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OwnerAndrew J. Bett, Michael Chastain +6inventors
InventorsAndrew J. Bett, Michael Chastain, Volker Sandig, Emilio A. Emini, John W. Shiver, Danilo R. Casimiro, David C. Kaslow, Manal Morsy
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Various methods for propagating and rescuing multiple serotypes of replication-defective adenovirus in a single adenoviral E1-complementing cell line are disclosed. Typically, replication-defective adenovirus vectors propagate only in cell lines which express E1 proteins of the same serotype or subgroup as the vector. The disclosed methods offer the ability to propagate vectors derived from multiple adenoviral serotypes in a single production cell line which expresses E1 proteins from a single serotype. Propagation in this manner is accomplished by providing all or a portion of an E4 region in cis within the genome of the replication-defective adenovirus. The added E4 region or portion thereof is cloned from a virus of the same or highly similar serotype as that of the E1 gene product(s) of the complementing cell line. Interaction between the expressed E1 of the cell line and the heterologous E4 of the replication-defective adenoviral vectors enables their propagation and rescue. The invention bypasses a need in the art to customize specific cell lines to the serotype or subgroup of the adenoviral vector being propagated and enables one to easily and rapidly develop alternative adenoviral serotypes as gene delivery vectors for use as vaccines or as a critical component in gene therapy.
A means for propagating replication-defective adenovirus in an adenoviral E1-complementing cell line expressing E1 gene product(s) which are non-native to the adenovirus, which comprises:
A replication-defective adenovirus comprising all or a portion of a heterologous E4 region comprising a heterologous adenoviral open reading frame 6 (ORF6).
A replication-defective adenovirus comprising all or a portion of a heterologous E4 region comprising a heterologous adenoviral open reading frame 6 (ORF6) and a gene encoding HIV-1 gag.
A replication-defective adenovirus comprising all or a portion of a heterologous E4 region comprising a heterologous adenoviral open reading frame 6 (ORF6) in place of a native E4 region or portion thereof comprising ORF6.
A replication-defective adenovirus comprising all or a portion of a heterologous E4 region comprising a complete heterologous E4 region in place of a complete native E4 region.
A replication-defective adenovirus comprising a heterologous E4 region or portion thereof comprising a complete heterologous E4 region including E4 promoter in place of a complete native E4 region.
A replication-defective adenovirus of serotype 35 comprising all or a portion of an adenovirus serotype 5 E4 region comprising open reading frame 6 (ORF6) and a heterologous gene of interest.
A replication-defective adenovirus of serotype 35 comprising all or a portion of an adenovirus serotype 5 E4 region comprising open reading frame 6 (ORF6) and a gene encoding HIV-1 gag.
A recombinant adenoviral vector of serotype 24 which comprises an E4 gene or a segment of an E4 gene comprising open reading frame 6 (“ORF6”) of an alternative serotype.
A recombinant adenoviral vector of serotype 24 which is at least partially deleted in E1 and devoid of E1 activity;
56 A recombinant adenoviral vector of serotype 34 which comprises an E4 gene or a segment of an E4 gene comprising open reading frame 6 (“ORF6”) of an alternative serotype.
A population of cells comprising the recombinant adenoviral vector of claim 56.
A method for producing recombinant, replication-defective adenovirus particles comprising:
A recombinant adenoviral vector in accordance with claim 56 which is at least partially deleted in E1 and devoid of E1 activity and comprises a heterologous nucleic acid.
A recombinant adenoviral vector of serotype 34 which is at least partially deleted in E1 and devoid of E1 activity;
Dependencies are inferred from the claim text.