New AAV Serotypes May Broaden the Therapeutic Pipeline to Human Gene Therapy
Authors: Terence R. Flotte, David A. Williams, Robert M. Frederickson
Journal: Molecular Therapy (2005)

Over the past several years, recombinant adeno-associated virus (rAAV, see [1Carter B.J. Adeno-associated virus and the development of adeno-associated virus vectors: a historical perspective.Mol. Ther. 2004; 10: 981-989Abstract Full Text Full Text PDF PubMed Scopus (107) Google Scholar] for a recent historical review) have been shown to mediate long-lasting gene transfer and expression in a variety of cell types without any reproducible adverse effects. This combination of features suggests that rAAV could be an ideal gene transfer vector for therapy of genetic diseases. However, a number of obstacles have been identified that may limit the clinical application of the prototype strain, AAV serotype 2 (AAV2). Most notable of these has been the relative paucity of AAV2 receptors on certain cell types and the presence of pre-existing immunity to AAV2 in most humans. This pre-existing immunity may have been a factor in limiting the efficacy of rAAV-mediated gene transfer in some clinical trials employing this vector, and may also have primed the development of anti-AAV2 immune responses that limit repeated administration of rAAV vectors. The recent discovery of over one hundred new serotypes and genomic variants of AAV from human and nonhuman primate samples has led to the development of new pseudotyped rAAV vectors that incorporate the basic AAV2 inverted terminal repeat sequences into the vector DNA cassette, yet allow the DNA to be packaged into any of a wide range of different capsids [2Gao G.P. et al.Clades of adeno-associated viruses are widely disseminated in human tissues.J. Virol. 2004; 78: 6381-6388Crossref PubMed Scopus (767) Google Scholar, 3Gao G.P. et al.Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy.Proc. Natl. Acad. Sci. USA. 2002; 99: 11854-11859Crossref PubMed Scopus (1235) Google Scholar]. Each of these new capsids may mediate binding to a different type of receptor, thus broadening the range of cellular targets that may be efficiently transduced by rAAV. Since many of these newly described AAV variants are immunologically distinct as well, rAAV particles pseudotyped with capsids from these new viruses may allow us to avoid pre-existing neutralizing antibodies and other adaptive immune constituents that are directed against AAV2. Two original papers in this issue demonstrate each of these advantages of the new AAV serotypes. In the paper by De et al. [4De B.P. et al.High levels of persistent expression of α1-antitrypsin mediated by the nonhuman primate serotype rh.10 adeno-associated virus despite pre-existing immunity to common human adeno-associated viruses.Mol. Ther. 2006; 13: 67-76Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar], the authors compared twenty-five distinct serotypes of rAAV, each encapsidating an identical α1 antitrypsin (α1AT) vector. α1AT deficiency is an autosomal recessive disorder associated with early onset emphysema. Although gene transfer is a possible strategy to treat α1AT deficiency, high levels of circulating α1AT would be required to protect alveolar structures from destruction by neutrophil elastase, the natural substrate of α1AT. High levels of preexisting immunity to the rAAV vector could compromise this effort. In the new study, identical doses of the various vectors were injected into the pleural space of the lungs of mice, and serum AAT levels were monitored over time. This systematic approach led to the identification of a serotype, AAVrh.10, that was substantially more efficient for systemic AAT expression and secretion from the very large surface area of the pleural space. The AAVrh.10-pseudotyped vector retained the safe and long-lasting expression pattern that is typical of rAAV vectors. Importantly, the authors demonstrated that prior immunization with AAV2 or AAV5 did not interfere with the efficacy of subsequent AAVrh.10-AAT vector dosing, although each serotype did elicit an immune response that was sufficient to block a second dose of a vector of the same serotype. In another study in this issue, by Gao et al. [5Gao G.P. et al.Biology of AAV serotype vectors in liver-directed gene transfer to nonhuman primates.Mol. Ther. 2006; 13: 77-87Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar], the authors present a comparison of the biological efficacy of a number of different AAV serotypes in nonhuman primates. In this study, the vectors were delivered to the livers of a nonhuman primate population in which some animals had pre-existing natural immunity to one or the other of the AAV serotypes. The authors were able to prove that prior natural immunity to an AAV serotype did limit the biological efficacy of vector administration of that particular serotype, but that it did not limit the efficacy of the other serotypes. In this case, they were able to confirm that AAV7 and AAV8 vectors tended to mediate higher levels of liver expression, and also showed that prior host immunity was an important factor dictating which of these two serotypes was more effective in this study. These results may seem somewhat at odds with other recent work indicating a conservation of T-cell epitopes between different AAV serotypes, such as that described in a paper published last month in Molecular Therapy, by Sabatino et al. [6Sabatino D.E. et al.Identification of mouse AAV capsid-specific CD8+ T cell epitopes.Mol. Ther. 2005; 12: 1013-1022Abstract Full Text Full Text PDF PubMed Scopus (74) Google Scholar]. In the latter study, the authors mapped murine CD8+ T-cell epitopes within the AAV2 and AAV8 capsids and identified common epitopes shared between these capsids. There are several important differences between these experiments and those described in the two above-mentioned articles published in this issue of the journal, however, that might account for the divergent results. Clearly, there could be differences between the extent of overlap between humoral immune epitopes and those epitopes involved in cytotoxic T-cell responses. In addition, Sabatino et al. made use of a recombinant adenovirus to express the AAV capsid proteins, which may have increased the probability of the development of CTL responses. This would be consistent with an earlier study in nonhuman primates in which it was shown that the presence of adenovirus was an important factor leading to the development of cell-mediated immune responses to wild-type AAV2 [7Hernandez Y.J. et al.Latent adeno-associated virus infection elicits humoral but not cell-mediated immune responses in a nonhuman primate model.J. Virol. 1999; 73: 8548-8558Google Scholar]. Taken together, these studies point to the great potential of the new AAV serotypes, both to enhance the efficiency of gene delivery and to provide alternatives that will allow one to evade pre-existing AAV immunity. Since many cell types, such as hepatocytes, muscle fibers, and neurons are nondividing, these cells could mediate long-lasting persistence of rAAV genomes without readministration of the vector. Thus, achieving an initial high efficiency of gene transfer and evading immunity on the first dose may be all that is required for life-long expression of the therapeutic transgene. This positive news is further enhanced by the recent development of rAAV pseudotyped vectors based on nonprimate AAVs in which the likelihood of prior immunity in humans is even more remote [8Schmidt M. et al.Cloning and characterization of a bovine adeno-associated virus.J. Virol. 2004; 78: 6509-65136Crossref PubMed Scopus (64) Google Scholar]. It appears likely that the number of potential therapeutic applications for this vector class is likely to increase substantially in the coming years, but the importance of examining the immune responses to these vectors cannot be overemphasized. This issue of Molecular Therapy is the first in the journal's thirteenth volume and seventh year. We are pleased to welcome five new associate editors to the journal: Andy Baker, Christopher Baum, Pedro Lowenstein, Kathy Ponder and Dave Russell. We are confident that the enthusiasm, insight and expertise of these new members will contribute strongly to the continued success of the journal. We would also like to take this occasion to recognize the contributions of the four associate editors whose three-year terms have recently ended: Jeffrey Chamberlain, Markus Grompe, Jeffrey Kordower, and Jon Wolff. We thank these editors for their generous contributions to the journal over the past several years and we look forward to working with them in their new roles on the jounal's editorial board. Here's looking forward to a successful new year!