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Generation of an adenovirus vector lacking E1, e2a, E3, and all of E4 except open reading frame 3
M I Gorziglia1, C Lapcevich, S Roy
1DNA Viral Vector Unit, Genetic Therapy, Inc., a Novartis Company, Gaithersburg, Maryland 20879, USA. mario.gorziglia@pharma.novartis.com
Journal of Virology
|June 11, 1999
Summary
This study introduces a novel adenovirus vector (Av4orf3nBg) with deleted E1, E2a, and most of E4 genes. This improved vector shows comparable gene transfer efficiency, enhanced expression, and reduced liver toxicity in mice for gene therapy.
Area of Science:
- Gene Therapy
- Virology
- Molecular Biology
Background:
- Adenovirus vectors are promising for gene therapy but face challenges with toxicity and immunogenicity due to backbone gene expression.
- Previous strategies involved deleting essential early genes like E1, E2a, E2b, or E4 to improve adenovirus vector safety.
- The role of specific adenovirus E4 open reading frames (ORFs) in vector performance and safety remains an area of investigation.
Purpose of the Study:
- To develop and characterize a novel adenovirus vector (Av4orf3nBg) with deletions in E1, E2a, and all E4 ORFs except ORF3.
- To evaluate the gene transfer efficiency, transgene expression, and toxicity profile of the novel vector compared to a control vector.
- To assess the potential benefits of E4 deletion, specifically retaining only ORF3, for improving liver-directed gene therapy.
Main Methods:
- Construction of the Av4orf3nBg vector lacking E1, E2a, and all E4 except ORF3, expressing a beta-galactosidase reporter gene.
- Production of the vector using a specialized E1-, E2a-, and E4-complementing cell line.
- In vivo studies in C57BL/6 mice to compare gene transfer efficiency, beta-galactosidase expression, and liver toxicity with a control vector (Av3nBg).
Main Results:
- The Av4orf3nBg vector demonstrated gene transfer efficiency comparable to the control Av3nBg vector in mouse liver.
- Av4orf3nBg exhibited significantly higher beta-galactosidase expression, suggesting E4 ORF3 enhances transgene expression.
- Mice injected with Av4orf3nBg showed reduced liver toxicity compared to those receiving the Av3nBg vector at similar DNA copy numbers.
Conclusions:
- Deleting E4, while retaining E1 and E2a, in an adenovirus vector background can enhance safety and reduce liver toxicity.
- The presence of E4 ORF3 alone may potentiate transgene expression, potentially due to enhanced RNA levels.
- This novel vector design holds promise for improving the safety, efficacy, and transgene capacity of adenovirus-based gene therapy, particularly for liver applications.