and Bglll and the released cassette was ligated into the Bglll site of pHAS4 ⁇ BX (Fig. 1).
  • the ligations were transformed into E. coli strain DH5 ⁇ .
  • pHAS4 ⁇ BXex-l and pHAS4 ⁇ BXex-3 Fig. 1
  • FIG. 1 Construction of shuttle vectors for inserting foreign genes into BHV-1.
  • PHAS4 is a 2.7kb subfragment from the BHV-1 Hindm-A fragment.
  • the BglH/XhoI subfragment to be deleted is shown.
  • the deletion derivative of pHAS4 is pHAS4 ⁇ BX.
  • the deleted thymidine kinase (tk) gene is shown as a dark stippled box.
  • the cassette containing the promoter and polyadenylation signal is shown just below pHAS4 ⁇ BX.
  • the CMV immediate early promoter is shown as a light stippled box, and the Bovine Growth Hormone (BGH) polyadenylation signal is shown as a striped box.
  • BGH Bovine Growth Hormone
  • Complementary oligonucleotides encoding any well characterized signal peptide can be used in this invention. Thirty-nine examples of well characterized signal peptide sequences found in Perlman, D., et al., J. Mol . Biol. Vol. 167 pp. 391-409 (1983). Incorporated by reference. These and any other well characterized signal peptides should be suitable for use as embodiments of this invention.
  • Section A Synthetic oligonucleotides corresponding to the signal peptide sequences of Bovine Growth Hormone (BGH), and Pseudorabies virus gill (PRV gill). Complementary oligonucleotides were synthesized such that the annealed pairs had Sallsites on the 5' ends and BamHI sites on the 3' ends. The deduced amino acid sequences of the signal peptides are also shown. In each case the predicted cleavage sites for the signal peptides are just after the alanine (A), three amino acids from the ends. Codons for two amino acid residues (F,P in BGH; P,S in gill) from the original native proteins were left on the signal peptide sequences to ensure correct cleavage.
  • BGH Bovine Growth Hormone
  • PRV gill Pseudorabies virus gill
  • Section B Site directed Mutagenesis of the cDNA encoding the BVDV gp53 gene.
  • the first 60 nucleotides of the gp53 cDNA and the corresponding amino acid sequence are shown.
  • a single base pair, shown by the arrow, was changed to create a BamHI restriction site in the sequence, shown in the box. This change does not change the amino acid sequence.
  • the cDNA was then digested with BamHI as shown, allowing in frame ligation to either of the signal peptide sequences shown in section A
  • BVDV gene fragments in addition to gp53.
  • Expression of other BVDV gene or gene combinations in a live virus vector are also embodiments of this invention. This would include any and all BVDV proteins to which a vaccinated animal could elicit an immune response. Examples include, but are not limited to, the other two BVDV surface glycoproteins, gp48 and gp25 (Collett, M.S., et al., Virology 165:200-208 (1988)), the pl4 capsid protein (Thiel, H.J., et al., J. Virol. 65:4705-4712 (1991)), and the p20 N-te ⁇ ninal protease.
  • BVDV protein candidate to express in a vaccine is the nonstructural pl25/p80 protein (Deregt, D., et al., Can. J. Microbiol. 37:815-122 (1991)), which elicits a significant antibody response in infected cows.
  • Insertion of the BVDV gp53 gene into the BHV-1 expression vectors The p53 gene, either with or without added signal peptide sequences, was ligated into the Hindlll insertion sites of pHAS4 ⁇ BXex-l and pHAS4 ⁇ BXex-3 by filling in all the respective cohesive ends of vectors and inserts followed by blunt end ligation. The ligations were transformed in E. coli strain DH5 ⁇ . We wanted to eventually evaluate the expression of gp53 in BHV-1 in various orientations and with at least two different signal peptides to ensure that we achieved the most efficient expression. The transformed colonies were screened by colony hybridization using as a probe the p53 insert labelled with Digoxygenin-dUTP.
  • EXAMPLE 1 pBHVtkex-3::p53: contains the BVDV gp53 gene inserted between the CMV promoter and the BGH polyadenylation signal of pHAS4 ⁇ BXex-3 with no added signal peptide. In this construct the original gp53 gene, PRIOR to site directed mutagenesis, was inserted. See Fig. 3A This plasmid was then used to construct the virus T2-3#.
  • pBHVtkex-l::BGH/p53 contains the mutagenized gp53 gene preceded by the BGH signal peptide sequence inserted into pHAS4 ⁇ BXex-
  • pBHVtkex-l::gIII/p53 contains the mutagenized gp53 gene preceded by the PRV gill signal peptide sequence inserted into pHAS4 ⁇ BXex-l. See Fig. 3C. This plasmid was used the create the virus
  • the gp53 gene is shown as a solid band, the BHV-1 sequences are shown as dark stippled bands, the CMV promoter region is shown as a light stippled band, and the BGH polyadenylation signal region is shown as a striped band.
  • the plasmid vector, pUCl ⁇ is shown as a thin line. In each case the direction of transcription of gp53 relative to the original direction of transcription of BHV-1 tk is shown. The various signal peptide sequences are indicated.
  • pBHVtkex-l::BGH/p53 c.
  • BVDV gp53 gene into the BHV- 1 tk gene are embodiments of this invention.
  • plasmids and any plasmids created in this manner are known as "Principal Plasmid Vectors" and are the plasmid vectors used to create the virus vaccines of this invention.
  • the five expression shuttle plasmids carrying gp53 were linearized by Xbal and cotransfected into Bovine Turbinate (BT) cells with unit length DNA from BHV- 1 strain Iowa (tk positive) by the standard CaPO 4 method (R. L. Graham, et al., Virology, 52:456-467 (1973 )) as modified by Cai (W. Cai, et al., J Virol, 61:714-721 (1987 )) .
  • the cells were obtained from ATCC.
  • the transfections were then subjected to two rounds of selection either on 143tk " cells (S. K Mittal, et al., J Gen Virol, 70:(1989 )), or on Rab (BU) cells (S.
  • BDUR lOOug ml 5-Bromo-2'-Deoxyuridine
  • Other tk " cell lines permissive for growth of BHV-1 can also be used.
  • transfections that still showed cytopathic effect were infected onto BT cells under complete media with 1% low melting agarose to obtain single plaques.
  • Tll-3 BHV-1 "Iowa” into which the insert sequences contained in pBHVtkex-l::gIII/p53 recombined.
  • EXAMPLE 5 Tll-8: BHV-1 "Iowa” into which the insert sequences contained in pBHVtkex-3::gIII/p53 recombined.
  • BVDV gp53 Contains the BVDV gp53 gene with the PRV gill signal peptide sequence situated between the CMV promoter and the BGH polyadenlyation signal, with transcriptional orientation in the same direction as the BHV-1 tk gene.
  • a virus was not isolated from cotransfections with "Iowa” DNA and plasmid pBHVtkex3::BGH/p53, EXAMPLE 4, but this prophetic virus, could be easily created, it and any other BHV-1 viruses containing the BVDVgp53 gene inserted into thymidine kinase gene are embodiments of this invention.
  • viruses were plaque purified twice by limiting dilution on BT cells.
  • a large number of existing cell lines are persistently infected with non- cytopathic BVDV from passage in media containing fetal bovine serum taken from infected calves.
  • viruses used as live, attenuated vaccines are free of contaminating BVDV.
  • we prepared DNA from each of the viruses including the parent strain Iowa and Iowa ⁇ BX) and subjected the DNA preps to extensive RNAse treatment using a cloned RNAse (RNAse ONE, Promega Corporation, Madison, Wisconsin).
  • BVDV has only RNA as its genetic material
  • this manipulation should eliminate any possible contaminating BVDV sequences from the viral DNA preps.
  • ATCC certified BVD-free MDBK cells
  • RNA from each of the recombinant viruses and the parent BHV- 1 strain Iowa was prepared and evaluated transcription of gp53 by Northern hybridization. A diagram of the possible message species and the probes used is shown in Fig. 4.
  • FIG. 5 Predicted transcripts of the BHV-l/gp53 recombinant viruses later shown in Figure. 5.
  • the two probes are 1) the gp53 cDNA and 2) the Sall/Bglll portion of pHAS4 (shown above the maps).
  • the first map shows the predicted transcripts from viruses Tll-3 and Tll-6, and the second map shows the predicted transcripts from Tll-8.
  • the sites of transcript initiation for tk and UL24 are shown for reference.
  • All of the gp53 recombinant viruses made a 1.6kb message that hybridized with a P-labelled gp53 probe, the size predicted for transcription initiation at the CMV promoter and termination at the BgH polyadenylation site , Fig. 5, probe 1.
  • the T2-3#3 and T2-2#5 virus are not shown.
  • Tll-3 and Tll-6 made an 8.5kb transcript and Tll-8 and T2-3#3 made a 5.6kb transcript. These transcripts were unique to the recombinant viruses, and were consistent with messages initiating at the CMV promoter, reading through the BgH poly adenylation signal and terminating at the UL24 or tk/gH polyadenylation signals, respectively.
  • Hybridization with the upstream and downstream probes confirmed the identity of these longer messages.
  • the p53 probe did not hybridize to Iowa, Ia ⁇ BX or mock infected RNAs.
  • pHAS6 an 867bp sail fragment that maps downstream of the tk open reading frame and is internal to the gH gene.
  • All of the viruses made equivalent amounts of the 3.1kb gH message (data not shown).
  • This probe also hybridized to the longer p53 messages in Tll-8 and T2-3-3, and to the 4.3kb tk message in Iowa, which is 3' coterminal with the gH transcript. L. J.
  • FIG. 5 Northern blots showing transcription of gp53 messenger RNAs in the BHV-1 recombinant viruses.
  • the first panel shows transcripts hybridizing to probe 1, the pg53 cDNA, and the second panel shows transcripts hybridizing to probe 2, the Sall/Bgll subfragment of pHAS4.
  • RNA size standards, in kilobases (kB) are given to the left of each panel. Expression of BVDV gp53 protein in BHV-1.
  • IP immunoprecipitation
  • Immunoprecipitin Gibco BRL, Gaithersburg, Maryland, or protein A sepharose 4B (Pharmacia, Uppsala, Sweden). Immunoreactive proteins were resolved by SDS- Polyacrylamide gel electrophoresis (SDS-PAGE) and fluorography.
  • Figure 6 shows that all three of the recombinant viruses carrying the gp53 gene preceded by a signal peptide sequence made significant amounts of the protein.
  • the clones t2-3#3 and T2-2#5 are independently isolated clones, which rules out the possibility that one particular virus had a defect that precluded gp53 expression (data not shown).
  • the possiblity remains that gp53 is being synthesized from T2-3, but is rapidly degraded, or that our antibody does not detect unprocessed forms of the protein.
  • the slight size difference between the recombinant and native proteins could be due to the fact that the gp53 gene in the BHV-1 viruses came from a different BVD strain which could have a gp53 of a slightly different size, or the cDNA gp53 clone might not contain the exact amino acids processed from the BVDV polyprotein into native gp53.
  • a virus was deposited with the American Type Culture Collection, 12301 Parklawn Drive, Rockville, Maryland, zip code 20852, USA. That deposit was designated UC VR-58 by the Upjohn Company and given the following number by the depository, ATCC No. VR2436, it corresponds to the virus described herein as "Tll-6,” also known as "Example 2.” This deposit was received by the American Type Culture Collection depository on 28 October 1993. Several plasmids were deposited with the Agricultural Research Service
  • coli culture UC 15086 referring to pBHVtkex-3::gIII ⁇ p53, it corresponds to the plasmid used to create the virus described herein as, "T-ll-8," also known as “Example 5.”
  • This plasmid was given the following number by the depository, NRRL B-21351. Both of the plasmids were received by the Agricultural Research Service Culture Collection depository on 26 October 1994.
  • NAME Wootton, Thomas A.
  • MOLECULE TYPE DNA (genomic)
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • ORIGINAL SOURCE
  • ATATCGTTTA CGGGGGATGG CGATAGACGC CTTTGGTGAC TTGGGCGATT CTGTGTCG 2160 CAAATATCGC AGTTTCGATA TAGGTGACAG ACGATATGAG GCTATATCGC CGATAGAGGC 2220
  • GCCAGCCCCG ACACCCGCCA ACACCCGCTG ACGCGCCCTG ACGGGCTTGT CTGCTCCCGG 6000 CATCCGCTTA CAGACAAGCT GTGACCGTCT CCGGGAGCTG CATGTGTCAG AGGTTTTCAC 6060
  • MOLECULE TYPE DNA (genomic)
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • ORIGINAL SOURCE
  • GAAAGACAAC AACGGGCCGC GCGCTCGCGG CCGCTTCCAC CGCTGGGGAG GGCGTGCTCT 1500 TTTTCCCGGA GCCGATGGCG TACTGGCGCA CGATGTTTGG TACGGACGCC TTAAGTGGGA 1560
  • ACCAGAGCAC GTATCTTCCA CCCAGTAAAG CTACCACCAC CACCAATATG GACTCGGCGA 2700 AGTAATCCCG ATGATGGTCA GTGACCTCCA GGTCGAACCA GTATTGATAA TCTCCTTTTA 2760
  • GTGTTGTATT GAACTTCCCT CTTACTACGG GATTGGCATC GCATGGGCAG AGTCCAAATT 3420 CAAAGTCGTC AGCCATTTCA AATGTTTCCC CTTGCCTTTG CCCCTCGAAA AGTTTTTTGA 3480
  • ATTTCCGTGT CGCCCTTATT CCCTTTTTTG CGGCATTTTG CCTTCCTGTT TTTGCTCACC 6420
  • MOLECULE TYPE DNA (genomic)
  • CTGAGCTTGC ATGCCTGAGG TCGACCCTGG ATAAGCTGAT CCTCAATCAA TCAAGGTGGT 2460
  • TGCCGTCTAC AAATCTGTAG CCAGTCTCAT TCTTCAACCT ACACTTGCCG ATGGGGTAGT 3060
  • TGATGCTCGT CAGGGGGGCG GAGCCTATGG AAAAACGCC GCAACGCGGC CTTTTTACGG 7980
  • MOLECULE TYPE DNA (genomic)
  • HYPOTHETICAL NO
  • ATATCGTTTA CGGGGGATGG CGATAGACGC CTTTGGTGAC TTGGGCGATT CTGTGTCG 2160 CAAATATCGC AGTTTCGATA TAGGTGACAG ACGATATGAG GCTATATCGC CGATAGAGGC 2220
  • TGACACCAAA CTTGGGCCTA TGCCTTGCAA GCCATATGAG ATCATACCAA GTGAGGGGCC 4080 TGTAGAAAAG ACGGCATGCA CCTTCAACTA CACGAGGACA TTAAAAAATA AATATTTTGA 4140
  • ATTTCCGTGT CGCCCTTATT CCCTTTTTTG CGGCATTTTG CCTTCCTGTT TTTGCTCACC 6420
  • AGCTAACCGC TTTTTTGCAC AACATGGGGG ATCATGTAAC TCGCCTTGAT CGTTGGGAAC 6840 CGGAGCTGAA TGAAGCCATA CCAAACGACG AGCGTGACAC CACGATGCCT GTAGCAATGG 6900
  • MOLECULE TYPE DNA (genomic)
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • ORIGINAL SOURCE
  • GAAAGACAAC AACGGGCCGC GCGCTCGCGG CCGCTTCCAC CGCTGGGGAG GGCGTGCTCT 1500 TTTTCCCGGA GCCGATGGCG TACTGGCGCA CGATGTTTGG TACGGACGCC TTAAGTGGGA 1560
  • GAT TCGCCA TTTTTCCAAA AGTTGATTTT TGGGCATACG CGATATCTGG CGATACGCTT 2100 ATATCGTTTA CGGGGGATGG CGATAGACGC CTTTGGTGAC TTGGGCGATT CTGTGTCG 2160
  • GGCACATCCC CTTCTCTGTG ACACACCCTG TCCACGCCCC TGGTTCTTAG TTCCAGCCCC 4860
  • GCCTGGTATC TTTATAGTCC TGTCGGGTTT CGCCACCTCT GACTTGAGCG TCGATTTTTG 7920 TGATGCTCGT CAGGGGGGCG GAGCCTATGG AAAAACGCCA GCAACGCGGC CTTTTTACGG 7980

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Abstract

Cette invention se rapporte au virus de la maladie des muqueuses (BVDV) et à des vaccins utilisés dans le traitement de cette maladie. Cette invention concerne la préparation de l'herpèsvirus des bovins vivants atténués, de type 1 (BHV-1), sous forme d'un virus, d'un vaccin et d'un vecteur d'expression des antigènes de BVDV. Un clone d'ADNc du BVDV contenant des séquences correspondant à la glycoprotéine gp53 est inséré dans un virus BHV-1 inactivé.
PCT/US1994/012198 1993-11-05 1994-10-31 Vecteur viral a antigenes du virus de la maladie des muqueuses (bvdv) WO1995012682A2 (fr)

Priority Applications (5)

Application Number Priority Date Filing Date Title
AU10423/95A AU688819B2 (en) 1993-11-05 1994-10-31 Viral vector with bovine viral diarrhea virus (BVDV) antigens
KR1019960702343A KR960705944A (ko) 1993-11-05 1994-10-31 소 바이러스성 설사 바이러스(bvdv) 항원을 갖는 바이러스 벡터(viral vector with bovine viral diarrhea virus(bvdv)antigens)
JP7513263A JPH09504435A (ja) 1993-11-05 1994-10-31 ウシウイルス性下痢ウイルス（ｂｖｄｖ）抗原を有するウイルスベクター
NZ276234A NZ276234A (en) 1993-11-05 1994-10-31 An attenuated replicating non pathogenic flavivirus (bovine viral diarrhoea virus - bvdv) vaccine
EP95901037A EP0725831A1 (fr) 1993-11-05 1994-10-31 Vecteur viral a antigenes du virus de la maladie des muqueuses (bvdv)

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US14781093A 1993-11-05 1993-11-05
US08/147,810 1993-11-05

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WO1995012682A3 WO1995012682A3 (fr) 1995-07-06

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Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6060457A (en) * 1996-06-20 2000-05-09 Universite De Montreal DNA plasmid vaccine for immunization of animals against BVDV
EP1026252A1 (fr) * 1999-02-02 2000-08-09 Akzo Nobel N.V. Gène synthétique du virus de la diarrhée virale des bovins
EP1104676A1 (fr) * 1999-11-30 2001-06-06 Boehringer Ingelheim Vetmedica Gmbh Virus de la diarrhée des bovins surs attenués pour utilisation chez des vaches gravides
WO2002000881A1 (fr) * 2000-06-27 2002-01-03 Bayer Aktiengesellschaft Pseudo-particules virales du virus de la diarrhee virale des bovins (bvd)
EP1743652A1 (fr) 2002-08-26 2007-01-17 Pfizer Products Inc. Vaccin contre les infections de l'appareil génital et du système respiratoire chez les bovins
WO2014151401A1 (fr) 2013-03-15 2014-09-25 Zoetis Llc Protection croisée de bovins contre une infection par b. trehalosi par un vaccin multivalent

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR100331176B1 (ko) * 1999-12-15 2002-04-06 대한민국(관리청:특허청장, 승계청:국립수의과학검역원장) 재조합 단백질을 항원으로 이용한 소 바이러스성 설사증의진단방법
BRPI0417225A (pt) * 2003-12-05 2007-03-06 Becton Dickinson Co métodos de aumentar a resposta imune no compartimento intradérmico e compostos úteis nos referidos métodos
KR101876535B1 (ko) * 2012-06-14 2018-07-09 베트올 (주) 소바이러스성 설사병 바이러스의 탐지용 항체, 이를 이용한 항원 검출 방법, 및 이를 포함하는 탐지키트
CN113913461A (zh) * 2021-11-15 2022-01-11 贵州大学 牛病毒性腹泻e0-e2基因重组腺病毒疫苗构建方法

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0119025A2 (fr) * 1983-02-25 1984-09-19 Baylor College of Medicine Mutant d'herpès virus-1 bovin thymidine kinase négatif résistant à la température, comme vaccin contre la rhinotrachéite infectieuse bovine
WO1990001337A1 (fr) * 1988-08-03 1990-02-22 Institute For Animal Health Limited Vaccin
EP0464010A1 (fr) * 1990-06-08 1992-01-02 Statens Veterinärmedicinska Anstalt Procédé pour la détection d'une infection due à un virus, d'un type spécifique, des amorces, des sondes et trousse d'essai
WO1994000586A2 (fr) * 1992-06-26 1994-01-06 Rhône Merieux Mutants et vaccins du virus de la rhinotracheite infectieuse bovine

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0119025A2 (fr) * 1983-02-25 1984-09-19 Baylor College of Medicine Mutant d'herpès virus-1 bovin thymidine kinase négatif résistant à la température, comme vaccin contre la rhinotrachéite infectieuse bovine
WO1990001337A1 (fr) * 1988-08-03 1990-02-22 Institute For Animal Health Limited Vaccin
EP0464010A1 (fr) * 1990-06-08 1992-01-02 Statens Veterinärmedicinska Anstalt Procédé pour la détection d'une infection due à un virus, d'un type spécifique, des amorces, des sondes et trousse d'essai
WO1994000586A2 (fr) * 1992-06-26 1994-01-06 Rhône Merieux Mutants et vaccins du virus de la rhinotracheite infectieuse bovine

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
VIROLOGY, vol.190, no.2, 1992 pages 666 - 673 BELLO, L. ET AL. 'Bovine herpesvirus 1 as a live virus vector for expression of foreign genes' *
VIROLOGY, vol.190, no.2, 1992 pages 763 - 772 PATON, D.J. ET AL. 'Epitope mapping of the gp53 envelope protein of bovine viral diarrhea virus' *
VIRUS RESEARCH, vol.34, no.2, 1994 pages 178 - 186 YU, M. ET AL. 'High level expression of the envelope glycoprotein (GP53) of bovine viral diarrhoea virus (singer) and its potential use as diagnostic reagent' *

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6060457A (en) * 1996-06-20 2000-05-09 Universite De Montreal DNA plasmid vaccine for immunization of animals against BVDV
EP1026252A1 (fr) * 1999-02-02 2000-08-09 Akzo Nobel N.V. Gène synthétique du virus de la diarrhée virale des bovins
EP1104676A1 (fr) * 1999-11-30 2001-06-06 Boehringer Ingelheim Vetmedica Gmbh Virus de la diarrhée des bovins surs attenués pour utilisation chez des vaches gravides
WO2001039801A2 (fr) * 1999-11-30 2001-06-07 Boehringer Ingelheim Vetmedica Gmbh Virus attenues et sans danger de la diarrhee virale bovine utilisable avec des vaches pleines
WO2001039801A3 (fr) * 1999-11-30 2002-02-07 Boehringer Ingelheim Vetmed Virus attenues et sans danger de la diarrhee virale bovine utilisable avec des vaches pleines
US6610305B1 (en) 1999-11-30 2003-08-26 Boehringer Ingelheim Vetmedica Gmbh Safe attenuated bovine viral diarrhea viruses for use in pregnant cows
AU777991B2 (en) * 1999-11-30 2004-11-11 Boehringer Ingelheim Vetmedica Gmbh Safe attenuated bovine viral diarrhea viruses for use in pregnant cows
WO2002000881A1 (fr) * 2000-06-27 2002-01-03 Bayer Aktiengesellschaft Pseudo-particules virales du virus de la diarrhee virale des bovins (bvd)
EP1170367A1 (fr) 2000-06-27 2002-01-09 Bayer Ag Particules de type viral du BVDV
EP2365082A1 (fr) 2000-06-27 2011-09-14 Pfizer Animal Health S.A. Particules de type viral du BVDV
EP1743652A1 (fr) 2002-08-26 2007-01-17 Pfizer Products Inc. Vaccin contre les infections de l'appareil génital et du système respiratoire chez les bovins
WO2014151401A1 (fr) 2013-03-15 2014-09-25 Zoetis Llc Protection croisée de bovins contre une infection par b. trehalosi par un vaccin multivalent

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WO1995012682A3 (fr) 1995-07-06
NZ276234A (en) 1998-01-26
AU688819B2 (en) 1998-03-19
KR960705944A (ko) 1996-11-08
EP0725831A1 (fr) 1996-08-14
CN1134175A (zh) 1996-10-23
CA2172815A1 (fr) 1995-05-11
JPH09504435A (ja) 1997-05-06
MXPA94008605A (es) 2004-11-11

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