+

WO1998032869A1 - Vecteurs d'expression et procedes d'expression in vivo de polypeptides therapeutiques - Google Patents

Vecteurs d'expression et procedes d'expression in vivo de polypeptides therapeutiques Download PDF

Info

Publication number
WO1998032869A1
WO1998032869A1 PCT/DK1998/000037 DK9800037W WO9832869A1 WO 1998032869 A1 WO1998032869 A1 WO 1998032869A1 DK 9800037 W DK9800037 W DK 9800037W WO 9832869 A1 WO9832869 A1 WO 9832869A1
Authority
WO
WIPO (PCT)
Prior art keywords
disease
expression vector
therapeutically active
recombinant
active polypeptide
Prior art date
Application number
PCT/DK1998/000037
Other languages
English (en)
Inventor
Teit E. Johansen
Original Assignee
Neurosearch A/S
Bavarian Nordic Research Institute A/S
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 Neurosearch A/S, Bavarian Nordic Research Institute A/S filed Critical Neurosearch A/S
Priority to EP98900847A priority Critical patent/EP0961830A1/fr
Publication of WO1998032869A1 publication Critical patent/WO1998032869A1/fr

Links

Classifications

    • 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
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • 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
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/13011—Gammaretrovirus, e.g. murine leukeamia virus
    • C12N2740/13041—Use of virus, viral particle or viral elements as a vector
    • C12N2740/13043—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/13011—Gammaretrovirus, e.g. murine leukeamia virus
    • C12N2740/13041—Use of virus, viral particle or viral elements as a vector
    • C12N2740/13045—Special targeting system for viral vectors

Definitions

  • the present invention relates to recombinant expression vectors carrying a gene encoding a therapeutically active polypeptide, which gene is under transcriptional control of a ubiquitin promoter.
  • the invention also relates to the use of a ubiquitin promoter to direct in vivo expression of therapeutic genes after transfer of such genes to the central nervous system.
  • Therapeutically active polypeptides may be delivered to the central nervous system (CNS) by implantation of polypeptide producing cells, or by injection of viral vectors capable of infecting brain cells and directing the expression of a therapeutically active polypeptide encoded by the viral vector in the brain cells.
  • CNS central nervous system
  • primary and immortalised cells have been successfully used for gene transfer applications.
  • primary cells of neuronal origin e.g. glia cells and astrocytes
  • non-neuronal origin e.g. fibroblasts, myoblasts and hepatocytes.
  • Such cells may not survive for longer periods in the CNS unless they are immortalised.
  • Intracerebral grafting of foetal tissue for the treatment of neurological disorders have also been investigated.
  • stem cells may be used.
  • cerebral endothelial cell Another cell type which has been proposed as gene transfer vehicle is the cerebral endothelial cell.
  • the direct implantation of immortalised and genetically transformed cerebral endothelial cells into various parts of the brain has been disclosed. It has also been suggested to deliver therapeutic agents by infecting endothelial cells of blood vessels located in the brain with a viral vector, as a result of intravascular administration of the vector to the host near the site of infection (WO 96/22112).
  • a broad range of viral vectors such as including adenovirus vectors (WO 95/26408), adeno-associated virus vectors (WO 95/34670), herpes virus vectors (Glorioso et al. Sem. Virol. 1992 3 265-276), vaccinia virus vectors, and retroviral vectors, including systems based on HIV, has been suggested as delivery vehicles for therapeutic genes to the CNS.
  • Such vectors can be administered to the CNS by the intravenous and the intracranial route, and by administration to the cerebrospinal fluid.
  • WO 95/09654 discloses a method for the treatment of adverse conditions of the CNS by administration of producer cells containing a retroviral vector to the cerebrospinal fluid.
  • the producer cells produce retroviral particles which is capable of transducing cells present in the nervous system.
  • Intracerebral implantation of encapsulated cells producing a therapeutic peptide has also been suggested.
  • Ubiquitin proteins are "house hold proteins", essential for the catabolism of all cellular proteins. The structure of ubiquitin proteins is highly conserved during evolution.
  • the ubiquitin promoter is a constitutive promoter which controls the expression of ubiquitin proteins. A number of transfection experiments have shown that this promoter is as strong as, or even stronger than the commonly used viral promoters. Furthermore, heterologous genes under the control of the ubiquitin promoter are actively transcribed in every cell line that has been transfected with this promoter (more than 20, including several cell lines of neuronal origin). In transgenic mice, the human ubiquitin C promoter (UbC) have been shown to direct in vivo expression of heterologous genes in several types of tissues, including brain-tissue (Nucleic Acids Research 1996 24 (9) 1787-1788).
  • the therapeutic gene In order to obtain efficient expression in the CNS, the therapeutic gene must be put under transcriptional control of a promoter which is active in the CNS.
  • viral promoters are down-regulated in vivo in mammalian cells. If efficient transfer of a therapeutic polypeptide to the CNS is to be obtained with a viral vector, it is should be replaced the with a promoter which will provide stable and efficient expression.
  • the ubiquitin promoter has proven to direct stable high level expression of heterologous genes in in vitro experiments. However, there is no indication of its efficiency in vivo.
  • the present invention is directed to the use of an ubiquitin promoter for stable and efficient expression of genes encoding therapeutically active polypeptides in the CNS.
  • An ubiquitin promoter of present invention includes the human ubiquitin promoter, which is indistinguishable from the endogenous ubiquitin promoter and should therefore not be susceptible to the in vivo down-regulation observed for viral promoters.
  • the present invention is directed towards a recombinant expression vector comprising a gene encoding a therapeutically active polypeptide, which gene is under transcriptional control of an ubiquitin promoter.
  • the invention provides an eukaryotic cell, transfected with the eukaryotic expression vector of the invention.
  • the invention provides a packaging cell line comprising the retroviral expression vector of the invention and one or more nucleotide constructs encoding the proteins required for the genome of the retroviral vector to be packaged.
  • the invention provides a method of producing a retroviral particle by culturing the packaging cell line of the invention, and a retroviral particle obtained according to this method.
  • the invention provides an eukaryotic cell, e.g. an immortalised cerebral endothelial cell line, infected by a retroviral particle obtained by the method of this invention.
  • the invention is directed towards the use of the recombinant expression vector of the invention for the manufacture of a pharmaceutical composition, useful for the treatment of a neurological disease or disorder.
  • the invention provides a method for the treatment of a living body suffering from a neurological disease or disorder which is responsive to a therapeutically active polypeptide, which method comprises administering to the living body, a retroviral particle obtained by the method of the invention, encoding the therapeutically active polypeptide; or implanting into the living body, the cells of the invention, producing the therapeutically active polypeptide.
  • Fig. 1 shows a graphic illustration of a UbC promoter containing plasmid based expression vector, pUbilz, obtained according to Example 2.
  • the present invention resides in the use of a ubiquitin promoter to direct in vivo expression of therapeutic genes after transfer of such genes to the central nervous system.
  • the invention provides a recombinant expression vector comprising a gene encoding a therapeutically active polypeptide, which gene is under transcriptional control of an ubiquitin promoter.
  • the recombinant expression vector of the invention may be any vector suitable for transferring a gene under transcriptional control of a ubiquitin promoter to mammalian cells.
  • the recombinant expression vector is an eukaryotic expression vector or a recombinant viral expression vector.
  • Genes can be transferred into cells using a variety of means including calcium phosphate precipitation [Graham et al., Virol. 1973 52 456-467; Wigler et al., Cell 1979 777-785], electroporation [Neumann et al., EMBO J. 1982 841-845], microinjection [Graessmann et al., Meth.
  • Viral vectors provide an efficient means of transferring genes into cells both in vivo and in vitro.
  • the efficiency of viral gene-transfer is due to the fact that transfer of DNA is an essential part of the natural life cycle of viruses and that DNA transfer is a receptor-mediated process.
  • viral systems including retrovirus, adenovirus, adeno-associated virus, vaccinia virus and herpes virus have been developed as in vivo therapeutic gene transfer vectors for gene therapy of CNS disorders.
  • Viral vectors have also been used to transform various forms of cells such as astrocytes, fibroblast cells and cerebral endothelial cells which is thereafter implanted into the CNS. Further WO 96/06942 describe genetically altered T-cells which enter the CNS and may be used as gene transfer vehicles.
  • the recombinant viral expression vector of the invention may be any viral expression vector suited for in vivo transfer and expression of genes.
  • Preferred viral expression vectors include retroviral vectors, recombinant adenovirus vectors, recombinant adeno-associated virus vectors, vaccinia virus vectors and recombinant herpes virus vectors.
  • Recombinant retroviral gene delivery methods are the most extensively used due in part to: (1) the efficient entry of genetic material (the vector genome) into replicating cells; (2) an active, efficient process of entry into the target cell nucleus; (3) relatively high levels of gene expression; (4) the potential to target particular cellular subtypes through control of the vector target cell binding and the tissue specific control of gene expression; (5) a general lack of pre-existing host immunity; and substantial knowledge and clinical experience which has been gained with such vectors.
  • the recombinant vector carrying a therapeutic gene under transcriptional control of an ubiquitin promoter is therefore a retroviral vector.
  • the retroviral genome consists of an RNA molecule with the structure R-U5- gag-pol-env-U3-R. During the process of reverse transcription, the U5 region is duplicated and placed at the right hand end of the generated DNA molecule, whilst the
  • U3 region is duplicated and placed at the left hand end of the generated DNA molecule.
  • the resulting terminal structure U3-R-U5 is called LTR (Long Terminal
  • the U3 region at the left hand end of the provirus harbours the promoter that is used to drive expression after infection has occurred. This promoter drives the synthesis of an RNA transcript initiating at the boundary between the left hand U3 and
  • the RNA is packaged into retroviral particles and transported into the target cell to be infected.
  • the RNA genome is reverse transcribed as described above.
  • Retroviral vector systems used for the generation of recombinant retroviral particles consist of two components.
  • the retroviral vector itself is a modified retrovirus (vector plasmid) in which the genes encoding for the viral proteins (gag, pol and/or env) have been replaced by therapeutic genes and/or marker genes to be transferred to the target cell. Since the replacement of the genes encoding for the viral proteins effectively cripples the virus it must be rescued by the second component in the system which provides the missing viral proteins to the modified retrovirus.
  • the second component is a cell line that produces large quantities of the viral proteins (gag, pol and/or env), however lacks the ability to produce replication competent virus.
  • This cell line is known as the packaging cell line and consists of a cell line transfected with one or more plasmids carrying the genes (gag, pol and/or env) enabling the modified retroviral vector to be packaged.
  • the vector plasmid is transfected into the packaging cell line.
  • the modified retroviral genome including the inserted therapeutic and marker genes is transcribed from the vector plasmid and packaged into modified retroviral particles (recombinant viral particles).
  • This recombinant virus is then used to infect target cells in vitro or in vivo and the vector genome and any carried marker or therapeutic genes become integrated into the target cell's DNA.
  • a cell infected with such a recombinant viral particle cannot produce new vector virus since no viral proteins are present in these cells.
  • the DNA of the vector carrying the therapeutic and marker genes is integrated in the cell's DNA as a provirus and can now be expressed in the infected cell.
  • WO-A1 -9607748 describes the principle and construction of a new type of retroviral vector.
  • the ProCon-vector plasmid the right-hand (3') U3 region is altered, but the normal left-hand (5') U3 structure is maintained; the vector can be normally transcribed into RNA utilising the normal retroviral promoter located within the left- hand (5') U3 region upon its introduction into packaging cells.
  • the generated RNA will only contain the altered right-hand (3') U3 structure.
  • this altered U3 structure will be present in both Long Terminal Repeat at either end of the retroviral structure.
  • any promoter including the human ubiquitin promoter, can be inserted and this promoter is then utilised exclusively in the target cell for expression of linked sequences encoding therapeutic polypeptides.
  • DNA segments homologous to one or more cellular sequences can also be inserted into the polylinker for the purposes of gene targeting, by homologous recombination.
  • the retroviral vectors of the invention need not be of the ProCon type, but can be any conventional retroviral vector carrying therapeutic genes under transcriptional control of the an ubiquitin promoter.
  • Such vectors include Self-lnactivating-Vectors (SIN) in which retroviral promoters are functionally inactivated in the target cell (WO-A 1-94/29437). Further modifications of these vectors include the insertion of promoter gene cassettes within the LTR region to create double copy vectors (WO-A1 -89/11539). In both of these vectors the heterologous promoters inserted either in the body of the vector, or in the LTR region are directly linked to the therapeutic gene.
  • the retroviral vectors of the invention are based preferably either on a BAG vector [Price, Turner J D, and Cepko C; Proc. Natl. Acad. Sci. USA 1987 84 156-160) or an LXSN vector [Miller A D, & Rossman G J; Biotechni ⁇ ues 1989 7 980-990].
  • Eukaryotic Expression Vectors include a BAG vector [Price, Turner J D, and Cepko C; Proc. Natl. Aca
  • the recombinant eukaryotic expression vector of the invention may be any eukaryotic expression vector suited for transferring a gene under transcriptional control of a ubiquitin promoter to mammalian cells.
  • Preferred eukaryotic expression vector of the invention are pTEJ-4, pTEJ-8, or pUbilZ.
  • the eukaryotic expression vector should contain sequences which facilitate the prokaryotic propagation along with two eukaryotic transcription units.
  • Prokaryotic sequences include a bacterial resistance gene under the transcriptional control of the prokaryotic EM7 promoter and a bacterial origin of DNA replication.
  • the eukaryotic transcription unit responsible for eukaryotic selection employ the SV 40 early promoter to drive the expression of a resistance gene and a polyadenylation signal.
  • the present expression vectors contain one of the following prokaryotic/eukaryotic selection markers: neomycin, hygromycin, pyromycin or zeocin resistance gene allowing the selection of bacterial clones under EM7 promoter and cellular clones under SV 40 early promoter.
  • the second eukaryotic transcription unit contain the UbC promoter, a polyadenylation signal, and finally a polylinker for insertion of nucleotide sequences encoding the protein in question (as an example see pUbilZ, of Example 2).
  • the ubiquitin promoter contemplated according to the present invention is an ubiquitin promoter derived from any convenient origin, e.g. derived from the 5' end
  • the ubiquitin promoter is of human origin, in particular the human ubiquitin C promoter (UbC).
  • the ubiquitin promoter of the invention is a human ubiquitin promoter having the sequence presented as SEQ ID NO: 1 in the attached sequence listing.
  • a therapeutically active polypeptide of the invention may be any peptide, polypeptide or protein that is capable of ameliorating neurological disorders.
  • the therapeutically active polypeptide of the invention is a Nerve Growth Factor (NGF); a Fibroblast Growth Factor (FGF), in particular an acidic or a basic Fibroblast Growth Factor (aFGF or bFGF); an Insulin-like Growth Factor, in particular IGF I or IGF II; an Endothelial Growth Factor (EGF), in particular a Vascular Endothelial Growth and Permeability Factor (VEGPF); a member of the Transforming Growth Factor (TGF) superfamily, including a Transforming Growth Factor- ⁇ and - ⁇ (TGF ⁇ and TGF ⁇ ); a Glial Derived Neurotrophic Factor (GDNF); a Ciliary Neurotrophic Factor (CNTF); a Brain Derived Neurotrophic Factor (BDNF); a Neurotrophin, in particular Neurotrophin 3, 4/5, 6 or 7; a Neuturin (NTN); a Percipin; a Tumor Necrosis Factor (TNF), in
  • the therapeutically active polypeptide of the invention may be employed in order to treat diseases or disorders in the brain and central nervous system.
  • diseases and disorders include ischemic strokes; angiogenesis; metabolic diseases of the brain; axonal injury; spinal cord injury; Alzheimer's disease; Amyothropic Lateral Sclerosis (ALS); Parkinson's disease; Huntington's disease; motor neuron diseases; central nervous system infections; epilepsy; post polio syndrome; mucopolysaccharidoses (MPS), in particular MPS types I to VII; lipidoses; in particular Gaucher's disease; Lesch-Nyhan syndrome; X-linked ADL; metachromatic leukodystrophy; Krabbe's disease; Charcot-Marie-Tooth disease; Fragile X; epilepsies; Down's syndrome; phenylketonuria; degenerative disorders; and mental disorders.
  • MPS mucopolysaccharidoses
  • the eukaryotic expression vectors and the recombinant retroviral vectors according to the invention may in addition to the therapeutic gene carry a gene encoding a marker.
  • the marker may in particular be proteins like ⁇ -galactosidase, neomycin, alcohol dehydrogenase, luciferace, puromycin, hypoxanthine phosphoribosyl transferase (HPRT), hygromycin, secreted alkaline phosphatase, or green or blue fluoroscent proteins (GFP).
  • the vector of the invention may be used to transduce cells to secrete the therapeutic polypeptide in vivo after being implanted into the CNS.
  • cells producing the therapeutically active polypeptide are generated by transfection with a plasmid vector
  • recipient cells e.g. immortalised neural stem cells, immortalised cerebral endothelial cell, or other immortalised cells compatible with the CNS.
  • transfection with a viral vector are used for transplantation of the packaging cell, several cells may be used, e.g. immortalised neural stem cells, immortalised cerebral endothelial cell, or other immortalised cell compatible with CNS.
  • immortalised neural stem cells e.g. immortalised neural stem cells, immortalised cerebral endothelial cell, or other immortalised cell compatible with CNS.
  • human cells may be used, e.g. immortalised neural stem cells, immortalised cerebral endothelial cell, or fibroblast-like human cell, e.g. HEK 293 or HeLa
  • the packaging cell line of the invention can be selected from an element of the group consisting of psi-2 [Mann R, Mulligan R C, & Buttimore D; CeN 1983 33 153- 159], psi-Crip [Danos O & Mulligan R C; Proc. Natl. Acad. Sci. USA 1988 85, 6460-
  • the packaging cell line is made from human cells, e.g. HT1080 cells (WO-A1-9621014), HEK 293, thereby allowing production of recombinant retrovirus that are capable of surviving inactivation by human serum.
  • the present invention resides in the use of a ubiquitin promoter to direct in vivo expression of therapeutic genes after transfer of such genes to the central nervous system. Accordingly, in a particular embodiment, the invention is directed to the use of the recombinant expression vector of the invention for the manufacture of a pharmaceutical composition, useful for the treatment of a neurological disease or disorder.
  • the pharmaceutical composition of the invention preferably is a composition suited for injection or implantation into the human brain. Such compositions may be provided in the form of vials of frosen cells, optionally provided in a freeze medium in suspension.
  • compositions suited for injection or implantation may be prepared by the skilled person according to conventional methods (see e.g. Scientia Medicinalis. 1997, Schering AG, ISSN 1433-190X).
  • the pharmaceutical compounds of the present invention may thus be formulated for in ampoules, pre-filled syringes, small volume infusion containers, etc.
  • the compositions may take such forms as suspensions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilising and/or dispersing agents.
  • the invention provides a method for the treatment of a living body suffering from a neurological disease or disorder which is responsive to a therapeutically active polypeptide, which method comprises administering to the living body a retroviral particle obtained by the method of the invention, encoding the therapeutically active polypeptide; or by implanting into the living body, cells of the invention, producing the therapeutically active polypeptide.
  • the neurological disorders contemplated according to the present invention are e.g.
  • ischemic stroke angiogenesis; metabolic disease of the brain; axonal injury; spinal cord injury; Alzheimer's disease; Amyothropic Lateral Sclerosis (ALS); Parkinson's disease; Huntington's disease; motor neuron disease; central nervous system infections; epilepsy; post polio syndrome; mucopolysaccharidoses (MPS), in particular MPS types I to VII; lipidosis, in particular Gaucher's disease; Lesch-Nyhan syndrome; X-linked ADL; metachromatic leukodystrophy; Krabbe's disease; Charcot- Marie-Tooth disease; Fragile X; epilepsy; Down's syndrome; phenylketonuria; 5 degenerative disorders; or mental disorders.
  • MPS mucopolysaccharidoses
  • the human UbC promoter was cloned into a modified version of pcDNA3.1/Zeo.
  • the unmodified pcDNA3.1/Zeo is commercial available from Invitogen.
  • the modified version is smaller than the parent vector, since the ampicillin gene (from position 3933 to 5015) and a sequence from position 2838 to 3134 were removed.
  • the ampicillin gene from position 3933 to 5015
  • a sequence from position 2838 to 3134 were removed.
  • EGFP Enhanced Green Fluorescence Protein
  • HiB5 cells 25 14 embryo hipocampus [Refranz et al; Cell 1991 66 713-729].
  • the HiB5 cells were transfected with Lipid 6 from Invitrogen according to manufactories recommendations.
  • GDNF The gene encoding GDNF was cloned using conventional techniques or as described in WO 93/06116 and inserted into the pUbilZ expression vector. Clones of Immortalised neural stem cells (rat and human) and immortalised cerebral endothelial cells (rat or human) expressing therapeutic genes have been generated. Clones will be studied in short-term experiments in order to establish the ability of each cell line to secrete the introduced neurotrophic factor (e.g. NGF, GDNF, NTN or CNTF) in vivo in sufficiently high amounts to produce full neuroprotective effects in established models.
  • the introduced neurotrophic factor e.g. NGF, GDNF, NTN or CNTF
  • the cells will be implanted into the septum in rats subjected to a unilateral fimbria-fornix lesion, and the magnitude of cholinergic cell loss in the medial septal nucleus will be established 2 weeks after operation. From previous studies on NGF-secreting rat HiB5 and RBE cells it is known that an implant of 2 x 10 5 cells with an estimated in vivo secretion rate of 50 ng NGF/day will be able to provide complete protection in this model.
  • GDNF and NTN secreting cell lines cells will be implanted in the substantia nigra and striatum, first, in C57 mice treated one week later with a single injection of MPTP (40 mg/kg s.c). The loss of dopamine neurones in the nigra and the reduction in dopamine levels and tyrosine hydroxylase immunoreactivity in the striatum will be analysed 4 weeks after injection.
  • nigral neurones will be pre-labelled with the retrograde tracer FluoroGold from the striatum according to the procedure established in the Swedish laboratory, in order to provide an accurate estimation of the magnitude of lesion induced nigral cell death.
  • transplants of cells expressing the reporter gene, EGFP will be used as controls. Since the minimum number of cells necessary to produce a full neuroprotective effect has not been established for GDNF and NTN, different cell numbers, from 2-8x10 5 cells, will be tested.
  • CNTF and NGF cells will be evaluated by their protective effect against quinolinic acid lesion (200 nmol) of striatal medium size projection neurones. This type of lesion induces complete neuronal death in 2 days to one week. The cells will be transplanted prior to the lesion and neuroprotection assessed one week post lesion, determining neuroprotection in the striatum, and preservation of innervation of target territories in the SNr.
  • the long-term functional effects and the stability of transgene expression will be studied in animal models relevant to the three clinical conditions, Parkinson's disease, Huntington ' s chorea and dementia. Three animal models will be used to study long-term functional effects of the therapeutic cell lines.
  • Rats with unilateral intrastriatal 6-hydroxydopamine lesions will receive implants of GDNF or NTN producing cells, either into the substantia nigra, 3 days after the lesion (i.e. prior to the onset of cell death), or into the striatum, 4 weeks after the lesion (at the time when dopamine neuron degeneration is well advanced).
  • the ability of the cell implants to reverse the Parkinsonian condition in tests of spontaneous motor behaviour (stepping and paw use tests) and to prevent the progressive neuro- degeneration long-term will be studied over a 6 month period.
  • NGF-secreting human neural and endothelial cells will be implanted into the nucleus basalis and septum in aged cognitively impaired rats (22-24 month old Sprague-Dawley rats) following the protocol used in previous studies on NGF secreting rat HiB5 cells in the Lund laboratory. Groups of young unimpaired rats, transplanted identically, will be studied in parallel. The ability of the cells to reverse the age dependent learning and memory impairment in the water maze test and the age-dependent cholinergic neuronal atrophy in basalis and septum will be studied over 1 and 6 months. EGFP expressing cells will be used as control.
  • the neuroprotective effects of the NGF or CNTF secreting human cell lines will be studied in rats receiving injections of quinolinic acid into the striatum, as above.
  • the cells will be implanted one week before the lesion, and the ability of the cells to block the excitotoxic striatal damage, as well as to prevent the development of impairments in motor behaviours, will be studied both short-term (4 days and 1 month) and long-term (6 months). Long-term expression of the transgene will be evaluated as above.
  • an UbC containing ProCon Vector and Production of Retroviral Particles by Packaging Cells In a preferred embodiment of the invention immortalised cerebral endothelial cells are transformed by a ProCon vector as described above carrying the gene encoding GDNF under transcriptional control of the human ubiquitin promoter.
  • the vector for transformation of the cerebral endothelial cells may be obtained by insertion of the human ubiquitin promoter prepared by PCR from the plasmid pTEJ-8 described in FEBS Letters. 1990 267 (2) 289-294, into the polylinker of the ProCon vector described in WO 96/07748.
  • a GDNF encoding gene may be cloned using conventional techniques or as described in WO 93/06116 and inserted into the body of the ProCon vector.
  • Retroviral particles are produced by introducing the ProCon vector obtained as above into a packaging cell line followed by isolation of the retroviral particles produced.
  • Immortalised cerebral endothelial cells may then be infected by the retroviral particles produced by the packaging cell line.
  • genes encoding GDNF or NGF may be introduced into rat cerebral endothelial cells (RBE) by transfection with an eukaryotic expression vector.
  • the eukaryotic expression vector may be constructed by insertion of GDNF cDNA or NGF cDNA into the polylinker of the plasmid pTEJ-8 or pUbMZ.
  • the resulting vector may be used to transfect immortalised cerebral endothelial cells of rat or human origin.
  • the genetically transformed immortalised endothelial cells may hereafter be implanted into the CNS of an individual suffering from Parkinson's disease or another disease susceptible to treatment with GDNF.
  • the immortalised neural stem cells or immortalised endothelial cells may be implanted into the striatum of the host but may also be implanted into other parts of the brain.
  • polylinker is used for a short stretch of artificially synthesised DNA which carries unique restriction sites allowing the easy insertion of any promoter or DNA segment.
  • GAGCGCAGCA AAATGGCGGC TGTTCCCGAG TCTTGAATGG AAGACGCTTG TGAGGCGGGC 960 TGTGAGGTCG TTGAAACAAG GTGGGGGGCA TGGTGGGCGG CAAGAACCCA AGGTCTTGAG 1020

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
  • Wood Science & Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Organic Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Biophysics (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Plant Pathology (AREA)
  • Biochemistry (AREA)
  • Physics & Mathematics (AREA)
  • Virology (AREA)
  • Medicinal Chemistry (AREA)
  • Microbiology (AREA)
  • Epidemiology (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

L'invention concerne des vecteurs d'expression de recombinaison portant un gène codant un polypeptide actif sur le plan thérapeutique, ledit gène étant placé sous la commande de transcription d'un promoteur d'ubiquitine. Elle concerne également l'utilisation d'un promoteur d'ubiquitine afin de diriger l'expression in vivo de gènes thérapeutiques après le transfert de ces gènes vers le système nerveux central.
PCT/DK1998/000037 1997-01-29 1998-01-29 Vecteurs d'expression et procedes d'expression in vivo de polypeptides therapeutiques WO1998032869A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
EP98900847A EP0961830A1 (fr) 1997-01-29 1998-01-29 VECTEURS D'EXPRESSION ET PROCEDES D'EXPRESSION $i(IN VIVO) DE POLYPEPTIDES THERAPEUTIQUES

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DK10297 1997-01-29
DK0102/97 1997-01-29

Publications (1)

Publication Number Publication Date
WO1998032869A1 true WO1998032869A1 (fr) 1998-07-30

Family

ID=8089764

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/DK1998/000037 WO1998032869A1 (fr) 1997-01-29 1998-01-29 Vecteurs d'expression et procedes d'expression in vivo de polypeptides therapeutiques

Country Status (2)

Country Link
EP (1) EP0961830A1 (fr)
WO (1) WO1998032869A1 (fr)

Cited By (41)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB2362884A (en) * 2000-05-30 2001-12-05 Isis Innovation Extended duration of airway gene therapy
WO2002006341A1 (fr) * 2000-07-14 2002-01-24 Children's Medical Center Corporation Facteur tropique capable de produire un effet salutaire sur le systeme nerveux chez un sujet
WO2002007774A2 (fr) * 2000-07-19 2002-01-31 The Regents Of The University Of California Methodes de traitement de maladies neurodegeneratives du cerveau
WO2002012514A2 (fr) * 2000-08-09 2002-02-14 Nsgene A/S Promoteur jet
WO2002024932A2 (fr) * 2000-09-18 2002-03-28 Genzyme Corporation Vecteurs d'expression contenant des promoteurs d'ubiquitine hybrides
WO2002036170A2 (fr) * 2000-11-03 2002-05-10 Oxford Biomedica (Uk) Limited Systeme vecteur
US6593133B1 (en) 1998-07-06 2003-07-15 Nsgene A/S Neurotrophic factors
US6683058B1 (en) 1998-04-15 2004-01-27 Regents Of The University Of California Methods for therapy of neurodegenerative disease of the brain
WO2004050879A1 (fr) * 2002-11-29 2004-06-17 Boehringer Ingelheim Pharma Gmbh & Co. Kg Vecteur d'expression, procedes pour realiser des produits geniques heterologues et procede de selection de cellules recombinantes hautement productives
EP1624067A2 (fr) * 2000-09-18 2006-02-08 Genzyme Corporation Vecteurs d'expression contenant des promoters d'ubiquitine hybrides
US7276580B2 (en) 2001-03-12 2007-10-02 Biogen Idec Ma Inc. Neurotrophic factors
US7384744B2 (en) 2002-11-29 2008-06-10 Boehringer Ingelheim Pharma Gmbh & Co., Kg Expression vector, methods for the production of heterologous gene products and for the selection of recombinant cells producing high levels of such products
US7442370B2 (en) 2001-02-01 2008-10-28 Biogen Idec Ma Inc. Polymer conjugates of mutated neublastin
WO2010083842A2 (fr) 2009-01-23 2010-07-29 Nsgene A/S Expression de neuropeptides dans des cellules mammaliennes
WO2010083841A2 (fr) 2009-01-23 2010-07-29 Nsgene A/S Lignées cellulaires améliorées et leur utilisation dans la biodélivrance de cellules encapsulées
WO2011011584A1 (fr) 2009-07-24 2011-01-27 Immune Design Corp Vecteurs lentiviraux pseudotypés avec une glycoprotéine d’enveloppe de virus sindbis
WO2012112691A1 (fr) 2011-02-15 2012-08-23 Immune Design Corp. Procédés d'amélioration des réponses immunitaires spécifiques d'un immunogène avec des vaccins vectorisés
WO2012141984A1 (fr) 2011-04-08 2012-10-18 Immune Design Corp. Compositions immunogènes et leurs procédés d'utilisation pour induire des réponses immunitaires humorales et cellulaires
WO2013149167A1 (fr) 2012-03-30 2013-10-03 Immune Design Corp. Particules de vecteur lentiviral ayant une meilleure efficacité de transduction pour des cellules exprimant dc-sign
WO2013173590A1 (fr) 2012-05-16 2013-11-21 Immune Design Corp. Vaccins contre le hsv-2
US8722862B2 (en) 2004-08-19 2014-05-13 Biogen Idec Ma Inc. Refolding transforming growth factor beta family proteins
WO2015123496A1 (fr) 2014-02-14 2015-08-20 Immune Design Corp. Immunothérapie du cancer par combinaison de stimulation immunitaire locale et systémique
US9138461B2 (en) 2007-05-01 2015-09-22 Biogen Ma Inc. Compositions and methods for increasing vascularization
WO2016011083A1 (fr) 2014-07-15 2016-01-21 Immune Design Corp. Régimes de primo-vaccination/rappel avec un adjuvant d'agoniste tlr4 et un vecteur lentiviral
WO2017068419A2 (fr) 2015-10-22 2017-04-27 Juno Therapeutics Gmbh Procédés, kits, agents et appareils de transduction
WO2017083356A1 (fr) 2015-11-09 2017-05-18 Immune Design Corp. Vecteur rétroviral pour l'administration et l'expression d'un réplicon arn exprimant des acides nucléiques hétérologues
WO2017083291A1 (fr) 2015-11-09 2017-05-18 Immune Design Corp. Compositions comprenant des vecteurs lentiviraux exprimant l'il-12 et leurs méthodes d'utilisation
CN106890342A (zh) * 2007-02-06 2017-06-27 俞泰俊 使用基因疗法治疗和预防神经变性疾病
WO2017147119A1 (fr) 2016-02-23 2017-08-31 Immune Design Corp. Préparations de vecteurs rétroviraux multigénomiques et procédés et systèmes pour les produire et les utiliser
WO2018023094A1 (fr) 2016-07-29 2018-02-01 Juno Therapeutics, Inc. Procédés d'évaluation de la présence ou de l'absence d'un virus compétent pour la réplication
WO2018106732A1 (fr) 2016-12-05 2018-06-14 Juno Therapeutics, Inc. Production de cellules modifiées pour une thérapie cellulaire adoptive
WO2018148180A2 (fr) 2017-02-07 2018-08-16 Immune Design Corp. Matériaux et méthodes pour l'identification et le traitement du cancer
CN108840943A (zh) * 2017-08-09 2018-11-20 安徽九川生物科技有限公司 一种重组鸡长效干扰素γ及制备此长效干扰素γ的融合蛋白及其制备方法
CN109053897A (zh) * 2017-08-09 2018-12-21 安徽九川生物科技有限公司 一种由鸡白细胞介素2、鸡干扰素γ和鸡干扰素α组成的融合蛋白及其制备方法
US10328125B2 (en) 2006-02-27 2019-06-25 Gloriana Therapeutics, Inc. Treatments for neurological disorders
WO2019152747A1 (fr) 2018-01-31 2019-08-08 Juno Therapeutics, Inc. Méthodes et réactifs d'évaluation de la présence ou de l'absence d'un virus compétent pour la réplication
US10973931B2 (en) 2014-09-16 2021-04-13 Universitat Autònoma De Barcelona Adeno-associated viral vectors for the gene therapy of metabolic diseases
US11001857B2 (en) 2010-07-12 2021-05-11 Universitat Autonoma De Barcelona Gene therapy composition for use in diabetes treatment
US11033638B2 (en) 2015-01-07 2021-06-15 Universität Autonoma De Barcelona Single-vector gene construct comprising insulin and glucokinase genes
WO2021154887A1 (fr) 2020-01-28 2021-08-05 Juno Therapeutics, Inc. Procédés pour la transduction de lymphocytes t
WO2023147515A1 (fr) 2022-01-28 2023-08-03 Juno Therapeutics, Inc. Procédés de fabrication de compositions cellulaires

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0342926A2 (fr) * 1988-05-17 1989-11-23 Mycogen Plant Science, Inc. Système de promoteur de l'ubiquitine végétale
WO1995009654A1 (fr) * 1993-10-01 1995-04-13 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Therapie genique concernant le systeme nerveux
WO1996007748A1 (fr) * 1994-09-02 1996-03-14 GSF-Forschungszentrum für Umwelt und Gesundheit GmbH Vecteurs retroviraux non auto-inactivants, utiles pour une expression ciblee
WO1997015664A1 (fr) * 1995-10-24 1997-05-01 Dr. Karl Thomae Gmbh Promoteur homologue puissant obtenu a partir de hamsters

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0342926A2 (fr) * 1988-05-17 1989-11-23 Mycogen Plant Science, Inc. Système de promoteur de l'ubiquitine végétale
WO1995009654A1 (fr) * 1993-10-01 1995-04-13 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Therapie genique concernant le systeme nerveux
WO1996007748A1 (fr) * 1994-09-02 1996-03-14 GSF-Forschungszentrum für Umwelt und Gesundheit GmbH Vecteurs retroviraux non auto-inactivants, utiles pour une expression ciblee
WO1997015664A1 (fr) * 1995-10-24 1997-05-01 Dr. Karl Thomae Gmbh Promoteur homologue puissant obtenu a partir de hamsters

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
JOHANSEN TE ET AL: "Biosynthesis of peptide precursors and protease inhibitors using new constitutive and inducible eukaryotic expression vectors.", FEBS LETT, JUL 16 1990, 267 (2) P289-94, NETHERLANDS, XP002069981 *
NENOI,M. ET AL.: "Heterogeneous structure of the polyubiquitin gene UbC of HeLa S3 cells", GENE, vol. 175, 1996, pages 179 - 185, XP004043312 *
SCHORPP, M. ET AL.: "The human ubiquitin C promoter directs high ubiquitous expression of transgenes in mice", NUCLEIC ACIDS RESEARCH, vol. 24, no. 9, 1996, pages 1787 - 1788, XP000001788 *

Cited By (71)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6683058B1 (en) 1998-04-15 2004-01-27 Regents Of The University Of California Methods for therapy of neurodegenerative disease of the brain
US8486385B2 (en) 1998-04-15 2013-07-16 Regents Of The University Of California Methods for therapy of neurodegenerative disease of the brain
EP2290079A2 (fr) 1998-07-06 2011-03-02 NsGene A/S Facteurs neurotrophiques
US6734284B1 (en) 1998-07-06 2004-05-11 Nsgene A/S Neublastin neurotrophic factors
US6593133B1 (en) 1998-07-06 2003-07-15 Nsgene A/S Neurotrophic factors
WO2001091800A1 (fr) * 2000-05-30 2001-12-06 Isis Innovation Limited Promoteur d'ubiquitine des vecteurs utilises dans la therapie genique des voies respiratoires
GB2362884A (en) * 2000-05-30 2001-12-05 Isis Innovation Extended duration of airway gene therapy
WO2002006341A1 (fr) * 2000-07-14 2002-01-24 Children's Medical Center Corporation Facteur tropique capable de produire un effet salutaire sur le systeme nerveux chez un sujet
WO2002007774A2 (fr) * 2000-07-19 2002-01-31 The Regents Of The University Of California Methodes de traitement de maladies neurodegeneratives du cerveau
EP2060267A3 (fr) * 2000-07-19 2009-08-05 The Regents Of The University Of California Procédés pour la thérapie d'une maladie neuro-dégénérative du cerveau
WO2002007774A3 (fr) * 2000-07-19 2003-01-16 Univ California Methodes de traitement de maladies neurodegeneratives du cerveau
AU2001261758B2 (en) * 2000-07-19 2005-07-21 Regents Of The University Of California Methods for therapy of neurodegenerative disease of the brain
US6555674B2 (en) 2000-08-09 2003-04-29 Nsgene A/S JeT promoter
WO2002012514A3 (fr) * 2000-08-09 2002-08-08 Nsgene As Promoteur jet
WO2002012514A2 (fr) * 2000-08-09 2002-02-14 Nsgene A/S Promoteur jet
WO2002024932A3 (fr) * 2000-09-18 2002-08-15 Genzyme Corp Vecteurs d'expression contenant des promoteurs d'ubiquitine hybrides
EP1624067A2 (fr) * 2000-09-18 2006-02-08 Genzyme Corporation Vecteurs d'expression contenant des promoters d'ubiquitine hybrides
EP1624067A3 (fr) * 2000-09-18 2006-03-15 Genzyme Corporation Vecteurs d'expression contenant des promoters d'ubiquitine hybrides
WO2002024932A2 (fr) * 2000-09-18 2002-03-28 Genzyme Corporation Vecteurs d'expression contenant des promoteurs d'ubiquitine hybrides
WO2002036170A2 (fr) * 2000-11-03 2002-05-10 Oxford Biomedica (Uk) Limited Systeme vecteur
WO2002036170A3 (fr) * 2000-11-03 2002-08-22 Oxford Biomedica Ltd Systeme vecteur
US7442370B2 (en) 2001-02-01 2008-10-28 Biogen Idec Ma Inc. Polymer conjugates of mutated neublastin
US7655463B2 (en) 2001-03-12 2010-02-02 Biogen Idec Ma Inc. Methods of activating RET receptor tyrosine kinase using neurotrophic factors
US7358228B2 (en) 2001-03-12 2008-04-15 Biogen Idec Ma Inc. Methods of treating pain using neurotrophic factors
US7276580B2 (en) 2001-03-12 2007-10-02 Biogen Idec Ma Inc. Neurotrophic factors
US7384744B2 (en) 2002-11-29 2008-06-10 Boehringer Ingelheim Pharma Gmbh & Co., Kg Expression vector, methods for the production of heterologous gene products and for the selection of recombinant cells producing high levels of such products
WO2004050879A1 (fr) * 2002-11-29 2004-06-17 Boehringer Ingelheim Pharma Gmbh & Co. Kg Vecteur d'expression, procedes pour realiser des produits geniques heterologues et procede de selection de cellules recombinantes hautement productives
US8722862B2 (en) 2004-08-19 2014-05-13 Biogen Idec Ma Inc. Refolding transforming growth factor beta family proteins
US8969042B2 (en) 2004-08-19 2015-03-03 Biogen Idec Ma Inc. Refolding transforming growth factor beta family proteins
US10328125B2 (en) 2006-02-27 2019-06-25 Gloriana Therapeutics, Inc. Treatments for neurological disorders
CN106890342A (zh) * 2007-02-06 2017-06-27 俞泰俊 使用基因疗法治疗和预防神经变性疾病
US9138461B2 (en) 2007-05-01 2015-09-22 Biogen Ma Inc. Compositions and methods for increasing vascularization
WO2010083842A2 (fr) 2009-01-23 2010-07-29 Nsgene A/S Expression de neuropeptides dans des cellules mammaliennes
WO2010083841A2 (fr) 2009-01-23 2010-07-29 Nsgene A/S Lignées cellulaires améliorées et leur utilisation dans la biodélivrance de cellules encapsulées
EP2770061A1 (fr) 2009-07-24 2014-08-27 Immune Design Corp. Vecteurs lentiviraux non intégrants
WO2011011584A1 (fr) 2009-07-24 2011-01-27 Immune Design Corp Vecteurs lentiviraux pseudotypés avec une glycoprotéine d’enveloppe de virus sindbis
US11001857B2 (en) 2010-07-12 2021-05-11 Universitat Autonoma De Barcelona Gene therapy composition for use in diabetes treatment
WO2012112691A1 (fr) 2011-02-15 2012-08-23 Immune Design Corp. Procédés d'amélioration des réponses immunitaires spécifiques d'un immunogène avec des vaccins vectorisés
US9044420B2 (en) 2011-04-08 2015-06-02 Immune Design Corp. Immunogenic compositions and methods of using the compositions for inducing humoral and cellular immune responses
EP3632463A1 (fr) 2011-04-08 2020-04-08 Immune Design Corp. Compositions immunogènes et leurs procédés d'utilisation pour induire des réponses immunitaires humorales et cellulaires
WO2012141984A1 (fr) 2011-04-08 2012-10-18 Immune Design Corp. Compositions immunogènes et leurs procédés d'utilisation pour induire des réponses immunitaires humorales et cellulaires
EP3480316A1 (fr) 2012-03-30 2019-05-08 Immune Design Corp. Particules de vecteur lentiviral ameliorée en efficacite de la transduction de cellules exprimant dc-sign
WO2013149167A1 (fr) 2012-03-30 2013-10-03 Immune Design Corp. Particules de vecteur lentiviral ayant une meilleure efficacité de transduction pour des cellules exprimant dc-sign
EP3388835A1 (fr) 2012-05-16 2018-10-17 Immune Design Corp. Vaccins contre le hsv-2
WO2013173590A1 (fr) 2012-05-16 2013-11-21 Immune Design Corp. Vaccins contre le hsv-2
EP3542816A1 (fr) 2014-02-14 2019-09-25 Immune Design Corp. Immunothérapie du cancer par combinaison de stimulation immunitaire locale et systémique
WO2015123496A1 (fr) 2014-02-14 2015-08-20 Immune Design Corp. Immunothérapie du cancer par combinaison de stimulation immunitaire locale et systémique
WO2016011083A1 (fr) 2014-07-15 2016-01-21 Immune Design Corp. Régimes de primo-vaccination/rappel avec un adjuvant d'agoniste tlr4 et un vecteur lentiviral
US10973931B2 (en) 2014-09-16 2021-04-13 Universitat Autònoma De Barcelona Adeno-associated viral vectors for the gene therapy of metabolic diseases
US11033638B2 (en) 2015-01-07 2021-06-15 Universität Autonoma De Barcelona Single-vector gene construct comprising insulin and glucokinase genes
WO2017068419A2 (fr) 2015-10-22 2017-04-27 Juno Therapeutics Gmbh Procédés, kits, agents et appareils de transduction
US12129477B2 (en) 2015-10-22 2024-10-29 Juno Therapeutics Gmbh Methods, kits, agents and apparatuses for transduction
US11135283B2 (en) 2015-11-09 2021-10-05 Immune Design Corp. Retroviral vector for the administration and expression of replicon RNA expressing heterologous nucleic acids
US11365230B2 (en) 2015-11-09 2022-06-21 Immune Design Corp. Compositions comprising lentiviral vectors expressing IL-12 and methods of use thereof
WO2017083291A1 (fr) 2015-11-09 2017-05-18 Immune Design Corp. Compositions comprenant des vecteurs lentiviraux exprimant l'il-12 et leurs méthodes d'utilisation
EP3738973A1 (fr) 2015-11-09 2020-11-18 Immune Design Corp. Compositions comprenant des vecteurs lentiviraux exprimant il-12 et leurs procédés d'utilisation
WO2017083356A1 (fr) 2015-11-09 2017-05-18 Immune Design Corp. Vecteur rétroviral pour l'administration et l'expression d'un réplicon arn exprimant des acides nucléiques hétérologues
WO2017147119A1 (fr) 2016-02-23 2017-08-31 Immune Design Corp. Préparations de vecteurs rétroviraux multigénomiques et procédés et systèmes pour les produire et les utiliser
US10584356B2 (en) 2016-02-23 2020-03-10 Immune Design Corp. Multigenome retroviral vector preparations and methods and systems for producing and using same
US10179919B2 (en) 2016-02-23 2019-01-15 Immune Design Corp. Multigenome retroviral vector preparations and methods and systems for producing and using same
US11421287B2 (en) 2016-07-29 2022-08-23 Juno Therapeutics, Inc. Methods for assessing the presence or absence of replication competent virus
WO2018023094A1 (fr) 2016-07-29 2018-02-01 Juno Therapeutics, Inc. Procédés d'évaluation de la présence ou de l'absence d'un virus compétent pour la réplication
WO2018106732A1 (fr) 2016-12-05 2018-06-14 Juno Therapeutics, Inc. Production de cellules modifiées pour une thérapie cellulaire adoptive
WO2018148180A2 (fr) 2017-02-07 2018-08-16 Immune Design Corp. Matériaux et méthodes pour l'identification et le traitement du cancer
CN109053897A (zh) * 2017-08-09 2018-12-21 安徽九川生物科技有限公司 一种由鸡白细胞介素2、鸡干扰素γ和鸡干扰素α组成的融合蛋白及其制备方法
CN108840943A (zh) * 2017-08-09 2018-11-20 安徽九川生物科技有限公司 一种重组鸡长效干扰素γ及制备此长效干扰素γ的融合蛋白及其制备方法
WO2019152747A1 (fr) 2018-01-31 2019-08-08 Juno Therapeutics, Inc. Méthodes et réactifs d'évaluation de la présence ou de l'absence d'un virus compétent pour la réplication
US11535903B2 (en) 2018-01-31 2022-12-27 Juno Therapeutics, Inc. Methods and reagents for assessing the presence or absence of replication competent virus
US12264371B2 (en) 2018-01-31 2025-04-01 Juno Therapeutics, Inc. Methods and reagents for assessing the presence or absence of replication competent virus
WO2021154887A1 (fr) 2020-01-28 2021-08-05 Juno Therapeutics, Inc. Procédés pour la transduction de lymphocytes t
WO2023147515A1 (fr) 2022-01-28 2023-08-03 Juno Therapeutics, Inc. Procédés de fabrication de compositions cellulaires

Also Published As

Publication number Publication date
EP0961830A1 (fr) 1999-12-08

Similar Documents

Publication Publication Date Title
WO1998032869A1 (fr) Vecteurs d'expression et procedes d'expression in vivo de polypeptides therapeutiques
Takamiya et al. An experimental model of retrovirus gene therapy for malignant brain tumors
Kwok et al. Retroviral transfer of genes into canine hemopoietic progenitor cells in culture: a model for human gene therapy.
US6872528B2 (en) Highly productive packaging lines
KR100246096B1 (ko) 유전자 요법을 위한 개선된 레트로바이러스 벡터
EP0848757B1 (fr) Utilisation de sequences regulatrices de la proteine acide du lactoserum ou du virus de la tumeur mammaire de la souris, aux fins d'expression ciblee de genes heterologues lies dans des cellules mammaires humaines et notamment dans des cellules du cancer mammaire humain
US6309878B1 (en) Glucose-inducible recombinant viral vector
US5736360A (en) Endothelial cell tropic compositions and methods of making and using the same
WO1997039629A1 (fr) Vecteurs viraux incluant des polynucleotides codant des facteurs neurotrophiques ainsi que leur utilisation
AU729934B2 (en) Retroviral vector and its use in gene therapy
JP2001500021A (ja) 新規な内部リボソームエントリー部位およびそれを含有するベクター
JP2002519069A (ja) レトロウイルスベクターを用いた染色体へのターゲッティングされた組込み
US6630322B1 (en) Generating replicating molecules in vivo
JPH11507244A (ja) Ｓｒｖ−３エンベロープ糖タンパク質配列で偽型化したレトロウイルスベクター
KR20030090609A (ko) 씨에이알피 유전자의 상류 서열, 이를 함유하는 벡터 및이들의 용도
US20020168760A1 (en) Retroviral vectors for gene transfer into neuronal cells
Baum et al. Salivary glands as a model for craniofacial applications of gene transfer
WO1994028151A1 (fr) Therapy genique s'appliquant a l'hemophilie
KR100720201B1 (ko) 트랜스진의 신경-특이성 발현을 위한 음성 조절요소의 용도
WO1998002564A1 (fr) Vecteur hybride de virus herpetique/virus de epstein-barr
KR100537142B1 (ko) 복제성분자의 생체내 제조방법
Lee et al. Liver-directed gene therapy: 2000 and beyond
Lea et al. The Scope of Viral Vectors for the Transduction of Haemopoietic Cells
Csonka Generation of prototype human satellite DNA-based artificial chromosomes
MXPA00007617A (en) Use of negative regulation elements for nerve-specific expression of transgenes

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): JP US

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FI FR GB GR IE IT LU MC NL PT SE

DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
121 Ep: the epo has been informed by wipo that ep was designated in this application
WWE Wipo information: entry into national phase

Ref document number: 1998900847

Country of ref document: EP

WWE Wipo information: entry into national phase

Ref document number: 09355457

Country of ref document: US

WWP Wipo information: published in national office

Ref document number: 1998900847

Country of ref document: EP

NENP Non-entry into the national phase

Ref country code: JP

Ref document number: 1998531504

Format of ref document f/p: F

WWW Wipo information: withdrawn in national office

Ref document number: 1998900847

Country of ref document: EP

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载