Drug – bio-affecting and body treating compositions – Antigen – epitope – or other immunospecific immunoeffector – Virus or component thereof
Reexamination Certificate
1999-02-25
2001-06-05
Mosher, Mary E. (Department: 1648)
Drug, bio-affecting and body treating compositions
Antigen, epitope, or other immunospecific immunoeffector
Virus or component thereof
C424S816000, C435S235100
Reexamination Certificate
active
06241991
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a new aviadenovirus.
BACKGROUND OF THE INVENTION
Aviadenoviruses can be subdivided into three groups:
Group I adenoviruses, a subclass of which comprises the fowl adenoviruses (FAV) having a genomic DNA length of ±43.8 kb and two fibres protruding from one pentonbase. The Group I viruses share a common antigen (‘group antigen’) which distinguishes them from Group II and Group III viruses (ref. 1,9).
Group I adenoviruses can be isolated from different avian species and have been classified as Fowl adenoviruses (FAV), Goose adenoviruses (GAV), Duck adenoviruses (DAV) and Turkey adenoviruses (TAV). The length of the two fibres of FAVs is different for each of the 12 serotypes. In addition to serological typic, the viruses can be characterized by analysis of their DNA. Based on BamHI and HindIII cleavage of the DNA of FAV serotypes 1-11, five subgroups are recognized (ref. 1,4,6,9).
Group II adenoviruses, having Group II specific antigen, comprise a small number of viruses, such as the turkey Haemorrhagic Enteritis virus. These viruses have a genomic DNA length of ±25.5 kb and sport one fibre per penton base.
Group III adenoviruses, of which the Egg Dropping Syndrome causing virus (EDS) is the only serotype, have a genomic DNA length of 33.2 kb and a single fibre per penton base. EDS is serologically not directly related to Group I viruses (ref. 11).
SUMMARY OF THE INVENTION
The aviadenovirus according to the present invention comprises group I-specific antigen and cannot be neutralized by antiserum against any of the FAV serotypes 1-12. Group I specific antigen has not been characterized, but an antiserum obtained using viruses from one of the FAVs is capable of recognizing the other FAVs. This immunoreaction can be detected by using immunofluorescence methods well-known in the art, such as described in ref. 13. In the present invention, and in accordance with ref. 7, the term neutralisation is defined as the ability of an antiserum raised against a particular virus to inhibit virus propagation of another virus.
The new aviadenovirus was isolated from the liver of a pigeon. It is well known that aviadenoviruses can pass species barriers. As yet, the new virus has not been isolated from other avians, which may indicate that it is a primary pigeon adenovirus. Hence, it has been given the name Pigeon AdenoVirus (PiAV).
DETAILED DESCRIPTION OF THE INVENTION
DNA analysis of the PiAV using restriction enzymes BamHI and HindIII result in patterns that do not correspond to any of the five DNA types A-E recognized so far. In fact it did not correspond to the pattern of all known Group I adenoviruses, including GAV and TAV. In addition, it has been found that antiserum against any of the FAV serotypes 1-12 could not neutralise the virus according to the present invention, further proving the unique nature of PiAV.
A sample of the aviadenovirus isolate PiAV 197/5 was deposited on Mar. 3, 1998 with CNCM of the Institute Pasteur (25, Rue du Docteur Roux, F-75724 Paris CEDEX 15, France) under accession number I-1988.
The invention also relates to an antibody or antiserum immunoreactive with a fibre of the aviadenovirus according to the invention.
Preferably, the aviadenovirus according to the invention is a PiAV which in a cross-neutralisation test has a homologous:heterologous titre ratio of 8 or more in both directions with PiAV isolate as deposited under I-1988. More preferably the PiAV according to the invention has a homologous:heterologous titre ratio of 16 or more, in particular of more than 16.
Such an antibody is useful for the immunological detection of PiAV in avians suspected of contracting it.
The invention also relates to a fibre or a synthetic or proteolytically obtained fragment thereof of the aviadenovirus according to the invention.
These fibres or fragments thereof, comprising an antigenic determinant, are useful for generation of monoclonal antibodies, and for the development of a vaccine.
Furthermore the invention relates to a nucleic acid sequence encoding an amino acid sequence, said amino acid sequence comprising an immunogenic determinant or a functional variant thereof of a fibre of the aviadenovirus according to the invention.
Such a nucleic acid sequence is useful for transformation of a host cell or a host virus, which may be used to develop a vaccine.
Accordingly, the present invention relates to a host cell transformed with a nucleic acid sequence according to the invention.
Such a host cell may be used for the preparation of a vaccine, or for the generation of a large amount of amino sequences comprising an immunogenic determinant, said amino sequences being useful for vaccination purposes and obtaining monoclonal and polyclonal antibodies.
Finally, the present invention relates to a vaccine against PiAV related diseases, comprising an attenuated virus, a chemically or physically inactivated virus, a fibre or a fragment thereof, a host cell transformed with the nucleic acid sequence together with a pharmaceutical carrier.
Aviadenoviruses according to the present invention can be obtained by conventional methods. Briefly, a susceptible substrate is inoculated with PiAV and propagated until the virus replicated to a desired titre after which PiAV containing material is harvested.
Every substrate which is able to support the replication of PiAV can be used in the present invention, for example chicken embryo liver cells.
The vaccine according to the invention containing live attenuated virus can be prepared and distributed in the form of a suspension or in a lyophilized form and additionally contains a pharmaceutically acceptable carrier or diluent customary used for such compositions. Carriers include stabilizers, preservatives and buffers. Suitable stabilizers are, for example SPGA, carbohydrates (such as sorbitol, mannitol, starch, sucrose, dextron, glutamate or glucose), proteins (such as dried milk serum, albumin or casein) or degradation products thereof. Suitable buffers are for example alkali metal phosphates. Suitable preservatives are thimerosal, merthiolate and gentamicin. Diluents include water, aqueous buffers (such as buffered saline) and polyols (such as glycerol).
If desired, the live vaccines according to the invention may contain an adjuvant.
The aim of inactivation of PIAV harvested after the propagation step is to eliminate reproduction of the viruses. In general, this can be achieved by chemical or physical means. Chemical inactivation can be effected by treating the viruses with, for example, enzymes, formaldehyde, &bgr;-propiolactone, ethyleneimine or a derivative thereof. If necessary, the inactivating compound is neutralized after-wards. Material inactivated with formaldehyde can, for example, be neutralized with thiosulphate or sodium metabisulphite. Physical inactivation can preferably be carried out by subjecting the viruses to energy-rich radiation, such as UV light, X-radiation or &ggr;-radiation. If desired, the pH can be brought back to a value of about 7 after treatment.
Preferably, an inactivated vaccine according to the invention comprises one or more compounds with adjuvant activity. Suitable compounds or compositions for this purpose include aluminium hydroxide, -phosphate or -oxide, oil-in-water or water-in-oil emulsion based on, for example a mineral oil, such as Bayol™ or Marcol 52™ or a vegetable oil such as vitamin E acetate and saponins.
To obtain attenuated PiAV, the virus can be used to infect embryonated eggs and subsequently propagation and passaging of the virus in embryonated eggs by methods known in the art for this purpose.
The vaccine according to the invention comprises an effective dosage of aviadenoviruses according to the present invention as the active component, i.e. an amount of immunizing PiAV material that will induce immunity in the vaccinated birds against challenge by a virulent PiAV. Immunity is defined herein as the induction of a significant higher level of protection in a population of birds after vaccination compared to an unva
Hess Michael Bernhard
Paul Guntram
Akzo Nobel N.V.
Blackstone William M.
Mosher Mary E.
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