AAV-8 Is More Efficient than AAV-9 in Transducing Neonatal Dog Heart

Abstract

Adeno-associated virus serotype-8 and 9 (AAV-8 and 9) are the leading candidate vectors to test bodywide neonatal muscle gene therapy in large mammals. We have previously shown that systemic injection of 2-2.5 · 10 14 viral genome (vg) particles/kg of AAV-9 resulted in widespread skeletal muscle gene transfer in newborn dogs. However, nominal transduction was observed in the heart. In contrast, robust expression was achieved in both skeletal muscle and heart in neonatal dogs with 7.14-9.06 · 10 14 vg particles/kg of AAV-8. To determine whether superior cardiac transduction of AAV-8 is because of the higher vector dose, we delivered 6.14 · 10 14 and 9.65 · 10 14 vg particles/kg of AAV-9 to newborn puppies via the jugular vein. Transduction was examined 2.5 months later. Consistent with our previous reports, we observed robust bodywide transduction in skeletal muscle. However, increased AAV dose only moderately improved heart transduction. It never reached the level achieved by AAV-8. Our results suggest that differential cardiac transduction by AAV-8 and AAV-9 is likely because of the intrinsic property of the viral capsid rather than the vector dose. IntroductionO ver the last two decades, adeno-associated virus (AAV) has become a leading vector for gene therapy. AAV is a single-stranded DNA virus discovered in mid1960s. 1 More than 100 AAV variants are now available for gene transfer studies. 2,3 These AAV variants are either isolated from natural resources (such as adenovirus stocks and animal tissues) or engineered in the laboratory by rational design and/or evolution. While all AAV variants have a similar icosahedral capsid, the difference in the amino acid composition has yielded distinctive biological properties that are now been capitalized for different gene therapy applications. Among these, the ability to escape from the vasculature while remaining transduction competent is particularly attractive for treating diseases like Duchenne muscular dystrophy (DMD). Muscle makes up *40% of the body mass and it is distributed all over the body. An effective therapy for DMD will require an efficient delivery of a therapeutic gene to the whole body.The breakthrough in systemic gene delivery was made about a decade ago. [4][5][6][7][8] Studies from several laboratories show that a single intravenous injection of AAV-6, 8, or 9 leads to sustained whole-body gene transfer in rodents. Subsequent studies in mouse models of DMD and other types of muscular dystrophies revealed excellent bodywide muscle transduction and disease amelioration. [9][10][11][12][13][14] We conducted the first systemic AAV gene delivery in a large mammal in 2008. 15 In the study, we delivered 1-2.5 · 10 14 viral genome (vg)/kg of AAV-9 to newborn dogs. We picked AAV-9 because this serotype worked extremely well in the rodent heart. We reasoned that the cardiac tropism would help to treat cardiomyopathy, a lethal complication of muscular dystrophy. Although robust bodywide skeletal muscle transduction was observed, surprisingly, very ...

Systemic delivery of adeno-associated viral vectors

“…Surprisingly, despite spectacular bodywide skeletal muscle transduction, few cardiomyocytes were transduced [66]. In sharp contrast, AAV-8 yielded robust transduction of both skeletal and cardiac muscles in dog puppies (Figure 1) [67,68]. AAV-1 and AAV-6 are two other serotypes that have shown good systemic transduction in rodents [26,30,31].…”

Viral vector-mediated gene therapies

“…However, considerable emerging evidence from (mostly unpublished) animal and human trials of various disorders using AAV vectors suggest that systemic gene delivery to striated muscles might be achieved using simple infusion of a high dose of vector delivered through single or several veins or arteries [e.g. (10, 18)].…”

“…These intra-species differences make it difficult to predict the optimal vector for clinical application. For example AAV9 was able to transfect rodent hearts well but in neonatal dogs AAV8 achieved a higher transfection rate (10). Similarly, while AAV6 displays better efficiency in rodent striated muscles and in canine cardiac muscle, AAV9 appears to work better in adult canine skeletal muscles (Seto, Ramos et al, in preparation).…”

Nanotherapy for Duchenne muscular dystrophy

“…Third, intravenous delivery by AAV‐8 and ‐9 leads to high‐level expression in the liver in mice . However, there is minimal expression in the liver of dogs (even with a ubiquitous promoter and a reporter gene) although a significant amount of the AAV genome is detected in the liver . Finally, a study from the Xiao lab suggests that systemic injection of AAV‐9 may result in a massive inflammatory response under certain condition (the ubiquitous promoter, human transgene, dystrophic puppy, or vector stock impurity) …”