Cardio-specific long-term gene expression in a porcine model after selective pressure-regulated retroinfusion of adeno-associated viral (AAV) vectors.
Cornerstone for an efficient cardiac gene therapy is the need for a vector system, which enables selective and long-term expression of the gene of interest. In rodent animal models adeno-associated viral (AAV) vectors like AAV-6 have been shown to efficiently transduce cardiomyocytes. However, since significant species-dependent differences in transduction characteristics exist, large animal models are of imminent need for preclinical evaluations. We compared gene transfer efficiencies of AAV-6 and heparin binding site-deleted AAV-2 vectors in a porcine model. Application of the AAVs was performed by pressure-regulated retroinfusion of the anterior interventricular cardiac vein, which has been previously shown to efficiently deliver genes to the myocardium (3.5 x 10(10) viral genomes per animal; n=5 animals per group). All vectors harbored a luciferase reporter gene under control of a cytomegalovirus (CMV)-enhanced 1.5 kb rat myosin light chain promoter (CMV-MLC2v). Expression levels were evaluated 4 weeks after gene transfer by determining luciferase activities. To rule out a systemic spillover peripheral tissue was analyzed by PCR for the presence of vector genomes. Selective retroinfusion of AAV serotype 6 vectors into the anterior cardiac vein substantially increased reporter gene expression in the targeted distal left anterior descending (LAD) territory (65 943+/-31 122 vs control territory 294+/-69, P<0.05). Retroinfusion of AAV-2 vectors showed lower transgene expression, which could be increased with coadministration of recombinant human vascular endothelial growth factor (1365+/-707 no vascular endothelial growth factor (VEGF) vs 38 760+/-2448 with VEGF, P<0.05). Significant transgene expression was not detected in other organs than the heart, although vector genomes were detected also in the lung and liver. Thus, selective retroinfusion of AAV-6 into the coronary vein led to efficient long-term myocardial reporter gene expression in the targeted LAD area of the porcine heart. Coapplication of VEGF significantly increased transduction efficiency of AAV-2.
Raake
P W
PW
Department of Internal Medicine I, Grosshadern University Hospital, Ludwig-Maximilians-University, Munich, Germany.
Hinkel
R
R
Müller
S
S
Delker
S
S
Kreuzpointner
R
R
Kupatt
C
C
Katus
H A
HA
Kleinschmidt
J A
JA
Boekstegers
P
P
Müller
O J
OJ
D000818:Animals; D003331:Coronary Vessels; D000229:Dependovirus / Q000235:genetics*; D017353:Gene Deletion; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D006331:Heart Diseases / Q000628:therapy*; D006493:Heparin / Q000031:analogs & derivatives / Q000235:genetics; D007262:Infusions, Intravenous / Q000379:methods; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D023421:Models, Animal; D009206:Myocardium / Q000201:enzymology; D011312:Pressure; D011509:Proteoglycans / Q000235:genetics; D013552:Swine; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics
D011509:Proteoglycans; D042461:Vascular Endothelial Growth Factor A; C036090:heparin proteoglycan; D006493:Heparin; D008156:Luciferases