[go: up one dir, main page]

WO2025046414A1 - Vecteurs de cdv via et leurs utilisations - Google Patents

Vecteurs de cdv via et leurs utilisations Download PDF

Info

Publication number
WO2025046414A1
WO2025046414A1 PCT/IB2024/058182 IB2024058182W WO2025046414A1 WO 2025046414 A1 WO2025046414 A1 WO 2025046414A1 IB 2024058182 W IB2024058182 W IB 2024058182W WO 2025046414 A1 WO2025046414 A1 WO 2025046414A1
Authority
WO
WIPO (PCT)
Prior art keywords
cdv
aiv
vector
recombinant
antigen
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/IB2024/058182
Other languages
English (en)
Inventor
Frederic Reynard
Michel Bublot
Teshome Mebatsion
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Boehringer Ingelheim Vetmedica GmbH
Original Assignee
Boehringer Ingelheim Vetmedica GmbH
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Boehringer Ingelheim Vetmedica GmbH filed Critical Boehringer Ingelheim Vetmedica GmbH
Publication of WO2025046414A1 publication Critical patent/WO2025046414A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • 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
    • A61P31/14—Antivirals for RNA viruses
    • 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
    • A61P31/14—Antivirals for RNA viruses
    • A61P31/16—Antivirals for RNA viruses for influenza or rhinoviruses
    • 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
    • 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
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/525—Virus
    • A61K2039/5256—Virus expressing foreign proteins
    • 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
    • A61K2039/55—Medicinal preparations containing antigens or antibodies characterised by the host/recipient, e.g. newborn with maternal antibodies
    • A61K2039/552—Veterinary vaccine
    • 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
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/16011—Orthomyxoviridae
    • C12N2760/16111—Influenzavirus A, i.e. influenza A virus
    • C12N2760/16122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • 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
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/16011—Orthomyxoviridae
    • C12N2760/16111—Influenzavirus A, i.e. influenza A virus
    • C12N2760/16134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • 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
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/18011—Paramyxoviridae
    • C12N2760/18411—Morbillivirus, e.g. Measles virus, canine distemper
    • C12N2760/18434—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • 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
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/18011—Paramyxoviridae
    • C12N2760/18411—Morbillivirus, e.g. Measles virus, canine distemper
    • C12N2760/18441—Use of virus, viral particle or viral elements as a vector
    • C12N2760/18443—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector

Definitions

  • the present invention relates to recombinant canine distemper virus (CDV) viral vectors comprising a heterologous polynucleotide encoding a foreign avian influenza (AIV) antigen, for example an AIV hemagglutinin serotype H9 (AIV H9-HA) antigen.
  • CDV canine distemper virus
  • the recombinant viral vectors are suitable for use in immunogenic compositions and vaccines, and can provide protection against avian influenza.
  • Avian influenza sometimes called avian flu, and commonly recognized as bird flu refers to influenza caused by influenza viruses adapted to birds.
  • AIV is a segmented, singlestrand, negative sense RNA virus belonging to the family of Orthomyxoviridae, and is classified as a type A influenza virus.
  • Type A virus is the most frequent cause of animal and human influenza. This type occurs in numerous strains or subtypes that are differentiated mainly on the basis of two surface lipid-enveloped membrane proteins, hemagglutinin (HA) and neuraminidase (NA).
  • HA facilitates entry of the virus into host cells
  • NA assists in the release of progeny virus from infected cells (de Jong et al., J Clin Virol.
  • Influenza type A viruses are divided into subtypes based on their specific HA and NA content. There are 16 different HA subtypes, and 9 different NA subtypes. Many different combinations of HA and NA proteins are possible. Subtypes of influenza A virus are named according to their HA and NA surface proteins.
  • Avian influenza H9N2 virus is considered a low-pathogenic virus that is endemic to poultry populations. Low pathogenicity avian influenza H9N2 has adverse effects on poultry production and poses a significant cross-species transmission and zoonotic threat.
  • Canine distemper virus (CDV) belongs to the Paramyxoviridae family and the Morbillivirus genus.
  • Canine distemper the disease caused by CDV, is a highly infectious, febrile disease of dogs and other mammal families, in particular those of the order Carnivora.
  • the transmission of CDV is usually via the oral or respiratory routes, with sneezing, coughing and sharing of food or water bowls identified as routes of transmission.
  • CDV replicates in the lymphatic tissue of the respiratory tract, followed by infection of respiratory, gastrointestinal and urogenital epithelium (Whitehouse, CABI Compendium “canine distemper”, 2015; Creevy K.E., Overview of Canine Distemper. In: The Merck Veterinary Manual. 2013).
  • the mortality rate of CDV in canines is high, ranging between 30 and 80 percent. CDV is not known to routinely infect avian populations.
  • CDV is an enveloped virus containing a single stranded RNA genome, 15,690 nucleotides in length.
  • the genomic RNA contains six genes that encode the following proteins in the order of: the nucleocapsid protein (NP), phosphoprotein (P), matrix protein (M), fusion protein (F), hemagglutinin glycoprotein (H), and large protein (L), which are sited between a short 3’ leader region and followed by a short 5’ trailer region.
  • the six genes are separated by intergenic regions of 3 nucleotides.
  • two non-structural proteins, V and C are encoded within the P gene (Zhao, et al., Viruses, 12(3):339, 2020)
  • WO 2019/057859 further describes an expression cassette for insertion between two adjacent essential genes of a Paramyxoviridae virus, creating a virus vector that can accommodate, maintain and express the foreign gene, encoded by the expression cassette, for long periods of time.
  • the disclosed vectors of WO 2019/057859 are designed for mammals, in particular swine, and describe canine, parvovirus VP2, H3 -subtype hemagglutinin of swine influenza virus, a spike protein of porcine epidemic diarrhoea virus, various coronavirus, and avian infectious bronchitis virus S protein.
  • Suitable Paramyxoviridae viruses include CDV, feline morbillivirus, and peste-des-petits-ruminants virus.
  • the disclosed studies are limited to the Lederle CDV vaccine strain and vaccination of pigs, resulting in some immunogenicity. Therefore, there is no indication that CDV vectors would be efficacious in other species.
  • the present application describes recombinant CDV vectors comprising a heterologous polynucleotide encoding an avian influenza (AIV) H9 haemagglutinin antigen (AIV H9-HA, also referred to simply as H9), derived from a Saudi Arabia AIV H9N2 strain.
  • AIV avian influenza
  • AIV H9-HA haemagglutinin antigen
  • the recombinant viral vectors can be used in immunogenic compositions and vaccines to provide animals with protection against avian influenza and/or CDV.
  • the present invention provides a recombinant canine distemper virus (CDV) vector comprising a heterologous polynucleotide encoding an Avian Influenza Virus (AIV) antigen.
  • CDV canine distemper virus
  • AIV Avian Influenza Virus
  • the present invention provides one or more nucleic acid molecules comprising the recombinant CDV vector of the invention.
  • the present invention provides a cell comprising the recombinant CDV vector or the nucleic acid molecules of the invention.
  • the present invention provides a composition comprising the recombinant CDV vector, the nucleic acid molecules or the cell of the invention.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of inducing a protective immune response against AIV and/or CDV, preferably AIV, in an animal.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of inducing a protective immune response against AIV in an avian.
  • the present invention provides a method of inducing a protective immune response against AIV and/or CDV, preferably AIV, in an animal, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the animal.
  • the present invention provides a method of inducing a protective immune response against AIV in an avian, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the avian.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for inducing a protective immune response against AIV and/or CDV, preferably AIV, in an animal.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for inducing a protective immune response against AIV in an avian.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of reducing AIV shedding in an avian.
  • the present invention provides a method of reducing AIV shedding in an avian, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the avian.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for reducing AIV shedding in an avian.
  • the present invention provides a method of manufacturing a recombinant CDV vector, the method comprising: a) providing one or more nucleic acids encoding a CDV vector; b) providing a polynucleotide encoding an AIV antigen; and c) recombinantly combining the nucleic acids encoding the CDV vector and the polynucleotide encoding the AIV antigen.
  • FIG. 1 shows a pCG219 plasmid map.
  • FIG. 2 shows a pCG171 plasmid map.
  • FIG. 3 shows a pCG172 plasmid map.
  • FIG. 4 shows a pCG173 plasmid map.
  • FIG. 5 shows expression results for vCDV-6.
  • FIG. 6 shows kinetics of mean clinical daily scores of different groups vaccinated (Gl) or not (G2) with vCDV6 vaccine candidate expressing H9N2 HA gene after H9N2 challenge at D21.
  • FIG. 7 shows dispersions of individual global clinical scores by groups.
  • Gl vCDV6 and G2: controls.
  • GCS Mann-Whitney- Wilcoxon test. Box plots represent the mini, the lower quartile, the median, the upper quartile and the maxi).
  • FIG. 8 shows H9N2 shedding after challenge.
  • Left panel mean oro-pharyngeal H9N2 RNA excretion by groups expressed as equivalent loglO EID50/mM; the limit of detection was 2.09 loglO equivalent EID50/ml.
  • FIG. 9 shows dispersions of individual AUC of viral RNA by groups; box plots represent the mini, the lower quartile, the median, the upper quartile and the maxi.
  • FIG. 10 shows dispersions of individual HAI titers against antigen IRAK on D20 by groups (loglO HAI titers); box plots represent the mini, the lower quartile, the median, mean (cross), the upper quartile and the maxi. Gl: vCDV6 and G2: controls. CTL group remained negative on D21. Seroconversion occurred in the CDV group. DETAILED DESCRIPTION
  • the present invention provides recombinant CDV vectors comprising a heterologous polynucleotide encoding an avian influenza AIV H9-HA antigen.
  • the present invention provides a novel recombinant CDV vector comprising a heterologous polynucleotide encoding an AIV H9-HA antigen, for example from a H9N2 Saudi Arabia strain of AIV.
  • the polynucleotide encoding the AIV H9-HA antigen of the present invention can be defined with reference to SEQ ID NO: 1 (H9 gene of H9N2 Saudi Arabia strain).
  • the AIV H9-HA antigen of the present invention can be defined with reference to polypeptide sequence of SEQ ID NO: 2 (H9 gene of H9N2 Saudi Arabia strain).
  • the recombinant CDV viral vectors of the invention can be used in immunogenic compositions and vaccines to provide animals with protection against avian influenza and/or canine distemper.
  • the recombinant viral vectors of the present invention provide a combined CDV and AIV vector vaccine which is surprisingly effective even in stringent clinical conditions, with an unexpected level of efficacy that can achieve advantageous results such as a significant decrease in viral excretion and a reduction in clinical signs.
  • the recombinant CDV viral vectors of the invention can be used in immunogenic compositions and vaccines to provide avians with protection against avian influenza.
  • Benefits of the viral vectors of the invention include the capability to induce a protective immune response against AIV infection, including a reduction in viral excretion, even when administered just once and to avians of six or fewer days of age, with demonstrated efficacy via the subcutaneous route of administration.
  • the present invention provides a recombinant canine distemper virus (CDV) vector comprising a heterologous polynucleotide encoding an Avian Influenza Virus (AIV) antigen.
  • CDV canine distemper virus
  • AIV Avian Influenza Virus
  • the AIV antigen is an AIV haemagglutinin antigen.
  • the AIV antigen is an AIV subtype H9 haemagglutinin (AIV H9- HA) antigen.
  • AIV H9-HA antigen is a Saudi Arabia strain H9-HA antigen.
  • the polynucleotide encoding the AIV H9-HA antigen is a Saudi Arabia strain H9-HA polynucleotide.
  • the polynucleotide encoding the AIV antigen comprises a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 95.0% 95.1%, 95.2%, 95.3%, 95.4%, 95.5%, 95.6%, 95.7%, 95.8%, 95.9%, 96%, 96.0%, 96.1%, 96.2%, 96.3%, 96.4%, 96.5%, 96.6%, 96.7%, 96.8%, 96.9%, 97%, 97.0%, 97.1%, 97.2%, 97.3%, 97.4%, 97.5%, 97.6%,
  • the AIV antigen comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 95.0% 95.1%, 95.2%, 95.3%, 95.4%, 95.5%, 95.6%, 95.7%, 95.8%, 95.9%, 96%, 96.0%, 96.1%, 96.2%, 96.3%, 96.4%, 96.5%, 96.6%, 96.7%, 96.8%, 96.9%, 97%, 97.0%, 97.1%, 97.2%, 97.3%, 97.4%, 97.5%, 97.6%, 97.7%, 97.8%, 97.9%, 98%, 98.0%, 98.1%, 98.2%, 98.3%, 98.4%, 98.5%, 98.6%, 98.7%, 98.0%,
  • the polynucleotide encoding the AIV antigen comprises a sequence having at least 80% identity to SEQ ID NO: 1, and/or the AIV antigen comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 2.
  • the polynucleotide encoding the AIV H9-HA antigen comprises a sequence having at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 95.0% 95.1%, 95.2%, 95.3%, 95.4%, 95.5%, 95.6%, 95.7%, 95.8%, 95.9%, 96%, 96.0%, 96.1%, 96.2%, 96.3%, 96.4%, 96.5%, 96.6%, 96.7%, 96.8%, 96.9%, 97%, 97.0%, 97.1%, 97.2%, 97.3%, 97.4%, 97.5%, 97.6%, 97.7%, 97.8%, 97.9%, 98%, 98.0%, 98.1%, 98.2%, 98.3%, 98.0%, 98.1%
  • the polynucleotide encoding the AIV H9-HA antigen comprises a sequence having at least 80% identity to SEQ ID NO: 1.
  • the AIV H9-HA antigen comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 95.0% 95.1%, 95.2%, 95.3%, 95.4%, 95.5%, 95.6%, 95.7%, 95.8%, 95.9%, 96%, 96.0%, 96.1%, 96.2%, 96.3%, 96.4%, 96.5%, 96.6%, 96.7%, 96.8%, 96.9%, 97%, 97.0%, 97.1%, 97.2%, 97.3%, 97.4%, 97.5%, 97.6%, 97.7%, 97.8%, 97.9%, 97%, 97.
  • the AIV H9-HA antigen comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 2.
  • the polynucleotide encoding the AIV H9-HA antigen comprises the sequence of SEQ ID NO: 1, and/or the AIV H9- HA antigen comprises the amino acid sequence of SEQ ID NO: 2.
  • the polynucleotide encoding the AIV H9-HA antigen consists of the sequence of SEQ ID NO: 1, and/or the AIV H9- HA antigen consists of the amino acid sequence of SEQ ID NO: 2.
  • the present invention relates to particular AIV antigens, which may be defined with reference to SEQ ID NOs 1 and 2, which are advantageous in a CDV vector context for the reasons outlined herein.
  • the avian influenza H9 HA polynucleotide is from strain A/avian/Saudi Arabia/910135/2006 (H9N2), and encodes a polypeptide defined by GenBank No: ACY80655.1, which is incorporated by reference herein in its entirety.
  • the genome of the CDV vector comprises a sequence having at least 80% identity to SEQ ID NO: 3, into which the heterologous polynucleotide encoding the AIV antigen has been inserted.
  • the genome of the CDV vector comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 3, into which the heterologous polynucleotide encoding the AIV antigen has been inserted.
  • the recombinant CDV vector of the invention comprises a CDV vector genome of SEQ ID NO: 3 (or a sequence having % identity to SEQ ID NO: 3 as defined herein), that a heterologous polynucleotide encoding an AIV antigen of the invention has been inserted into.
  • the recombinant CDV vector of the invention comprises a CDV vector genome sequence defined in relation to SEQ ID NO: 3 that is interrupted by the polynucleotide encoding the AIV antigen of the invention as well as any other recombinant elements included in the AIV antigen expression cassette.
  • the AIV antigen is an AIV H9-HA antigen.
  • the CDV vector is an Onderstepoort strain CDV vector.
  • the CDV vector is an Onderstepoort strain CDV vector comprising a genome sequence having at least 80% identity to SEQ ID NO: 3.
  • the recombinant CDV vector of the invention is a CDV Onderstepoort strain vector comprising a genome sequence having at least 80% identity to SEQ ID NO: 3, into which the heterologous polynucleotide encoding the AIV antigen has been inserted.
  • the recombinant CDV vector of the invention is a CDV Onderstepoort strain vector comprising a genome sequence having at least at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 3, into which the heterologous polynucleotide encoding the AIV antigen has been inserted.
  • the recombinant Onderstepoort strain CDV vector of the invention comprises an Onderstepoort strain CDV vector genome of SEQ ID NO: 3 (or a sequence having % identity to SEQ ID NO: 3 as defined herein), that a heterologous polynucleotide encoding an AIV antigen of the invention has been inserted into.
  • the CDV vector is an Onderstepoort strain CDV vector comprising the genome sequence set forth in ENA/EBI accession no. AF014953.1 (which is incorporated by reference herein).
  • the recombinant Onderstepoort strain CDV vector of the invention comprises an Onderstepoort strain CDV vector genome sequence defined in relation to SEQ ID NO: 3 that is interrupted by the AIV heterologous polynucleotide of the invention as well as any other recombinant elements included in the AIV antigen expression cassette.
  • the AIV antigen is an AIV H9-HA antigen.
  • SEQ ID NO: 3 is the sequence of pCGl 77, a transfer vector comprising transfer vector elements (e.g. a T7 promoter, terminator, rop (repressor of primer) and ori (origin of replication) sequences) as well as a CDV Onderstepoort genome.
  • transfer vector elements may be present in the recombinant CDV vector of the invention or may be absent from the recombinant CDV vector of the invention.
  • the recombinant CDV vector of the invention comprises transfer vector elements.
  • the recombinant CDV vector of the invention does not comprise transfer vector elements.
  • the polynucleotide sequence of the recombinant CDV vector of the invention consists of one or more CDV genomic sequences and an AIV H9-HA sequence.
  • embodiments of the present invention relate to a CDV vector comprising SEQ ID NO: 3 or a sequence with at least 80% identity thereto, into which a polynucleotide encoding an AIV H9-HA antigen has been inserted, preferably between the CDV P and M genes. In essence, this results in the provision of pCG219 as illustrated in Fig. 1.
  • the transfer vector elements are not present in SEQ ID NO: 3.
  • non-CDV elements are not present in SEQ ID NO: 3.
  • sequence of the invention defined in respect of SEQ ID NO: 3 or a sequence with identity thereto is defined in respect of positions 3914-19603 of SEQ ID NO: 3 or a sequence with identity to positions 3914-19603 of SEQ ID NO: 3.
  • the reference to SEQ ID NO: 3 is understood as solely referring to positions 3914-19603 of SEQ ID NO: 3.
  • the present invention relates to a recombinant CDV Onderstepoort vector comprising a heterologous polynucleotide.
  • the heterologous polynucleotide in such a vector is not limited, e.g. to AIV.
  • the recombinant CDV Onderstepoort vector is as defined herein, e.g. with reference to SEQ ID NO: 3.
  • the heterologous polynucleotide encodes a polypeptide.
  • the heterologous polynucleotide encodes a heterologous antigen.
  • the heterologous antigen is derived from a pathogen other than CDV.
  • the heterologous polynucleotide is expressible or expressed.
  • the recombinant CDV vector is capable of eliciting heterologous protection against the pathogen from which the heterologous antigen is derived.
  • the heterologous polynucleotide is inserted between the P gene and the M gene in the CDV genome.
  • the heterologous polynucleotide encoding the AIV antigen is inserted between the P and M genes of the CDV vector genome.
  • the heterologous polynucleotide encoding the AIV antigen is inserted between the P gene and the M gene of the CDV vector genome defined in relation to SEQ ID NO: 3.
  • the AIV antigen is an AIV H9-HA antigen.
  • the recombinant CDV vectors comprise a promoter.
  • the promoter is selected from a murine cytomegalovirus (mCMV) promoter, a vaccinia virus H6 promoter, a T7 promoter, and a CDV genome promoter.
  • the promoter driving the AIV antigen expression is a CDV genome promoter.
  • the expression of the AIV antigen is regulated by the promoter.
  • the expression of the AIV antigen is regulated by the CDV genome promoter.
  • the AIV antigen is an AIV H9-HA antigen.
  • the transcription of the entire CDV genome is under the control of the CDV genome promoter.
  • Each gene in the CDV genome is flanked by transcription start and transcription stop sequences. Therefore, the inserted heterologous polynucleotide encoding the AIV antigen is also flanked by virus- specific transcription start and transcription stop sequences. It should thus be understood in this context that it is the genomic RNA of the CDV virus that is under the control of the CDV genome promoter and it does not mean that the heterologous polynucleotide encoding the AIV antigen is flanked by the CDV genome promoter.
  • a recombinant promoter e.g.
  • a heterologous non-CDV promoter may be used to control the expression of the polynucleotide encoding the AIV antigen.
  • the recombinant promoter is located 5’ of, such as directly 5’ of, the polynucleotide encoding the AIV antigen.
  • the heterologous polynucleotide encoding the AIV antigen is inserted between the P and M genes and the expression of the AIV antigen is regulated by the CDV genome promoter.
  • the heterologous polynucleotide encoding the AIV antigen is inserted between the P and M genes of the CDV vector genome of SEQ ID NO: 3 (or a sequence having % identity to SEQ ID NO: 3 as defined herein) and the expression of the AIV antigen is regulated by the CDV genome promoter.
  • the AIV antigen is an AIV H9-HA antigen.
  • the recombinant CDV vector comprises a polyadenylation (poly A) signal.
  • the polyA signal is a simian virus 40 (SV40) polyA tail.
  • expression of the AIV antigen is influenced by the polyA signal.
  • the polyA signal is located downstream of the heterologous polynucleotide encoding the AIV antigen.
  • the AIV antigen is an AIV H9-HA antigen.
  • the recombinant CDV vector of the invention is useful as a vaccine against CDV and/or AIV, which can be administered to an animal to induce heterologous protection against CDV and/or AIV.
  • the recombinant CDV vector is capable of inducing a protective immune response against CDV.
  • the recombinant CDV vector is capable of inducing a protective immune response against AIV.
  • the recombinant CDV vector is capable of inducing a protective immune response against CDV and AIV.
  • the CDV vector is capable of eliciting an immune response against AIV in an avian.
  • the CDV vector is capable of eliciting a protective immune response against AIV in an avian.
  • the protective immune response comprises a reduction of clinical signs caused by CDV and/or AIV.
  • the clinical signs are respiratory signs.
  • the protective immune response comprises reducing shedding caused by AIV in an avian.
  • the recombinant CDV vector of the invention is capable of reducing shedding caused by AIV in an avian.
  • the recombinant CDV vector of the invention is capable of reducing shedding caused by AIV in an avian following AIV infection or challenge.
  • the protective immune response comprises reducing clinical signs caused by CDV and/or AIV as well as reducing shedding caused by AIV.
  • the reduction is relative to the clinical signs of an equivalent unvaccinated animal or avian, such as an animal or avian that has not received a CDV and/or AIV vaccine and has been infected or challenged with CDV and/or AIV.
  • one dose of the recombinant CDV vector is capable of inducing a protective immune response against AIV and/or CDV.
  • one dose of the recombinant CDV vector is capable of inducing a protective immune response against AIV.
  • one dose of the recombinant CDV vector is capable of inducing a protective immune response against CDV. In an embodiment, this means that a protective immune response against AIV and/or CDV will be induced in an animal that has received (only) a single administration of the recombinant CDV vector of the invention.
  • a protective immune response against AIV and/or CDV will be induced in an animal that has received (only) a single administration of any AIV and/or CDV vaccine or antigen, wherein the single administration is of the recombinant CDV vector of the invention.
  • one dose alone of the recombinant vector is capable of inducing a protective immune response against AIV and/or CDV.
  • the protective immune response is induced when (only) a single dose of the recombinant CDV vector of the invention has been administered.
  • “one dose” means that a second or further dose of the recombinant CDV vector of the invention is not necessary for the induction of the protective immune response. In an embodiment, “one dose” means that a second or further dose of any CDV and/or AIV vaccine or antigen is not necessary for the induction of the protective immune response. In an embodiment “one dose” means that the animal has not been primed against the same pathogen(s) to which the invention relates. For example, the animal has not been primed with the same vaccine, vector or antigen to which the invention relates, or any other type of antigen (e.g. killed vaccine, subunit vaccine, etc). In an embodiment, the one dose is about 7.7 log 10 PFU.
  • the recombinant CDV vector is capable of inducing a protective immune response against AIV and/or CDV when administered to an avian of six or fewer days of age, five or fewer days of age, four or fewer days of age, three or fewer days of age, two or fewer days of age, one or fewer day of age, or one day of age, preferably one day of age.
  • the recombinant CDV vector of the invention is capable of inducing a protective immune response against AIV and/or CDV even when administered to an avian of one day of age.
  • the present invention provides one or more nucleic acid molecules comprising the recombinant CDV vector of the invention.
  • the present invention provides a cell comprising the recombinant CDV vector or the nucleic acid molecules of the invention.
  • the present invention provides a composition comprising the recombinant CDV vector, the nucleic acid molecules or the cell of the invention.
  • the present invention also provides immunogenic compositions and vaccines comprising any of the recombinant vectors described herein.
  • the recombinant viral vectors of the present invention can be used in immunogenic compositions and vaccines to elicit an immune response against avian influenza.
  • the recombinant viral vectors of the present invention can be used in immunogenic compositions and vaccines to provide animals, such as avians with protection against avian influenza.
  • the recombinant viral vectors of the present invention can be used in immunogenic compositions and vaccines to elicit an immune response against CDV.
  • the recombinant viral vectors of the present invention can be used in immunogenic compositions and vaccines to provide animals with protection against CDV.
  • the immunogenic compositions are effective to elicit an immune response in an animal, such as an avian, when administered to the animal. In some embodiments, the immunogenic compositions are effective to induce an immune response in an animal, such as an avian, when administered to the animal. In some embodiments, the immunogenic compositions are effective to stimulate an immune response in an animal, such as an avian, when administered to the animal. In embodiments, the immunogenic compositions of the present invention are formulated such that they are safe and effective to elicit immunity against avian influenza, and/or canine distemper when administered to an animal. In embodiments, the immunogenic compositions of the present invention are formulated such that they are safe and effective to elicit immunity against avian influenza when administered to an avian.
  • the vaccines are effective to elicit a protective immune response in an animal, such as an avian, when administered to the animal.
  • the vaccines are effective to induce a protective immune response in an animal, such as an avian, when administered to the animal.
  • the vaccines are effective to stimulate a protective immune response in an animal, such as an avian, when administered to the animal.
  • the vaccines of the present invention are formulated such that they are safe and effective to elicit protective immunity against avian influenza when administered to an avian.
  • the immunogenic compositions and vaccines comprise a recombinant CDV vector comprising an avian influenza H9 heterologous polynucleotide that encodes an H9 antigen.
  • the immunogenic compositions and vaccines comprise a pharmaceutically or veterinarily acceptable carrier, excipient, and/or adjuvant.
  • suitable pharmaceutically or veterinarily acceptable carriers include 0.9% NaCl (e.g., saline) solution, phosphate buffer, poly-(L-glutamate), lactated Ringer’s injection diluent (sodium chloride, sodium lactate, potassium chloride and calcium chloride), and polyvinylpyrrolidone.
  • Examples of pharmaceutically or veterinarily acceptable adjuvants include, (1) polymers of acrylic or methacrylic acid, maleic anhydride and alkenyl derivative polymers, (2) immunostimulating sequences (ISS), such as oligodeoxyribonucleotide sequences having one or more non-methylated CpG units, (3) an oil in water emulsion, (4) cation lipids containing a quaternary ammonium salt, e.g., DDA (5) cytokines, (6) aluminum hydroxide or aluminum phosphate, and/or (7) saponin.
  • ISS immunostimulating sequences
  • ISS immunostimulating sequences
  • oligodeoxyribonucleotide sequences having one or more non-methylated CpG units such as oligodeoxyribonucleotide sequences having one or more non-methylated CpG units
  • an oil in water emulsion such as an oil in water emulsion
  • the immunogenic compositions and vaccines are formulated for administration to an animal, such as an avian.
  • the immunogenic compositions and vaccines are formulated for administration by one or more of the following routes: aerosol (spray), ocular, nasal, oculonasal, oral, subcutaneous injection, and/or intramuscular injection.
  • the immunogenic compositions and vaccines are formulated for administration by subcutaneous injection.
  • the immunogenic compositions and vaccines can contain a titer of recombinant vector from 10 2 to 10 3 , 10 3 to 10 4 , 10 4 to 10 5 , 10 5 to 10 6 , 10 6 to 10 7 , 10 7 to 10 8 , 10 8 to 10 9 , 10 9 to IO 10 , 10 3 to 10 8 , 10 4 to 10 8 , or 10 5 to 10 7 , per dose.
  • the recombinant vector may be titrated based on any virus titration methods including, but not limited to, FFA (Focus Forming Assay) or FFU (Focus Forming Unit), TCID50 (50% Tissue Culture Infective Dose), EID50 (50% Egg Infective Dose), PFU (Plaque Forming Units), and FAID50 (50% Fluorescent Antibody Infectious Dose).
  • the dose volumes can be between 0.01 and 10 ml, between 0.01 and 5 ml, between 0.01 and 1 ml, or 0.01 and 0.5 ml.
  • the recombinant CDV vector of the present invention surprisingly induces a protective immune response against AIV due to the presence of the AIV (H9-HA) antigen.
  • the recombinant CDV vector of the present invention will induce a protective immune response against CDV due to the presence of the CDV vector genome (e.g. the backbone of the recombinant CDV vector of the invention), which is preferably SEQ ID NO: 3 (or a sequence with % identity to SEQ ID NO: 3 as defined herein).
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of inducing a protective immune response against AIV and/or CDV, preferably AIV, in an animal.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of inducing a protective immune response against AIV in an avian.
  • the present invention provides a method of inducing a protective immune response against AIV and/or CDV, preferably AIV, in an avian, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the animal.
  • the present invention provides a method of inducing a protective immune response against AIV in an avian, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the avian.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for inducing a protective immune response against AIV and/or CDV, preferably AIV, in an animal.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for inducing a protective immune response against AIV in an avian.
  • inducing a protective immune response comprises reducing clinical signs associated with infection or challenge by AIV and/or CDV.
  • the clinical signs are respiratory clinical signs.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in a method of reducing AIV shedding in an avian.
  • the present invention provides a method of reducing AIV shedding in an avian, comprising administering the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention to the avian.
  • the present invention provides the recombinant CDV vector, the nucleic acid molecules, the cell or the composition of the invention, for use in the manufacture of a medicament for reducing AIV shedding in an avian.
  • the recombinant CDV vector is administered once. In an embodiment, this means that the recombinant CDV vector is only administered once. In an embodiment, this means that second/further administrations of the recombinant CDV vector are not carried out. In an embodiment, the recombinant CDV vector is administered once and the protective immune response and/or reduction in shedding is obtained after such single administration. In another embodiment, this means that the initial administration of the recombinant CDV vector only comprises one dose to induce a protective immune response, but the administration of one or more booster administrations comprising further doses at a substantially later date are not excluded. In an embodiment, the one dose is about 7.7 log 10 PFU.
  • the recombinant CDV vector is administered to an avian at six or fewer days of age, five or fewer days of age four or fewer days of age, three or fewer days of age, two or fewer days of age, one or fewer days of age, or one day of age.
  • the recombinant CDV vector is administered to an avian at six or fewer days of age, preferably one day of age.
  • the recombinant CDV vector of the invention is administered to an avian of one day of age and induces a protective immune response against AIV.
  • the recombinant CDV vector is administered subcutaneously.
  • the present invention also provides methods of immunizing, methods for eliciting an immune response and methods for eliciting a protective immune response in an animal using any of the recombinant vectors, immunogenic compositions, and/or vaccines described herein.
  • the animal is an avian.
  • the animal is a chicken, for example a one day old chick.
  • the methods comprise administering to an animal, such as an avian, a recombinant vector according to the present invention.
  • the methods comprise administering to an animal, such as an avian, an immunogenic composition according to the present invention.
  • the methods are effective to elicit, induce, and/or stimulate an immune response against avian influenza and/or canine distemper in an animal.
  • the methods are effective to elicit, induce, and/or stimulate an immune response against avian influenza in an avian.
  • the methods comprise administering to an animal a vaccine comprising an effective amount of a recombinant vector according to the present invention.
  • the animal is vaccinated/immunized against avian influenza and/or CDV.
  • the methods are effective to elicit, induce, and/or stimulate a protective immune response against avian influenza and/or canine distemper in an animal, and thereby reduce and/or prevent clinical signs associated with subsequent avian influenza and/or CDV exposure, infection, challenge and/or disease in the animal, relative to a non-vaccinated control animal of the same species.
  • the methods are effective to elicit, induce, and/or stimulate a protective immune response against avian influenza in an avian, and thereby reduce and/or prevent clinical signs associated with subsequent avian influenza exposure, infection, challenge and/or disease in the avian, relative to a non-vaccinated control avian of the same species.
  • the protective immune response is effective to provide the animal with protection against subsequent avian influenza virus and/or CDV infection or challenge, and clinical disease and signs associated therewith.
  • the protective immune response is effective to provide the avian with protection against subsequent avian influenza virus infection or challenge, and clinical disease and signs associated therewith.
  • the recombinant vectors, immunogenic compositions, and/or vaccines may be administered in ovo 1 to 4 days before hatching.
  • the recombinant vectors, immunogenic compositions, and/or vaccines may be administered to a 1 day old, 2 day old, 3 day old, 4 day old, 5 day old, 6 day old, 7 day old, 8 day old, 9 day old, 10 day old, 11 day old, 12 day old, 13 day old, 14 day old, 15 day old, 16 day old, 17 day old, 18 day old, 19 day old, 20 day old, or 21 day old chicken.
  • a variety of administration routes may be used such as aerosol (spray), ocular, nasal, oculonasal, oral, subcutaneous injection, and intramuscular injection. In an embodiment, the administration routes are used in 1 day old chicks.
  • the animals that the recombinant CDV vectors of the invention are administered to are chickens.
  • the animals are SPF chickens.
  • the animals are 1 day old SPF chickens.
  • the present invention provides a method of manufacturing a recombinant CDV vector, the method comprising: a) providing one or more nucleic acids encoding a CDV vector; b) providing a polynucleotide encoding an AIV antigen; and c) recombinantly combining the nucleic acids encoding the CDV vector and the polynucleotide encoding the AIV antigen.
  • the AIV antigen is an AIV haemagglutinin antigen.
  • the AIV antigen is an AIV subtype H9 haemagglutinin (AIV H9-HA) antigen.
  • the polynucleotide encoding an AIV antigen is a polynucleotide of the invention as defined herein as SEQ ID NO: 1 (or a sequence with % identity to any of SEQ ID NO: 1 as defined herein, such as 80% identity to SEQ ID NO: 1).
  • the one or more nucleic acids encoding a CDV vector encode the CDV vector of SEQ ID NO: 3 (or a sequence with % identity to SEQ ID NO: 3 as defined herein, such as 80% identity to SEQ ID NO: 3)- [0085]
  • the method of manufacture further comprises step d) generating recombinant CDV particles from the recombinant nucleic acid obtained in step c).
  • the CDV particles are infectious CDV particles.
  • step d) of the method of manufacture involves a reverse genetics system for CDV.
  • step d) of the method of manufacture involves transcribing the recombinant nucleic acid obtained in step c).
  • the recombinant nucleic acid is comprised in a transcription plasmid.
  • the recombinant nucleic acid is transcribable/transcribed from a T7 promoter (i.e. a promoter recognised by T7 RNA polymerase).
  • step d) of the method of manufacture comprises expressing one or more nucleic acids encoding the CDV N, P and L proteins.
  • the method steps a), b), c) (and d)) are typically in this order (step a) followed by step b) followed by step c) followed by step d) when present).
  • the present invention also provides methods of making recombinant CDV vectors comprising an avian influenza H9 heterologous polynucleotide that encodes a hemagglutinin antigen (AIV H9-HA).
  • the methods comprise inserting a polynucleotide encoding an avian influenza H9 antigen into a CDV backbone vector.
  • the H9 antigen is from strain A/avian/Saudi Arabia/910135/2006(H9N2).
  • the methods further comprise inserting a promoter into the backbone vector.
  • the avian influenza H9 heterologous polynucleotide is operably linked to a promoter, and expression of the H9 antigen is regulated by the promoter.
  • the methods further comprise inserting a polyadenylation (poly A) signal into the backbone vector.
  • the polyA signal is a simian virus 40 (SV40) polyA tail.
  • the polyA signal is inserted downstream of the H9 heterologous polynucleotide.
  • the term “animal” includes all mammals and birds.
  • the animal may be selected from equine (e.g., horse), canine (e.g., dogs, wolves, foxes, coyotes, jackals), feline (e.g., lions, tigers, domestic cats, wild cats, other big cats, and other felines including cheetahs and lynx), bovine (e.g., cattle), swine (e.g., pig), ovine (e.g., sheep, goats, lamas, bison), avian (e.g., chicken, duck, goose, turkey, quail, pheasant, parrot, finches, hawk, crow, ostrich, emu and cassowary), primate (e.g., prosimian, tarsier, monkey, gibbon, ape), and humans.
  • equine e.g., horse
  • canine e.g., dogs
  • animal also includes an individual animal in all stages of development, including embryonic and fetal stages.
  • the animal referred to herein is a dog.
  • the animals referred to herein are avians.
  • an avian is a chicken.
  • the term “about” means approximately, in the region of, roughly, or around. When the term “about” is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth. In general, the term “about” is used herein to modify a numerical value above and below the stated value by a variance of 10%. Therefore, about 50 means in the range of 45-55. Numerical ranges recited herein by endpoints include all numbers and fractions subsumed within that range (e.g. 1 to 5 includes 1, 1.5, 2, 2.75, 3, 3.90, 4, and 5).
  • an “adjuvant” is a substance that is able to favor or amplify the cascade of immunological events, ultimately leading to a better immunological response (e.g., the integrated bodily response to an antigen).
  • An adjuvant is in general not required for the immunological response to occur, but favors or amplifies this response.
  • the terms “antigen” or “immunogen” mean a substance that induces a specific immune response in a host animal (e.g., an immune response of the humoral and/or cellular type directed against the antigen).
  • the antigen may be a whole organism, killed, attenuated or live; a subunit or portion of an organism; a recombinant vector containing an insert with immunogenic properties; a piece or fragment of DNA capable of inducing an immune response upon presentation to a host animal; a polypeptide, an epitope, a hapten, and the like.
  • avian includes, for example, chicken, chick, broiler, capon, duck, goose, turkey, grouse, quail, swan, squab, pigeon, pheasant, parrot, finches, hawk, crow, ostrich, emu and cassowary.
  • the term “avian” also includes an individual avian in all stages of development, including embryonic and fetal stages. In a preferred embodiment, an avian is a chicken.
  • carrier refers to a solvent or diluent in which a recombinant vector is formulated and/or administered.
  • Pharmaceutically and veterinarily acceptable carriers can be sterile liquids such as water and/or oils.
  • suitable oil-based carriers can include petroleum oils, animal oils, vegetable oils, and oils of synthetic origin (e.g., peanut oil, soybean oil, mineral oil, sesame oil, etc.).
  • suitable aqueous carriers can include water and aqueous solutions (e.g. aqueous saline solution, aqueous dextrose solution, glycerol solution, etc.).
  • fragments thereof relate to polypeptide or polynucleotide sequences having an equivalent function to the polypeptide or polynucleotide sequence in respect of which they are defined, i.e. SEQ ID NOs: 1 to 2.
  • equivalent function means that a polynucleotide fragment encodes an AIV H9-HA antigen.
  • equivalent function means that a polypeptide fragment comprises an AIV H9-HA antigen.
  • an AIV H9-HA antigen is defined as a sequence that is capable of inducing an immune response, preferably a protective immune response, against AIV, preferably H9N2 AIV.
  • the term “gene” is used broadly to refer to any segment of polynucleotide associated with a biological function.
  • the term “genome” refers to the heritable genetic information of a host organism.
  • the genome contemplated in the present invention can refer to the DNA or RNA of a virus or pathogenic organism.
  • the RNA may be a positive strand or a negative strand RNA.
  • heterologous means derived from a genetically distinct entity from the rest of the entity to which it is being compared.
  • a polynucleotide may be placed by genetic engineering techniques into a plasmid or vector derived from a different source, and is a heterologous polynucleotide.
  • a promoter removed from its native coding sequence and operatively linked to a coding sequence other than the native sequence is a heterologous promoter.
  • sequence identity refers to a relationship between two or more sequences, namely a reference sequence and a given sequence to be compared with the reference sequence. Sequence identity is determined by comparing the given sequence to the reference sequence after the sequences have been optimally aligned to produce the highest degree of sequence similarity, as determined by the match between strings of such sequences. Upon such alignment, sequence identity is ascertained on a position-by-position basis, e.g., the sequences are “identical” at a particular position if at that position, the nucleotides/amino acids are identical.
  • Sequence identity can be readily calculated by known methods, including but not limited to, those described in Computational Molecular Biology, Lesk, A. N., ed., Oxford University Press, New York (1988), Biocomputing: Informatics and Genome Projects, Smith, D. W., ed., Academic Press, New York (1993); Computer Analysis of Sequence Data, Part I, Griffin, A. M., and Griffin, H.
  • immunogenic composition refers to a composition that comprises at least one antigen which elicits an immunological response in a host to which the immunogenic composition is administered.
  • a “protective immune response” comprises an “immunological response” to a composition or vaccine is the development in the host of a cellular and/or antibody- mediated immune response to a composition or vaccine of interest.
  • an “immunological response” includes but is not limited to one or more of the following effects: the production of antibodies, B cells, helper T cells, and/or cytotoxic T cells, directed specifically to an antigen or antigens included in the composition or vaccine of interest.
  • the host will display a 1 protective or therapeutic immunological response such that resistance to new infection will be enhanced and/or the clinical severity of subsequent disease reduced.
  • Such protection can be demonstrated by a reduction in clinical disease signs relative to those normally displayed by an infected host, a lack of clinical disease signs relative to those normally displayed by an infected host, a quicker recovery time relative to that normally displayed by an infected host, and/or a lowered pathogen count relative to that normally found in an infected host.
  • nucleic acid and “polynucleotide” refer to RNA, DNA, and derivatives thereof. It is to be understood that unless otherwise specified each reference to a “heterologous polynucleotide”, “polynucleotide sequence”, “nucleotide sequence”, “nucleic acid” or equivalent term encompasses not only the nucleotide sequence per se, but also the complement of that nucleotide sequence, the reverse of that nucleotide sequence, and the reverse complement of that nucleotide sequence.
  • references further encompass the RNA equivalent of that nucleotide sequence, the RNA complement (or transcript) of that nucleotide sequence, the reverse of the RNA equivalent of that nucleotide sequence and the reverse of the RNA complement (or transcript) of that nucleotide sequence. Unless otherwise indicated, such references further encompass any modifications or routine or non- substantive variations to any of those sequences, which are well-known in the art and further described herein.
  • the terms “pharmaceutically acceptable” and “veterinarily acceptable” are used adjectivally to mean that the modified noun is appropriate for use in a pharmaceutical or veterinary product.
  • pharmaceutical or veterinary vaccine when it is used, for example, to describe an excipient in a pharmaceutical or veterinary vaccine, it characterizes the excipient as being compatible with the other ingredients of the composition and not disadvantageously deleterious to the intended recipient.
  • polypeptide and “protein” are used interchangeably to refer to a polymer of consecutive amino acid residues.
  • protection does not require complete protection from any indication of infection.
  • protection can mean that, after challenge, clinical signs of the underlying infection are at least reduced, and/or that one or more of the underlying cellular, physiological, or biochemical causes or mechanisms causing the clinical signs are reduced and/or eliminated.
  • reduced means relative to the state of the infection, including the molecular state of the infection, not just the physiological state of the infection.
  • the term “recombinant” in the context of a polynucleotide or protein means a polynucleotide or protein with a semisynthetic or synthetic origin which either does not occur in nature or is linked to another a polynucleotide or protein in an arrangement not found in nature.
  • a recombinant polynucleotide or protein can be made by modifying, altering, or engineering a polynucleotide or protein from its native form or structure to a non-native form or structure.
  • the modification, alteration or engineering of a polynucleotide or protein may include, for example, the deletion of one or more nucleotides or amino acids, the substitution of one or more nucleotides or amino acids, and/or the insertion of one or more nucleotides or amino acids.
  • a “vaccine” is an immunogenic composition that is suitable for administration to an animal which, upon administration to the animal, induces an immune response strong enough to aid in the protection from a clinical disease arising from an infection with a wildtype pathogenic micro-organism (e.g., strong enough for aiding in the curing of, ameliorating of, protection against, and/or prevention of a clinical disease and/or clinical signs associated therewith).
  • This Example details the construction and characterization of a canine distemper virus (CDV) vector expressing avian influenza virus H9 gene from an H9N2 Saudi Arabia strain (vCDV-6).
  • CDV canine distemper virus
  • a plasmid harboring the CDV Onderstepoort Haig genome was generated in which was inserted the avian influenza virus H9 gene which comes from an H9N2 Saudi Arabia strain.
  • Vector CDV transcription plasmid pCG177 (comprising a CDV Onderstepoort vaccine strain; SEQ ID NO: 3).
  • Insert H9 gene from Influenza A virus (A/avian/Saudi Arabia/910135/2006(H9N2)) strain (Genbank: ACY80655.1). The gene has SEQ ID NO: 1.
  • the H9 Saudi Arabia gene was synthesized by GeneArt (no. 1166890) and sub cloned in pIV29. Then the H9 Saudi Arabia gene was amplified by PCR and cloned into pCG177 to replace the existing CPIV2 HA gene using the Gibson Assembly Master Mix (NEB, France).
  • Resistance gene Ampicillin.
  • Cloning The H9 Saudi Arabia gene was inserted in the pCG177 by cloning using the Gibson Assembly Master Mix (NEB, France).
  • Host cells Escherichia coli KI 2 strain NEB 10-beta (NEB, France).
  • Plasmid Map A map of plasmid pCG 219 is shown in FIG. 1.
  • the CDV is a negative RNA virus and the generation of genetically modified CDV virus can be accomplished with a reverse genetics system.
  • the transcription of a full length genomic viral RNA and the simultaneous expression of N, P and L proteins permit the assembly of RNP and the transcription of positive RNA into negative RNA genome. It initiates the normal replication cycle of CDV virus and permit the generation of infectious particles.
  • modified CDV particles the following reagents and conditions were used:
  • Transcription plasmid pCG219 The plasmid map is shown in FIG. 1. The amount transfected was 5 pg.
  • Expression N plasmid pCG171 The plasmid map is shown in FIG. 2. The amount transfected was 0.5 pg.
  • Expression P plasmid pCG172 The plasmid map is shown in FIG. 3. The amount transfected was 0.5 pg.
  • Expression L plasmid pCG173 The plasmid map is shown in FIG. 4. The amount transfected was 0.5 pg.
  • T7 RNA polymerase plasmid pNS151 The amount transfected was 0.7 pg.
  • Cells and transfection (a) Name and quantity of cells: CHO cells, 1.10 6 cells/well (6- well plate); (b) Medium used : MEM glutamax + SVF (GIBCO); (c) Confluency: subconfluent; (d) Transfection Medium: MEM (Gibco); (e) Transfecting agent: Lipofectamine LTX plus reagent (Invitrogen); (f) Incubation medium: Opti MEM medium (Gibco) without SVF; (g) Incubation time: 48h; (h) Co-culture on VERO cells (0.7 x 10 6 cells/well (6-well plate)); (i) Incubation 7 days; (j) Harvest: supernatant and cells.
  • Cell passages VERO cells.
  • Cell passage 2 had a titer of 5.28 loglO DICC50/ml.
  • Cell passage 3 had a titer of 5.27 loglO DICC50/ml.
  • the objective of this study was to assess the efficacy of a recombinant CDV candidate vaccine expressing the H9 of Avian Influenza H9N2, when administered to SPF chicks at one day of age, against a H9N2 virulent challenge (strain Saudi Arabia 2010) carried out 21 days postvaccination.
  • a H9N2 virulent challenge strain Saudi Arabia 2010
  • the following parameters were monitored after challenge: (i) respiratory and ocular symptoms (clinical monitoring); and (ii) oro-pharyngeal viral excretion (assessed by qRT-PCR).
  • G1 was treated with the CDV vaccine candidate expressing the H9 gene from the A/avian/Saudi Arabia/910135/2006 strain, and G2 was the unvaccinated control group, as indicated in Table 1 below:
  • Table 1 Group constitution and treatment [0136] Generation of vCDV6 is described in Example 1. The dose of the vaccine is shown in Table 1. The vCDV6 vaccine candidate was thawed, but not diluted before administration and was administered by the SC route.
  • mice from a same group were gathered in a nebulization box and nebulized with 4.2 ml for about 20 minutes. Animals were kept in closed contact in the nebulization box for about 3 to 5 minutes and then, they were moved back to their isolators. Chickens of all groups were daily monitored from D21 (before challenge) to D31. For this monitoring, particular attention was paid to respiratory and ocular symptoms. All other observed clinical signs were recorded as well. Respiratory and ocular symptoms were scored as detailed in Table 2.
  • Table 2 Description and score of respiratory and ocular clinical signs. If an animal was found dead or euthanized on ethical grounds after challenge, it was attributed a score of 6 daily until the end of the study, when clinical signs or necropsy result could be associated to the H9N2 challenge. Clinical signs Score
  • Ocular Mild conjunctivitis with oedema of the eyelid and excessive weeping+ 2 periorbital oedema
  • a daily clinical score was calculated by adding the scores associated with respiratory and ocular symptoms (see Table 2) from D22 to D31.
  • a global clinical score was calculated by adding all of the daily clinical scores.
  • Each group was compared to the control group on the criterion “Global clinical score” using a Mann- Whitney -Wilcoxon’s test. On D31, all surviving animals were humanely euthanized.
  • Virus- excreting groups were compared to the control group on the criterion “AUC” using a Student’s T test (assuming the homogeneity of the variances (Fisher- Snedecor’s test).
  • AUC Average-to-Unwinding Average

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Virology (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Medicinal Chemistry (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Engineering & Computer Science (AREA)
  • Veterinary Medicine (AREA)
  • General Engineering & Computer Science (AREA)
  • Biophysics (AREA)
  • Wood Science & Technology (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biomedical Technology (AREA)
  • Zoology (AREA)
  • Biotechnology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Communicable Diseases (AREA)
  • Oncology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Pulmonology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Plant Pathology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Mycology (AREA)
  • Epidemiology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

La présente invention concerne des vecteurs viraux recombinants du virus de la maladie de Carré (CDV) comprenant un polynucléotide hétérologue codant pour un antigène de la grippe aviaire. Les vecteurs viraux recombinants sont appropriés pour être utilisés dans des compositions immunogènes et des vaccins, et peuvent assurer une protection contre la grippe aviaire et d'autres pathogènes lorsqu'ils sont administrés à un animal.
PCT/IB2024/058182 2023-08-25 2024-08-22 Vecteurs de cdv via et leurs utilisations Pending WO2025046414A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202363578893P 2023-08-25 2023-08-25
US63/578,893 2023-08-25

Publications (1)

Publication Number Publication Date
WO2025046414A1 true WO2025046414A1 (fr) 2025-03-06

Family

ID=92842982

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/IB2024/058182 Pending WO2025046414A1 (fr) 2023-08-25 2024-08-22 Vecteurs de cdv via et leurs utilisations

Country Status (1)

Country Link
WO (1) WO2025046414A1 (fr)

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007104782A1 (fr) * 2006-03-15 2007-09-20 Intervet International B.V. Virus recombinant de la maladie de newcastle exprimant l'hémagglutinine h5 du virus de la grippe aviaire
WO2019057859A1 (fr) 2017-09-23 2019-03-28 Boehringer Ingelheim Vetmedica Gmbh Système d'expression de paramyxoviridae

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007104782A1 (fr) * 2006-03-15 2007-09-20 Intervet International B.V. Virus recombinant de la maladie de newcastle exprimant l'hémagglutinine h5 du virus de la grippe aviaire
WO2019057859A1 (fr) 2017-09-23 2019-03-28 Boehringer Ingelheim Vetmedica Gmbh Système d'expression de paramyxoviridae

Non-Patent Citations (12)

* Cited by examiner, † Cited by third party
Title
"Biocomputing: Informatics and Genome Projects", 1993, ACADEMIC PRESS
"Computer Analysis of Sequence Data", 1994, HUMANA PRESS
"Genbank", Database accession no. ACY80655.1
"Sequence Analysis in Molecular Biology", 1987, ACADEMIC PRESS
"Sequence Analysis Primer, Gribskov", 1991, STOCKTON PRESS
CARILLO, H.LIPMAN, D.: "Computational Molecular Biology", vol. 48, 1988, OXFORD UNIVERSITY PRESS, pages: 1073
DATABASE EMBL 1 January 2010 (2010-01-01), KIM L. M.: "Influenza A virus (A/avian/Saudi Arabia/910135/2006(H9N2)) hemagglutinin", XP093138362 *
DE JONG ET AL., J CLIN VIROL., vol. 35, no. 1, 2006, pages 2 - 13
KAUFMANN, L ET AL., JOVE, vol. 130, 2017, pages e55833
PARKS CHRISTOPHER L ET AL: "Expression of a foreign gene by recombinant canine distemper virus recovered from cloned DNAs", VIRUS RESEARCH, AMSTERDAM, NL, vol. 83, no. 1-2, 1 January 2002 (2002-01-01), pages 131 - 147, XP002205764, ISSN: 0168-1702, DOI: 10.1016/S0168-1702(01)00430-0 *
WANG ET AL., VACCINE, vol. 30, 2012, pages 5067 - 5072
ZHAO ET AL., VIRUSES, vol. 12, no. 3, 2020, pages 339

Similar Documents

Publication Publication Date Title
Paillot et al. Vaccination against equine influenza: quid novi?
ES2399845T3 (es) Herpesvirus de pavo vectorial recombinante que contiene genes de la influencia aviar
EP2629794B1 (fr) Vaccin basé sur le vecteur de l'herpèsvirus de la dinde contre la grippe aviaire chez la volaille
PL188409B1 (pl) Szczepionki do leczenia koni, zastosowanie plazmidu do wytwarzania szczepionki i zestaw do szczepienia koni
KR20000065258A (ko) 조류 폴리뉴클레오티드 백신 제제
US11730805B2 (en) Paramyxoviridae expression system
TW201701900A (zh) 二價豬流感病毒疫苗
US11744888B2 (en) Method of treating or preventing clinical signs caused by infectious bronchitis virus with 4/91 IBV vaccine having heterologous spike protein
WO2020058341A1 (fr) Vaccin à vecteur par voie intranasale contre la diarrhée épidémique porcine
CN105251000A (zh) 猪伪狂犬病病毒疫苗组合物及其制备方法和应用
JP2023545524A (ja) 組換えｈｖｔ及びその使用
US11696947B2 (en) H52 IBV vaccine with heterologous spike protein
AU2008364200B2 (en) Recombinant inactivated viral vector vaccine
CN119040279A (zh) 基于水疱性口炎病毒载体的狂犬病毒疫苗
US9370563B2 (en) Newcastle disease virus and the use thereof as a vaccine
WO2025046414A1 (fr) Vecteurs de cdv via et leurs utilisations
KR20210035676A (ko) 뉴캐슬병 바이러스 벡터를 이용한 고역가 조류메타뉴모바이러스 백신
US20090258035A1 (en) Avian virus vaccines and uses thereof
WO2025046413A1 (fr) Vecteurs de via de vmn et leurs utilisations
US20250064917A1 (en) Hvt aiv vectors and uses thereof
Guo et al. Recombinant avian metapneumovirus subtype C expressing HA protein of H9N2 avian influenza virus are stable and induce protection
KR20210035675A (ko) 뉴캐슬병 바이러스 벡터를 이용한 고역가 조류메타뉴모바이러스 백신
US20250064920A1 (en) Ibv vaccine with heterologous dmv/1639 spike protein
HK1164691A (en) Recombinant inactivated viral vector vaccine
JP2015120709A (ja) 遺伝子組み換え不活性化ウィルスベクターワクチン

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 24773557

Country of ref document: EP

Kind code of ref document: A1