Abstract: The present invention relates to methods of amplifying a nucleotide sequence. The nucleotide sequence to be amplified is flanked by CARE elements. The invention provides nucleic acid molecules (e.g. plasmids and vectors) comprising first and second CARE elements, flanking the nucleotide sequence to be amplified. The invention also provides host cells comprising such nucleic acid molecules and methods of amplification using such nucleic acid molecules. The invention is particularly applicable to the amplification of viral genes and the production of recombinant adeno-associated viruses (AAVs).

Abstract: The present invention relates to an adenoviral vector comprising a regulatable Major Late Promoter and an exogenous transgene. The invention also provides cells comprising such adenoviral vectors, and processes using such vectors.

Abstract: The present invention relates to a process for producing recombinant adeno-associated virus (AAV) particles, described herein as trans-pseudotyping. The process involves the production of recombinant AAV particles in first host cells and then in second host cells, wherein first and second AAV cap genes are expressed in the first and second host cells, respectively, thus producing first and second recombinant AAV particles which are encapsidated by first and second AAV capsid polypeptides. The first and second recombinant AAV particles have different cell tropisms, preferably towards production cell lines (for high efficiency production of AAVs) and for cells associated with a therapeutic indication (for treatment of such an indication), respectively.

Abstract: The present invention relates to a process for producing a library of recombinant adeno-associated vims (AAV) particles which are encapsidated by a variety of different capsid polypeptides. Each recombinant AAV particle in the library comprises an AAV genome which comprises AAV ITRs flanking a first AAV cap gene encoding a first Cap polypeptide, wherein the particles in the library differ in the nucleotide sequences of their first AAV cap genes, and wherein each particle in the library is encapsidated by a Cap polypeptide which is encoded by the cap gene within its AAV genome. The library may be used to screen for recombinant AAV particles which have high specificity for cells of a target therapeutic tissue.

Abstract: The present invention relates to a cell for use in producing recombinant adenoviruses (AVs), wherein DNA molecules which are capable of producing antisense RNAs against AAV cap and AAV rep mRNAs are stably integrated into the cell's genome. The invention also relates to processes for the production of such cells and processes for using such cells in the production of recombinant AVs.

Abstract: The invention relates to a nucleic acid molecule encoding at least one AAV Rep polypeptide, wherein one or more of the AAV (p5), (p19) and (p40) promoters have been modified to reduce or eliminate expression of one or more of the Rep polypeptides, or the nucleic acid molecule does not encode functional (Rep52) or (Rep40) polypeptides, or the nucleic acid molecule does not encode a functional adenovirus inhibitor sequence. The invention also relates to a process for producing recombinant AAV vectors through the use of a 2-adenovirus system, wherein all of the genes required for AAV replication and packaging (i.e. an AAV rep sequence of the invention, AAV cap and the AAV transfer vector comprising a transgene) may be encoded within two adenoviruses.

Abstract: Methods of determining the titre of recombinant or wild-type adeno-associated viruses (AAVs) in a sample of recombinant or wild-type AAVs, respectively, are provided. The methods utilise recombinant adenoviruses to amplify the number of AAVs. In some embodiments, the genome of each recombinant adenovirus comprises a rep gene. In some embodiments, the genome of each recombinant adenovirus comprises a repressor element in the Major Late Promoter (MLP).

Abstract: The present invention relates to the production of plasmids which are useful in the production of Adeno-Associated Virus (AAV) particles. In particular, the invention provides nucleic acid molecules comprising capgenes and repgenes, wherein the capand repgenes are both operably-associated with the same promoter. The invention also provides host cells comprising nucleic acid molecules of the invention and methods for their use.

Abstract: The invention relates to a method for promoting the modification, preferably by homology-dependent repair (HDR), of a target site in a genome of a cell. The method comprises the steps of introducing a template DNA molecule and one or more DNA repair inhibitors into a cell which comprises or is capable of expressing a site-specific DNA endonuclease (e.g. Cas12a). The DNA repair inhibitors comprise BAY598, together with one or more other inhibitors. The invention also relates to kits comprising the aforementioned DNA repair inhibitors.

Abstract: The invention relates to a method for promoting the modification, preferably by homology-dependent repair (HDR), of a target site in a genome of a cell. The method comprises the steps of introducing a template DNA molecule and one or more DNA repair inhibitors into a cell which comprises or is capable of expressing a site-specific DNA endonuclease (e.g. Cas9). The DNA repair inhibitors comprise one or more aurora kinase inhibitors, wherein the aurora kinase inhibitors are selected from the group consisting of AT9283, PHA-680632, TAK-901 and CCT137690, together with one or more other inhibitors. The invention also relates to kits comprising the aforementioned DNA repair inhibitors.

Abstract: The present invention relates to nucleic acid molecules comprising viral genes or derivatives thereof for use in the production of retroviral packaging vectors, and retroviral packaging and producer cell lines. In one embodiment, the nucleic acid molecules comprise env and gag-pol genes wherein the coding sequences of the env and gag-pol genes are in opposing orientations.

Abstract: The present invention relates to a method of amplifying a DNA molecule which is operably-linked to a CARE element in a host cell. The method comprises the step of culturing a host cell which comprises a CARE element operably-linked to the DNA molecule, a nucleotide sequence encoding a L4 22K polypeptide or a variant thereof, a nucleic acid molecule comprising a nucleotide sequence encoding an AAV Rep polypeptide or a variant thereof, and optionally one or more further nucleic acid molecules. The invention also relates to nucleic acid molecules encoding a L4 22K polypeptide or a variant thereof, operably-linked to a heterologous promoter; nucleic acid molecules encoding a CARE element operably-linked to viral genes; processes for producing adenoviral vectors and host cells; and processes for producing viral particles, more preferably AAV particles, in host cells.

Abstract: The invention relates to nucleic acid molecules, vectors and plasmids comprising AAV cap genes and rep genes, wherein the cap and rep genes are both operably-associated with an inducible promoter which comprises one or more cumate operator (CuO) sequences. The invention further relates to producer and packaging cell lines which are useful in the production of Adeno-Associated Virus (AAV) particles.

Abstract: The present invention relates to a method for identifying specific binding partners (e.g. antibodies or antibody mimetics) which bind to a desired target polypeptide. In particular, the method involves expressing a library of specific binding partners in a population of mammalian cells, wherein each cell in the population of cells displays the target polypeptide on the outer surface of the cell, and identifying or isolating cells within the population of cells to which specific binding partners are bound.

Abstract: The present invention relates to the production of plasmids which are useful in the production of Adeno-Associated Virus (AAV) particles. In particular, the invention provides nucleic acid molecules comprising capgenes and repgenes, wherein the capand repgenes are both operably-associated with the same promoter. The invention also provides host cells comprising nucleic acid molecules of the invention and methods for their use.

Abstract: The present invention relates to nucleic acid molecules comprising viral genes or derivatives thereof for use in the production of retroviral packaging vectors, and retroviral packaging and producer cell lines. In one embodiment, the nucleic acid molecules comprise env and gag-pol genes wherein the coding sequences of the env and gag-pol genes are in opposing orientations.

Abstract: The present invention relates to an adenoviral vector comprising a regulatable Major Late Promoter and an exogenous transgene. The invention also provides cells comprising such adenoviral vectors, and processes using such vectors.

Abstract: The present invention relates to a viral vector comprising a transposon and a nucleic acid encoding a transposase. The transposon comprises a transgene, or insertion site for a transgene, for integration into the genome of a target cell. The expression of the transposase is controlled such that the transposase is not expressed during production or packaging of the viral vector. Furthermore, the transposon comprises a packaging signal for the virus genome, thus preventing the packaging of any viral genome from which the transposon has been removed. Also provided is a process for producing a modified mammalian cell and a process for producing a mammalian cell with a modified genome, using a viral vector of the invention.

Abstract: The present invention relates to nucleic acid molecules which are capable of promoting transcription of operably-linked heterologous polynucleotides in mammalian cells. The invention also relates to expression vectors and host cells which comprise the nucleic acid molecules of the invention. Such expression vectors may be used to produce recombinant proteins, e.g. antibodies and lentiviral polypeptides.

Abstract: The present invention relates to a method for identifying specific binding partners (e.g. antibodies or antibody mimetics) which bind to a desired target polypeptide. In particular, the method involves expressing a library of antibodies or antibody mimetics in a population of mammalian cells, wherein each cell in the population of cells displays the target polypeptide on the outer surface of the cell, and identifying or isolating cells within the population of cells to which antibodies or antibody mimetics are bound.