280 results about "Viral Genes" patented technology

The present invention relates methods of generating infectious negative-strand virus in host cells by an entirely vector-based system without the aid of a helper virus. In particular, the present invention relates methods of generating infectious recombinant negative-strand RNA viruses intracellularly in the absence of helper virus from expression vectors comprising cDNAs encoding the viral proteins necessary to form ribonucleoprotein complexes (RNPs) and expression vectors comprising cDNA for genomic viral RNA(s) (vRNAs) or the corresponding cRNA(s). The present invention also relates to methods of generating infectious recombinant negative-strand RNA viruses which have mutations in viral genes and/or which express, package and/or present peptides or polypeptides encoded by heterologous nucleic acid sequences. The present invention further relates the use of the recombinant negative-strand RNA viruses or chimeric negative-strand RNA viruses of the invention in vaccine formulations and pharmaceutical compositions.

Disclosed and claimed are methods of non-invasive genetic immunization in an animal and/or methods of inducing a systemic immune or therapeutic response in an animal, products therefrom and uses for the methods and products therefrom. The methods can include contacting skin of the animal with a vector in an amount effective to induce the systemic immune or therapeutic response in the animal. The vector can include and express an exogenous nucleic acid molecule encoding an epitope or gene product of interest. The systemic immune response can be to or from the epitope or gene product. The nucleic acid molecule can encode an epitope of interest and/or an antigen of interest and/or a nucleic acid molecule that stimulates and/or modulates an immunological response and/or stimulates and/or modulates expression, e.g., transcription and/or translation, such as transcription and/or translation of an endogenous and/or exogenous nucleic acid molecule; e.g., one or more of influenza hemagglutinin, influenza nuclear protein, influenza M2, tetanus toxin C-fragment, anthrax protective antigen, anthrax lethal factor, rabies glycoprotein, HBV surface antigen, HIV gp 120, HIV gp 160, human carcinoembryonic antigen, malaria CSP, malaria SSP, malaria MSP, malaria pfg, and mycobacterium tuberculosis HSP; and/or a therapeutic, an immunomodulatory gene, such as co-stimulatory gene and/or a cytokine gene. The immune response can be induced by the vector expressing the nucleic acid molecule in the animal's cells. The animal's cells can be epidermal cells. The immune response can be against a pathogen or a neoplasm. A prophylactic vaccine or a therapeutic vaccine or an immunological composition can include the vector. The animal can be a vertebrate, e.g., a mammal, such as human, a cow, a horse, a dog, a cat, a goat, a sheep or a pig; or fowl such as turkey, chicken or duck. The vector can be one or more of a viral vector, including viral coat, e.g., with some or all viral genes deleted therefrom, bacterial, protozoan, transposon, retrotransposon, and DNA vector, e.g., a recombinant vector; for instance, an adenovirus, such as an adenovirus defective in its E1 and/or E3 and/or E4 region(s). The method can encompass applying a delivery device including the vector to the skin of the animal, as well as such a method further including disposing the vector in and/or on the delivery device. The vector can have all viral genes deleted therefrom. The vector can induce a therapeutic and/or an anti-tumor effect in the animal, e.g., by expressing an oncogene, a tumor-suppressor gene, or a tumor-associated gene. Immunological products generated by the expression, e.g., antibodies, cells from the methods, and the expression products, are likewise useful in in vitro and ex vivo applications, and such immunological and expression products and cells and applications are disclosed and claimed. Methods for expressing a gene product in vivo and products therefor and therefrom including mucosal and/or intranasal administration of an adenovirus, advantageously an E1 and/or E3 and/or E4 defective or deleted adenovirus, such as a human adenovirus or canine adenovirus, are also disclosed and claimed.

The present invention relates to genetically engineered attenuated viruses and methods for their production. In particular, the present invention relates to engineering live attenuated viruses which contain a modified NS gene segment. Recombinant DNA techniques can be utilized to engineer site specific mutations into one or more noncoding regions of the viral genome which result in the down-regulation of one or more viral genes. Alternatively, recombinant DNA techniques can be used to engineer a mutation, including but not limited to an insertion, deletion, or substitution of an amino acid residue(s) or an epitope(s) into a coding region of the viral genome so that altered or chimeric viral proteins are expressed by the engineered virus.

The innate immune system of eukaryotes is able to recognise foreign genetic material by means of Toll-like receptors and to initiate signal transduction cascades that trigger an antiviral state of cell populations by way of an interferon response. That antiviral state is also a barrier for non-viral gene delivery systems. If the signal transduction cascade is interrupted intracellularly or intercellularly, transfection efficiencies of non-viral gene delivery systems can be increased and undesirable changes in the expression profile can be avoided. Since RNA-interference is to be attributed to the antiviral state, the RNAi machinery is likewise activated after activation of the innate immune system. In that way, knock-down efficiencies on transfection with siRNA can be increased.

A method for producing viral gene delivery vehicles which can be transferred to pre-selected cell types by using targeting conjugates. The gene delivery vehicles comprise: 1) the gene of interest; and 2) a viral capsid or envelope carrying a member of a specific binding pair, the counterpart of which is not directly associated with the surface of the target cell. These vehicles can be rendered unable to bind to their natural cell receptor. The targeting conjugates include the counterpart member of the specific binding pair, linked to a targeting moiety which is a cell-type specific ligand (or fragments thereof). The number of the specific binding pair present on the viral vehicles can be, for example, an immunoglobulin binding moiety (e.g., capable of binding to a Fc fragment, protein A, protein G, FcR or an anti-Ig antibody), or biotin, avidin or streptavidin. The virus' outer membrane or capsid may contain a substance which mediates entrance of the gene delivery vehicle into the target cell. Due to the specificity of the ligand, the binding pair's high affinity, and the gene delivery vehicle's inability to be targeted when used alone, the universality of the method for gene delivery, together with its high cell type selectively can be achieved by using various targeting conjugates.

A genetically disabled mutant virus has a genome which is defective in respect of a selected gene that is essential for the production of infectious new virus particles, and which carries heterologous genetic material encoding an immunomodulatory protein such as GM-CSF, IL-2, or others, such that the mutant virus can infect normal host cells and cause expression of immunomodulatory protein, but the mutant virus cannot cause production of infectious new virus particles except when the virus infects recombinant complementing host cells expressing a gene that provides the function of the essential viral gene; the site of insertion of the heterologous genetic material encoding the immunomodulatory protein preferably being at the site of the defect in the selected essential viral gene. Uses include prophylactic and therapeutic use in generating an immune response in a subject treated therewith; use in the preparation of an immunogen such as a vaccine for use in tumor therapy; use in the in-vitro expansion of (e.g. virus-specific) cytotoxic T cells; and therapeutic or prophylactic use in corrective gene therapy.

The present invention relates to the engineering of recombinant influenza viruses that express tumor-associated antigens. Expression of tumor-associated antigens by these viruses can be achieved by engineering specific epitopes into influenza virus proteins, or by engineering viral genes that encode a viral protein and the specific antigen as independent polypeptides. Tumor-bearing patients can be immunized with the recombinant influenza viruses alone, or in combination with another treatment, to induce an immune response that leads to tumor reduction. The recombinant viruses can also be used to vaccinate high risk tumor-free patients to prevent tumor formation in vivo.

African swine fever virus (ASFV) is the etiological agent of a contagious and often lethal viral disease of domestic pigs. Control of ASF has been hampered by the unavailability of vaccines. Experimental vaccines have been derived from naturally occurring, cell culture-adapted, or genetically modified live attenuated ASFVs; however, these vaccines are only successful when protecting against homologous viruses. Among viral genes reported to be involved in virulence are components of the multi gene family (MGF). Here we report the construction of a recombinant ΔMGF virus derived from the highly virulent ASFV Georgia 2007 (ASFV-G) isolate. In vivo, ASFV-G ΔMGF administered intramuscularly (IM) to swine at either 10² or 10⁴ HAD₅₀ are completely attenuated; the inoculated animals are completely asymptomatic. Animals infected with 10² or 10⁴ HAD50 of ASFV-G ΔMGF are protected against the presentation of clinical disease when challenged at 28 days post infection with the virulent parental strain Georgia 2007.

The invention discloses a glutathione-modified chitosan copolymer serving as a non-viral gene carrier material and preparation and application thereof, which belong to the fields of gene therapy and novel materials. A preparation method of the copolymer comprises the following steps of: (1) synthesizing an allyl-modified chitosan derivative; (2) synthesizing brush-like PEG (Polyethylene Glycol) polymer chains with different molecular weights through RAFT (Reversible Addition-Fragmentation Chain Transfer); (3) grafting the brush-like PEG onto a chitosan framework by adopting a free radical coupling method; and (4) linking glutathione to the chain end of the brush-like PEG by adopting an EDC (1-Ethyl-3-(3-Dimethyllaminopropyl) Carbodiimide hydrochloride)/NHS (N-hydroxysuccinimide) activation method to obtain a glutathione-modified chitosan copolymer carrier material. The glutathione-modified chitosan copolymer obtained by adopting the technology serves as the non-viral gene carrier material. By adopting the copolymer, the endocytosis function of a composite nanoparticle formed from the copolymer and DNA (Deoxyribonucleic Acid) can be remarkably enhanced, the releasing mechanism of DNA from a composite particle after cell entrance is improved, and the non-viral gene carrier material with a high transfection efficiency is further obtained.

It is an object of the present invention to provide a polyion complex used as a non-viral gene vector, which achieves sufficiently high gene expression efficiency to a target cell. The polyion complex of the present invention comprises a block copolymer formed by binding polyethylene glycol to polycation via a disulfide group and a nucleic acid.

The invention belongs to technology field of carrier material for non-viral gene transfection, and relates to a carrier material for non-viral gene transfection, plasmid DNA compound particle and preparation method and administration. The weight ratio of polyethylene imine and hydrophobic unit containing benzene ring is 1:9-4:6.The polyethylene imine is superbranched polyethylene imine (PEI) with average molecular weight of 1.8K-25K.The hydrophobic unit containing benzene ring is one or more of gamma -benzyl-L-glutamic acid, epsi -carbobenzoxy lysine and phenylalanine at any ratio. The water soluble polyethylene imine modified substance of hydrophobic unit containing benzene ring can compound with genic material to form a particle with positive charge on surface and particle diameter of 50-300nm.The compound particle can transfer loaded genic material into cell to realize expression of genic material and complete transfection process. The invention prepared non-gene transfection carrier material has the advantages of low toxicity and high up transfection efficiency; and can be used as transfection agent in vivo and in vitro.

The invention relates to a novel cationic graft copolymer, and a preparation method and an application of a multiple composite non-viral gene vector. The novel cationic graft copolymer PCL-g-LPEI (polycaprolactone-g-linear polyethylenimine) is synthesized by ring-opening copolymerization and amidation reaction. The multiple composite non-viral gene vector comprises: (A) a plasmid DNA (deoxyribonucleic acid)-supported PCL-g-LPEI cationic compound serving as a kernel of a multiple gene compound; (B) a tumor-targeting ligand HA (hyaluronic acid) wrapping the cationic compound under the interaction of positive and negative charges to form a shell of the multiple gene compound. A multiple composite gene constructed by the non-viral gene vector has the advantages of low cytotoxicity, high blood compatibility, high transfection efficiency, tumor targeting effects and the like, and is expected to be applied to clinical gene therapy.

The invention provides biodegradable, cationic compositions based on cationic α-amino acid-containing PEA, PEUR and PEU polymers for use in preparation of non-viral gene transfer compositions. In the invention gene transfer compositions a poly nucleic acid is condensed with the polymer to form a soluble unit wherein the electrical charge of the poly nucleic acid is neutralized by the polymers. The invention gene transfer compositions can be used to transfect target cells by contact with the target cells.

The invention provides isolated polynucleotide molecules that comprise a DNA sequence encoding an infectious RNA sequence encoding a genetically-modified North American PRRS virus, methods to make it and related polypeptides, polynucleotides, and various components. Vaccines comprising the genetically modified virus and polynucleotides and a diagnostic kit to distinguish between naturally infected and vaccinated animals are also provided.

The invention discloses a tetravinyl-based Gemini type amphiphilic compound as well as a preparation method and application thereof. The compound disclosed by the invention is mainly prepared through McMurry coupling reaction, nucleophilic substitution reaction and Click reaction; the structure of the compound is also confirmed through infrared, nuclear magnetic and mass-spectrum ways; in an aqueous solution, molecules of a derivative of the compound disclosed by the invention self-assemble to form aggregation-induced fluorescence-enhanced micelles (AIE micelles); the compound acts with a nucleic acid, and then can co-aggregate to form nano particles easy for cellular uptake; green fluorescent protein (GFP) and luciferase (Luciferase) expression assays prove that the compound self and a liposome formed with dioleoyl phosphatidyl ethanolamine (DOPE) can be used as non-viral gene vectors; meanwhile, the derivative is successfully used for tracing the cellular uptake and release processes of pGL-3 and FAM-DNA by utilizing the reversible transformation between the self-assembly of the compound and the co-assembly with DNA (Deoxyribonucleic Acid).

The invention discloses a target quaternary ammonium salt cationic polymer lipid gene carrier, a preparation method and an application thereof. The target quaternary ammonium salt cationic polymer lipid genetic carrier is characterized in that: a polymeric quaternary ammonium salt and lipid are adopted for preparing a quaternary ammonium salt cationic polymer lipid genetic carrier according to a mass ratio, wherein the mass ratio of the polymeric quaternary ammonium salt to the lipid is 0.05-20:1; then a assembly method or a modification method is adopted for modifying to prepare a folic acid or EGFR antibody modified cationic polymer lipid gene carrier. Results of gene transfection experiments show that: gene transfection efficiencies of the target quaternary ammonium salt cationic polymer lipid gene carrier in 293T cells and NIH-3T3 cells are the same as the gene transfection efficiencies of positive control lipofectamine of lipofectamine<TM>2000 in the 293T cells and the NIH-3T3 cells; the gene transfection efficiencies of the EGFR antibody modified cationic polymer lipid genetic carrier in liver cancer Huh-7 cells and breast cancer MCF-7 cells are higher than the gene transfection efficiencies of the lipofectamine<TM>2000 in the liver cancer Huh-7 cells and the breast cancer MCF-7 cells. The cationic polymer lipid genetic carrier system provided by the present invention has good biocompatibility and low cytotoxicity, and can be as an excellent non-viral gene delivery carrier.

No antiviral regimen has been consistently successful in treating H5N1 virus infection. We demonstrate that a group of highly effective siRNAs targeting different H5N1 viral genes shares a unique motif, GGAGU/ACUCC. We further demonstrate that the effectiveness of siRNAs containing this motif is not sequence specific. The results suggested that the structure of the unique motif is critical in determining the potency of siRNA-mediated protective effects against viral infection and this potent in vivo protection is associated with early productions of β-defensin and IL-6 induced by the motif. Provided are methods and prophylactic and therapeutic agents useful against other viral infections in addition to the H5N1 influenza virus.

The present invention relates to a method for producing helper-free stocks of recombinant adeno-associated virus (rAAV) which can be used to efficiently and stably transduce foreign genes into host cells or organisms. The method comprises the cotransfection of eukaryotic cells with rAAV and with helper AAV DNA in the presence of helper virus (e.g. adenovirus or herpesvirus) such that the helper AAV DNA is not associated with virion formation. The crux of the invention lies in the inubility of the helper AAV DNA to recombine with rAAV vector, thereby preventing the generation of wild-type virus. In a specific embodiment of the invention, the vector comprises a recombinant AAV genome containing only the terminal regions of the AAV chromosome bracketing a non-viral gene, and the helper AAV DNA comprises a recombinant AAV genome containing that part of the AAV genome which is not present in the vector, and in which the AAV terminal regions are replaced by adenovirus sequences. In a further embodiment of the invention, cell lines are created which incorporate helper AAV DNA which can directly produce substantially pure recombinant AAV virus. The pure stocks of recombinant AAV produced according to the invention provide an AAV viral expression vector system with increased yield of recombinant virus, improved efficiency, higher definition, and greater safety than presently used systems.