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AAV serotype-dependent apolipoprotein A-I Milano gene expression.
Behrooz G Sharifi1, Kaijin Wu, Lai Wang
1Division of Cardiology, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA.
Atherosclerosis
|July 26, 2005
Summary
Gene therapy using adeno-associated virus (AAV) vectors shows promise for delivering apolipoprotein A-I Milano (ApoA-I Milano) to treat atherosclerosis. Optimizing AAV serotypes and promoters enhances ApoA-I Milano expression for potential therapeutic benefits.
Area of Science:
- Gene Therapy
- Cardiovascular Research
- Molecular Biology
Background:
- Apolipoprotein A-I Milano (ApoA-I Milano) complex with phospholipids demonstrated significant coronary atheroma regression in acute coronary syndromes.
- Previous studies confirmed atherosclerosis regression in an animal model.
- A viral vector-based gene delivery system is explored for ApoA-I Milano gene therapy.
Purpose of the Study:
- To develop and optimize a gene delivery system for ApoA-I Milano using adeno-associated virus (AAV) vectors.
- To evaluate the efficacy of different AAV serotypes and promoter constructs for ApoA-I Milano expression in vivo.
- To assess the impact of delivery routes (intravenous vs. intramuscular) on gene expression levels and duration.
Main Methods:
- Recombinant serotype 2 adeno-associated virus (rAAV2) vectors were engineered with ApoA-I Milano or enhanced green fluorescent protein (EGFP) genes.
- Combinations of cytomegalovirus (CMV) promoters were tested to identify optimal constructs for gene expression.
- In vivo studies compared intravenous (IV) and intramuscular (IM) delivery routes using optimized rAAV2 constructs and different AAV serotypes (rAAV1, rAAV2, rAAV5).
Main Results:
- A promoter construct with two CMV core promoters and a CMV enhancer showed higher activity than single CMV promoters.
- Intravenous (IV) delivery of rAAV2 resulted in high circulating ApoA-I Milano levels for extended periods (up to 220 ng/ml at 22 weeks), while intramuscular (IM) delivery yielded low, transient expression.
- Among IM delivery routes, rAAV1 demonstrated significantly higher ApoA-I Milano expression (15-fold higher than rAAV2, 9-fold higher than rAAV5), while IV delivery with all tested serotypes produced substantial expression.
Conclusions:
- Modifications to AAV promoter strength and serotype can significantly enhance transgene expression efficiency.
- Optimized AAV vectors, particularly with specific serotypes and IV delivery, hold potential for effective ApoA-I Milano gene therapy.
- These findings support the advancement of AAV-mediated gene therapy for cardiovascular diseases like atherosclerosis.