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AssigneeThe Scripps Research Institute
InventorsDaniel J. Von Seggern, Glen R. Nemerow, Paul Hallenbeck, Susan Stevenson, Yelena Skripchenko
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The present invention relates to methods for gene therapy, especially to adenovirus-based gene therapy, and related cell lines and compositions. In particular, novel nucleic acid constructs and packaging cell lines are disclosed, for use in facilitating the development of high-capacity and targeted vectors. The invention also discloses a variety of high-capacity adenovirus vectors and related compositions and kits including the disclosed cell lines and vectors. Finally, the invention discloses methods of preparing and using the disclosed vectors, cell lines and kits.
An isolated nucleic acid molecule comprising an adenovirus tripartite leader (TPL) nucleotide, said TPL nucleotide sequence comprising (a) first and second different TPL exons or (b) first, second and third same or different TPL exons, said TPL exons selected from the group consisting of complete TPL exon 1, partial TPL exon 1, complete TPL exon 2 and complete TPL exon 3.
A recombinant adenovirus particle comprising a recombinant adenovirus vector genome wherein said genome does not encode or does not express sufficient adenovirus fiber protein to support packaging of a fiber-containing adenovirus particle without complementation of said fiber gene.
A helper-independent fiberless recombinant adenovirus vector genome comprising genes which:
A method for producing an adenovirus vector particle containing a helper-independent fiberless recombinant adenovirus vector genome, said method comprising providing a packaging cell line which complements replication and packaging of said genome and a helper-independent fiberless recombinant adenovirus vector genome which is deficient in expressing sufficient functional fiber protein to support assembly of fiber-containing particles and harvesting said particle produced by said cell line.
A method for pseudotyping recombinant viral vectors comprising complementing a missing fiber gene of a helper-independent or helper dependent fiberless recombinant adenovirus vector genome by expressing in packaging cells a fiber gene from a different adenoviral serotype than said recombinant adenovirus vector, thereby pseudotyping said vector.
A method for specifically targeting an adenovirus vector to a cell of choice comprising introducing a helper-independent or helper-dependent fiberless recombinant adenovirus vector genome into a packaging cell line for producing a fiber gene-deleted adenovirus vector, wherein a gene for a missing fiber protein is complemented with a gene for a desired modification for targeting the vector to a cell of choice.
A method for producing a modified adenovirus comprising providing in vitro an exogenous fiber protein to a fiberless adenovirus.
A method for delivering a heterologous gene to EBV-infected B cells comprising infecting said B cells with a pseudotyped Ad5βgal.ΔF particle or other fiber-deleted adenovirus particle, said particle having a chimeric fiber including the receptor-binding knob domain of the adenovirus type 3 fiber.
A method for producing a modified adenovirus comprising providing a packaging cell line for producing a fiberless adenovirus helper-dependent fiberless recombinant adenovirus vector genome and a helper virus vector, wherein said cell line complements at least a deficient fiber protein gene, thereby producing the modified adenovirus.
A composition for preparing a therapeutic vector, said composition comprising a plasmid comprising an adenovirus genome lacking a nucleotide sequence encoding a fiber protein or a genome that is incapable of expressing sufficient fiber to result in packaging.
A helper adenovirus particle comprising an adenovirus vector genome that does not encode or does not express sufficient adenovirus fiber protein to support packaging of a fiber-containing adenovirus particle without complementation of said fiber gene, wherein said genome has a mutation in its packaging sequence that renders said genome substantially incapable of being packaged.
A helper adenovirus particle comprising an adenovirus vector genome with recombinase sites flanking its packaging sequence, wherein said vector genome does not encode or does not express sufficient adenovirus fiber protein to support packaging of a fiber-containing adenovirus particle without complementation of said fiber gene.
An adenovirus particle comprising a gutless adenoviral vector genome and a fiberless capsid.
An adenovirus particle comprising a gutless adenoviral vector genome and a capsid comprising a modified fiber protein.
Dependencies are inferred from the claim text.