Chemistry: molecular biology and microbiology – Virus or bacteriophage – except for viral vector or...
Reexamination Certificate
1997-04-25
2001-11-20
Mosher, Mary E. (Department: 1648)
Chemistry: molecular biology and microbiology
Virus or bacteriophage, except for viral vector or...
C435S236000, C424S199100, C424S229100, C424S231100, C424S093200, C424S205100
Reexamination Certificate
active
06319703
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to mutant viruses that can be used as recombinant virus vectors. The invention also relates to mutant virus vectors that can be used for the delivery to cells of nucleotide sequence(s) encoding polypeptide(s). The invention also relates to cells infected by such mutant viruses and to materials and methods for delivering nucleotide sequence(s) encoding polypeptide(s) to host cells ex vivo or to treated subjects in vivo such as human patients by use of a recombinant virus vector based on such mutant herpesvirus.
BACKGROUND OF THE INVENTION
Many disorders which manifest in symptoms such as respiratory distress, growth abnormalities, muscular insufficiency, and/or mental retardation result from the inheritance of genetic material which is defective in that a gene coding for the synthesis of a protein is either completely or partially absent or of an incorrect coding sequence. Thus the defect results in the disruption of the normal activities of cells which are dependent upon the normal protein for correct functioning.
The affected protein can be for example an enzyme, a storage protein, a transport protein, a hormone, a recognition protein such as a cell surface receptor or a cell pore protein.
Typical examples of disorders of this type are cystic fibrosis, Tay-Sachs disease. B-thalassaemia and Hurler disease.
If the particular genetic defect can be identified, there is the possibility that it can be correctable by providing the patient with a correct version of the gene.
Herpes simplex virus (HSV) has been considered to have potential as a vector for such gene therapy, because it has the potential to carry large inserts of foreign DNA and because it is capable of existing as a latent, quiescent genetic element in cells (particularly neurones) for the lifetime of the host. Nevertheless, a disadvantage of wild-type HSV is that it is a lytic virus whose growth results in cell damage or cell death. Therefore, the use of an unmodified form of an HSV virus vector is unacceptable, and even some modified forms can carry significant risks.
The construction and characterization of a HSV-1 mutant (in1814) unable to transinduce immediate early gene expression, and essentially avirulent when injected into mice is described by C I Ace et al. J Virol 63(5) 1989 pp 2260-2269, and specification WO 91/02788 (C M Preston & C I Ace: University of Glasgow) describes HSV1 mutants including in1814 capable of establishing latent infection in a neuronal host cell and of causing expression of an inserted therapeutic gene.
In mutants such as in1814 the gene for VP16 is modified: VP16 is a structural component of the virus particle which acts as a transcriptional activator of immediate-early genes of the infecting genome. This mutant enters cells normally. although it has a reduced ability to enter the productive lytic cycle.
SUMMARY AND DESCRIPTION OF THE INVENTION
The present inventor considers that although the mutants of the prior art include an essentially avirulent (in mice) mutant such as in1814, it remains desirable to provide a further genetic defect in a virus to be used as a vector for gene therapy and for other purposes.
The present invention provides mutant herpesvirus which (a) carries a mutation such that it has a reduced ability to enter the productive lytic cycle in an infected cell or cause lysis of an infected cell, and (b) has a genome which is defective in respect of a gene essential for the production of infectious virus; the invention also provides methods for its production and its uses. The mutant herpesvirus genome thus carries (a) a mutation such that the mutant virus has a reduced ability in comparison with a parent type to cause lysis of an infected cell, and (b) an inactivating mutation in a gene essential for the production of infectious virus.
The further genetic defect in the form of inactivating mutation (b) brings a safety advantage in that this mutation can be such that it is not susceptible of complementation within the host cell, and is practically free from risk of reversion. Such a further genetic defect is known per se, being of the kind described in specification WO 92/05263 (S C Inglis et al: Immunology Ltd) and corresponding GB 2 263 480 (Cantab Pharmaceuticals Ltd) entitled “Viral Defective Vaccine Produced by Transcomplementing Cell Line” which describe a mutant virus for use as a vaccine, having a genome which is defective in respect of a gene essential for the production of normal infectious virus. Mutant virus of this kind can be propagated on a recombinant complementing cell which provides the virus with the product of the deleted gene, thus making it possible to grow the virus in tissue culture.
A mutant herpesvirus of the present invention can be used for example as a vector for a gene, heterologous to the virus itself, to be administered to and expressed in cells of a treated subject by way of gene therapy.
Embodiments of the invention can in this connection have an advantage in that they can combine an increased margin of safety over known herpesvirus vectors in respect of the incidence of cytopathic effects and/or of the risk of reversion to virulence, along with useful persistence of expression. in cells of the treated subject, of the gene carried by the vector.
The mutant herpesvirus can also be used as a vector for a gene encoding an antigen against which an immune response is desired in the treated subject.
The mutant herpesvirus can be a mutant of any herpesvirus e.g. a mutant of a mammalian herpesvirus, e.g. a human herpesvirus. Examples of herpesviruses of which mutants are provided according to embodiments of the invention include herpes simplex virus of type 1 (HSV-1) or type 2 (HSV-2). a human or animal cytomegalovirus (CMV). e.g. human cytomegalovirus (HCMV), varicella zoster virus (VZV), Epstein-Barr virus (EBV), and human herpesvirus 6 and 7. Animal viruses of which mutants are provided according to embodiments of the invention include pseudorabies virus (PRV), equine and bovine herpesvirus including EHV and BHV types such as IBRV, and Marek's disease virus (MDV) and related viruses. The nomenclature of the genes of herpesviruses and their many corresponding homologues is diverse and where the context admits. mention of a gene in connection with a herpesvirus includes reference, in connection with other herpesviruses possessing a homologue of that gene, to the corresponding homologue.
A mutant herpesvirus of the invention carries at least one mutation that reduces its ability to cause lysis of an infected cell. Where for example the herpes virus is HSV-1, the defect in the genome which reduces the mutant's ability to enter a productive lytic cycle as compared to wild-type virus can for example comprise a mutation in the VP16 gene.
A preferred example of such a mutation is a mutation in the gene corresponding to VP16 (otherwise designated Vmw65 or alpha-TIF) in herpes simplex virus type 1, which is effective to reduce (by comparison with a parent type), or substantially remove the transinducing properties of the protein encoded by that gene, especially while retaining its structural role. A particular example of such a mutation is the in1814 mutation in HSV1. Further examples of such mutations include VP16 mutations such as a transition or transversion alteration of 1 to 72 base pairs, an oligonucleotide insert of 3 to 72 base pairs, or a deletion of 3 to 72 base pairs, e.g. at a position within the region corresponding to aminoacids 289 and 412 of the HSV1 VP16 protein, as described in specification WO 91/02788, hereby incorporated by reference, or other mutations in the gene corresponding to VP16 that allow growth of the mutant virus in cell culture in the presence of hexamethylene bisacetamide (HMBA).
Thus the present invention also provides a mutant herpesvirus e.g.
HSV-1 which comprises a genome which (i) has a defect such that in comparison to the wild-type form of the herpesvirus, or other corresponding parent strain in relation to the mutation, the mutant has a reduced a
Klarquist & Sparkman, LLP
Mosher Mary E.
LandOfFree
Recombinant virus vectors does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Recombinant virus vectors, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Recombinant virus vectors will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2569046