Chemistry: molecular biology and microbiology – Vector – per se
Reexamination Certificate
2000-07-05
2003-10-14
Housel, James (Department: 1648)
Chemistry: molecular biology and microbiology
Vector, per se
C435S320100, C435S235100, C424S199100, C424S204100, C424S201100, C424S202100, C424S229100, C424S810000
Reexamination Certificate
active
06632664
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a recombinant herpesvirus of turkeys (hereinafter referred to as BVT) and a recombinant Marek's disease virus (hereinafter referred to as MDV), in which an foreign gene has been integrated into a non-essential region of the genome of HVT or MDV, and vaccines employing said recombinant.
BACKGROUND ART
Conventionally known virus vector vaccines that are prepared using gene recombinant technology include vaccines that employ a virus of the genus poxvirus as a vector (Ogawa R. et al., Vaccine, 8:486-490 (1990)), vaccines that employ adenovirus as a vector (HSU, K. H. et al., Vaccine, 12;607-612 (1994)), vaccines that employ baculovirus as a vector, as well as vaccines that employ a virus of the genus herpesvirus as a vector (Shin, M. -F. et al., Proc. Natl. Acad. Sci. U.S.A., 81:5867-5870 (1984)). Among them, recombinant vector vaccines based on the genus herpesvirus are under intensive study in recent years.
As virus vectors that permit the expression of a gene for a foreign antigen, there are known human herpesvirus (HSV), Aujeszky's disease virus (pseudorabies virus; PRV) (Van Zijl M. et al., J. Virol., 65:2761-2765 (1991)), herpesvirus of turkey (HVT) (Morgan R. W. et al., Avian Dis. 36:858-870 (1992)), Marek's disease virus (MDV), and the like. Since, HVT virus and vaccine strain MDV, among them, have high safety in poultry that are the subject of vaccination and have good vaccine characteristics, they are attracting attention as vector viruses for avians.
Unlike poxvirus that has a mode of infection in which the virus is once released into the blood from the infected cell and then it infects another call during its infection from the infected cell to another cell, HVT and MDV establish infection via a cell-cell interaction to an adjacent cell. Thus, they are relatively free from the influences of HVT- or MDV-specific antibodies present in the circulating blood.
Conventionally, problems have been recognized in that the efficacy of live virus vaccines is attenuated by the presence of maternal antibody from the mother bird, resulting in the failure of exhibiting their full effects.
In recent years, methods of inoculating vaccines into developing chicken eggs have been developed as one of the methods of vaccination to chickens, and the usefulness of HVT or MDV as a vaccine is gaining recognition.
However, conventionally known genetic regions so far reported for construction of recombinant BVT or MDV for foreign antigen were only genetic regions that are considered non-essential for survival of HVT such as the TK region (Ross L. et al., 16th International Herpes virus Workshop (1991)), the US10region (Sakaguchi M. et al., Vaccine, 12:953-957 (1994)), and the US2 region (Sondermeijer, P. J. et al., Vaccine, 11:349-358 (1993)). Such integration into non-essential regions can attenuate the antigenicity of HVT or MDV since it causes the expression of a foreign gene in stead of a gene that, though non-essential, should naturally be expressed in HVT i.e. a gene that will make an antigenic determinant. In addition, the possibility cannot be ruled out that the genetic machineries (enhancers, promoters, terminators, etc.) involved in the transcription or translation of the open reading frame for the inserted region may adversely affect the expression of the inserted gene.
In fact, there are reports that the deletion of genetic regions encoding proteins that are considered to be non-essential or the integration therein of foreign genes resulted in the modification of viral morphology or the reduction in antigenicity. Furthermore, the integration of foreign genes has been used, in some cases, as a method of preparing attenuated vaccines.
Another report demonstrates that the insertion of a foreign gene into the TK region and the expression thereof resulted in the decreased antigenicity of the expressed gene (Ross L. et al., J. Gen. Virol., 74:371-377 (1993)). This method also has a number of problems as a vaccine in that many antigen genes cannot be inserted since the length of antigen genes that can be inserted into specific ORFs is limited.
As a result of intensive study to solve the above problems, the inventors of the present invention have found that there are the gene insertion regions of HVT or MDV into which a variety of genes for foreign antigen can be inserted and the antigen protein can be stably expressed, i.e. the untranslated region of HVT or MDV mentioned above, that genes of various foreign antigens can be inserted therein, and that by preparing recombinant HVT or MDV into which these genes of foreign antigen have been inserted and then infecting these recombinant viruses to the hosts, adequate vaccination effect cat be conferred to the host, and thereby have completed the present invention.
DISCLOSURE OF THE INVENTION
As a result of intensive study to solve the above problems, the inventors of the present invention have prepared a recombinant virus in which a foreign gene has been inserted into a specific site in a untranslated region of a virus DNA belonging to the avian infectious herpesvirus and found that said recombinant virus can be used as a vaccine, and thereby have completed the present invention.
Thus, the present invention relates to avian infectious recombinant herpesviruses in which a foreign gene has been inserted into a genetic region that is an untranslated region in the genome. Preferably, the virus mentioned above is herpesvirus of turkeys (HVT) and Marek's disease virus (MDV).
The above untranslated region is preferably an untranslated region present in the open reading frame of herpesvirus of turkeys or the open reading frame of Marek's disease virus, each corresponding to the open reading frame of human herpes simplex virus. A specifically preferred insertion site for a foreign gene is at least one insertion site selected from the group consisting of sites in (1) between UL44 and UL45, (2) between UL45 and UL46, (3) between UL41 and UL42, (4) between UL40 and UL41, (5) a region located downstream of the gB gene, (6) between UL53 and UL54, and (7) between UL36 and UL37.
The above foreign gene is preferably a gene derived from a pathogen of avian infectious diseases, and most preferably an antigen gene derived from a pathogen selected from the group consisting of viruses, bacteria, fungi, and protozoa. Furthermore, the above foreign gene is preferably a gene derived from a pathogen selected from the group consisting of Newcastle disease virus (NDV), Gumboro disease virus (infectious bursal disease virus: IBDV), infectious laryngotracheitis virus (ILTV), infectious bronchitis virus (IBV), mycoplasma (MG), and coccidia.
The present invention also relates to a chicken vaccine comprising the above recombinant virus as an active ingredient.
EBODIMENT FOR CARRYING OUT THE INVENTION
The present invention will now be explained hereinbelow in more details.
Viruses for Use in the Present Invention
Of the viruses that infect avians, viruses for use in the present invention are preferably those that belong to the genus herpesvirus (avian infectious herpesvirus), because viruses that belong to the genus herpesvirus have a property of permanently surviving in the body of an infected animal in the state of latent infection or persistent infection.
Of the avian infectious herpesviruses, specifically herpesvirus of turkey or Marek's disease virus (MDV) is preferred. Their effectiveness as a vaccine can be expected because the above viruses have a long infection period and are expected to confer to avians vaccination effects for a long period of time.
HVT or MDV
HVT or MDV for use in the present invention may include, but not limited to, naturally occurring ones or those available from ATCC etc. with or without charge.
Preferred examples of HVT include those that belong to the gamma herpesvirus subfamily, that are originally nonpathogenic, and that are noncarcinogenic and are used as a vaccine for poultry. Specifically, there can be mentioned FC126 (ATCC VR-584B), PB-THV1, H-2, YT-7, WTHV
Okuda Takashi
Saitoh Shuji
Armstrong Westerman & Hattori, LLP
Housel James
Li Bao Qun
Nippon Zeon Co. Ltd.
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