Improved cardiac gene transfer by transcriptional and transductional targeting of adeno-associated viral vectors.
OBJECTIVE
Vectors based on recombinant adeno-associated virus 2 (AAV-2) are a promising tool for cardiac gene transfer. However, potential therapeutic applications need to consider the predominant transduction of the liver once AAV-2 vectors enter the systemic circulation. We therefore aimed to increase efficiency and specificity of cardiac vector delivery by combining transcriptional and cell surface targeting.


METHODS
For analysis of transcriptional targeting, recombinant AAV vectors were generated harboring a luciferase reporter gene under control of the cytomegalovirus (CMV) promoter or the 1.5-kb cardiac myosin light chain promoter fused to the CMV immediate-early enhancer (CMV(enh)/MLC1.5). Luciferase activities were determined in representative organs three weeks after intravenous injection of the vector into adult mice. Transductional targeting was studied using luciferase-reporter constructs crosspackaged into capsids of AAV serotypes 1 to 6 and modified AAV-2 capsids devoid of binding their primary receptor heparan sulfate proteoglycan.


RESULTS
Intravenous injections of AAV-2 vectors harboring the CMV(enh)/MLC1.5 promoter enabled a specific and 50-fold higher reporter gene expression in left ventricular myocardium of adult mice compared to vectors containing the CMV promoter. Comparison of AAV-2 vector genomes crosspackaged into capsids of AAV-1 to -6 showed that AAV-1, -4, -5, and -6 capsids increased cardiac transduction efficiency by about 10-fold. However, transduction of other organs such as the liver was also increased after systemic administration. In contrast, AAV-2-based vectors with ablated binding to their primary receptor heparan sulfate proteoglycan enabled a significantly increased efficiency of cardiac gene transfer and reduced transduction of the liver.


CONCLUSIONS
Combining transcriptional targeting by the CMV(enh)/MLC1.5 promoter and AAV vectors devoid of binding the AAV-2 primary receptor results in an efficient cardiac gene transfer with a significantly reduced hepatic transduction.
Müller
Oliver J
OJ
Innere Medizin III, Universitätsklinikum Heidelberg, Im Neuenheimer Feld 410, 69120 Heidelberg, Germany. o.mueller@dkfz-heidelberg.de
Leuchs
Barbara
B
Pleger
Sven T
ST
Grimm
Dirk
D
Franz
Wolfgang-M
WM
Katus
Hugo A
HA
Kleinschmidt
Jürgen A
JA
D000818:Animals; D002213:Capsid; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics; D000229:Dependovirus / Q000235:genetics*; D005260:Female; D018390:Gene Targeting / Q000379:methods*; D005818:Genetic Engineering; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000235:genetics; D006331:Heart Diseases / Q000628:therapy; D006493:Heparin / Q000031:analogs & derivatives / Q000235:genetics; D007275:Injections, Intravenous; D008099:Liver / Q000201:enzymology; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D051379:Mice; D008815:Mice, Inbred Strains; D032383:Myocytes, Cardiac / Q000201:enzymology*; D018994:Myosin Light Chains / Q000235:genetics; D011401:Promoter Regions, Genetic; D011509:Proteoglycans / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D014161:Transduction, Genetic / Q000379:methods
D018994:Myosin Light Chains; D011509:Proteoglycans; C036090:heparin proteoglycan; D006493:Heparin; D008156:Luciferases