Adenovirus and method of use thereof
AssigneeThe Trustees of the University of Pennsylvania
InventorsJames M. Wilson, Krishna J. Fisher, Shu-Jen Chen, Matthew Weitzman
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Abstract
A recombinant adenovirus and a method for producing the virus are provided which utilize a recombinant shuttle vector comprising adenovirus DNA sequence for the 5′ and 3′ cis-elements necessary for replication and virion encapsidation in the absence of sequence encoding viral genes and a selected minigene linked thereto, and a helper adenovirus comprising sufficient adenovirus gene sequences necessary for a productive viral infection. Desirably the helper gene is crippled by modifications to its 5′ packaging sequences, which facilitates purification of the viral particle from the helper virus.
Claims12
A method for producing an AdΔ virus which comprises the steps of:
- (a) introducing into a selected host cell:
- (i) a recombinant shuttle vector which comprises adenovirus nucleic acid sequences and a minigene, wherein said adenovirus sequences are of a first serotype and consist of the adenovirus 5′ and 3′ cis-elements necessary for replication and virion encapsidation;
- and wherein said minigene comprises a selected gene operatively linked to regulatory sequences which direct expression of said selected gene in a target cell, said adenovirus cis-elements flanking said minigene, and
- (ii) a helper virus comprising adenovirus gene sequences necessary for a productive viral infection and encoding an adenoviral capsid of a second serotype;
- (b) culturing said host cell containing the shuttle vector and the helper virus so as to permit the formation of an AdΔ virus comprising the adenovirus sequences and minigene of (i) in an adenoviral capsid of a second serotype which differs from said first serotype.
A method for producing an AdΔ virus which comprises the steps of:
- (a) introducing into a selected host cell
- (i) a recombinant shuttle vector which comprises adenovirus nucleic acid sequences and a minigene, wherein said adenovirus sequences are of a first serotype and consist of the adenovirus 5′ and 3′ cis-elements necessary for replication and virion encapsidation;
- and wherein said minigene comprises a selected gene operatively linked to regulatory sequences which direct expression of said selected gene in a target cell, said adenovirus cis-elements flanking said minigene;
- and
- (ii) a helper virus comprising adenovirus gene sequences of a serotype which differ s from the serotype of the adenovirus sequences of (i);
- (b) culturing said host cell containing the shuttle vector and the helper virus, wherein the helper virus and the host cell provide the adenovirus genes necessary for a productive viral infection and which permit the formation of an AdΔ virus comprising the adenovirus sequences and minigene of (i) in an adenoviral capsid of a second serotype which differs from said first serotype.
A recombinant AdΔ virus comprising:
- (a) adenovirus nucleic acid sequences and a minigene, wherein said adenovirus sequences are of a first serotype and consist of the adenovirus 5′ and 3′ cis-elements necessary for replication and virion encapsidation;
- and wherein said minigene comprises a selected gene operatively linked to regulatory sequences which direct expression of said selected gene in a target cell, said adenovirus cis-elements flanking said minigene;
- and
- (b) a capsid of a second adenovirus serotype which differs from said first serotype and which encapsidates said adenovirus sequences and minigene of (a).
A method for producing a pseudotyped AdΔ virus comprising the steps of:
- culturing, under conditions which permit the formation of a pseudotyped AdΔ virus, a host cell containing:
- (i) adenovirus 5′ and 3′ cis-elements necessary for replication and virion encapsidation and a transgene under the control of regulatory elements which control expression thereof in a host cell, wherein said adenovirus 5′ and 3′ cis-elements flank said transgene and are of a first adenovirus serotype;
- (ii) an adenovirus capsid, wherein said adenovirus capsid is of a second adenovirus serotype, which differs from said first adenovirus serotype;
- and
- (iii) adenovirus sequences necessary for a productive viral infection.
Dependencies are inferred from the claim text.