Methods for producing an adenovirus type 5 gene transfer vector
OwnerRichard L. Atkinson, Suzanne E. Barbour +1inventors
InventorsRichard L. Atkinson, Suzanne E. Barbour, W. Palmer Wilkins
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Abstract
pZerotgCMV, an Ad-5-based expression vector in current use for gene transfer stimulates lipogenic enzymes within HepG2 cells (human hepatic carcinoma cells) and primary rat hepatocytes in vitro. Evidence indicates increased lipid accumulation in infected cells compared to uninfected cells. Therefore, inactivation of the E4 ORF1 gene or E4 gene cluster whether by replacement, removal, mutation, or use of antisense RNA encoded by the Ad-5 genome may prevent activation of lipogenic genes and subsequent lipid accumulation. Removal of just the E4 gene from pZerotgCMV may prevent both replication and stimulation of lipogenic enzymes.
Claims21
A method of preventing an adenovirus type 5 vector from stimulating lipogenic enzymes in a subject requiring gene therapy, said method comprising the step of:
- inactivating an adenovirus type 5 E4 protein by modifying the adenovirus type 5 vector, such that E4 protein is incapable of stimulating lipogenic enzymes in the subject.
A recombinant adenovirus type 5 vector comprising a modified E4 region where a nucleic acid encoding an adenovirus type 5 E4 protein has been replaced with a nucleic acid encoding an E4 protein of adenovirus type 2.
A recombinant adenovirus type 5 vector comprising:
- a precursor RNAi molecule capable of inhibiting the expression of an adenovirus type 5 E4 specific messenger RNA (mRNA) to which it corresponds wherein the RNAi molecule includes a first oligonucleoide strand having a length in a range of about 22 nucleotides to about 30 nucleotides, a second oligonucleotide strand having a length in a range of about 22 nucleotides to about 20 nucleotides, and wherein the nucleotide sequence is complementary to a sequence of an RNA of a target gene to direct target-specific RNAi.
Dependencies are inferred from the claim text.