Preliminary evaluation of a self-complementary AAV2/8 vector for hepatic gene transfer of human apoE3 to inhibit atherosclerotic lesion development in apoE-deficient mice.

Hepatic gene transfer of atheroprotective human apoE by recombinant viral vectors can reverse hypercholesterolaemia and inhibit atherogenesis in apoE-deficient (apoE(-/-)) mice. Here, in preliminary studies we assess the effectiveness of a recently developed self-complementary adeno-associated virus (scAAV) serotype 8 vector, driven by a hepatocyte-specific promoter (LP1), for liver-directed gene delivery of human apoE3. Vector viability was validated by transducing cultured HepG2 cells and measuring secretion of apoE3 protein. Male and female apoE(-/-) mice, 6-month old and fed on normal chow, were intravenously injected with 1x10(11) vg (vector genomes) of scAAV2/8.LP1.apoE3; age-matched untreated mice served as controls. In male mice, plasma apoE3 levels were sufficiently high (up to 17 microg/ml) to normalize plasma total cholesterol and ameliorate their proatherogenic lipoprotein profile, by reducing VLDL/LDL and increasing HDL 5-fold. At termination (12 weeks) development of aortic atherosclerosis was significantly retarded by 58% (aortic lesion area 8.2+/-1.4% vs. 19.3+/-2.4% in control males; P<0.001). Qualitatively similar anti-atherogenic effects were noted when female mice were treated, but the benefits were less marked and aortic lesions, for example, were reduced by only 33% (15.7+/-3.7% vs. 23.6+/-6.9%). Although group numbers were small (n=4/5), this gender-specific difference reflected two to three times less apoE3 in plasma of female mice at weeks 3 and 6, implying that gene transfer to female liver using scAAV vectors may require additional optimization, despite their established superior potency to conventional single-stranded (ssAAV) vectors.

Adenosine monophosphate-activated protein kinase disease mimicks hypertrophic cardiomyopathy and Wolff-Parkinson-White syndrome: natural history.

OBJECTIVES: The aim of this study was to investigate the clinical expression of adenosine monophosphate-activated protein kinase (AMPK) gene mutations (PRKAG2) in adenosine monophosphate (AMP) kinase disease based on 12 years follow-up of known mutation carriers and to define the prevalence of PRKAG2 mutations in hypertrophic cardiomyopathy (HCM). BACKGROUND: Adenosine monophosphate-activated protein kinase gene mutations cause HCM with Wolff-Parkinson-White syndrome and conduction disease. METHODS: Clinical evaluation of 44 patients with known AMP kinase disease was analyzed. Mutation analysis of PRKAG2 was performed by fluorescent single-strand confirmation polymorphism analysis and direct sequencing of abnormal conformers in 200 patients with HCM. RESULTS: Only one additional mutation was identified. The mean age at clinical diagnosis in the 45 gene carriers was 24 years (median 20 years, range 9 to 55 years). Symptoms of palpitation, dypspnea, chest pain, or syncope were present in 31 (69%) gene carriers; 7 (15%) complained of myalgia and had clinical evidence of proximal myopathy. Skeletal muscle biopsy showed excess mitochondria and ragged red fibers with minimal glycogen accumulation. Disease penetrance defined by typical electrocardiogram abnormalities was 100% by age 18 years. Thirty-two of 41 adults (78%) had left ventricular hypertrophy (LVH) on echocardiography, and progressive LVH was documented during follow-up. Survival was 91% at a mean follow-up of 12.2 years. Progressive conduction disease required pacemaker implantation in 17 of 45 (38%) at a mean age of 38 years. CONCLUSIONS: The AMP kinase disease is uncommon in HCM and is characterized by progressive conduction disease and cardiac hypertrophy and includes extracardiac manifestations such as a skeletal myopathy, consistent with a systemic metabolic storage disease. Defects in adenosine triphosphate utilization or in specific cellular substrates, rather than mere passive deposition of amylopectin, may account for these clinical features.