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Oct 1, 2011 (Vol. 31, No. 17)

The History and Promise of Gene Therapy

A Scientist's View from the Front Lines of Basic and Applied Clinical Research

  • Second-Generation Vector Technology

    Unfortunately, the first-generation vector technologies deployed in these clinical successes still had the limitations described in this article and will be of limited value beyond these test cases. The good news is that the field has been quietly developing the technology of the future, which is now ready for prime time.

    One of the most important limitations of vectors based on murine retroviruses was the need for the target cell to be dividing at the time of exposure to the vector for transduction to occur. This complicates ex vivo applications of stem cells, which often persist in a quiescent state. Furthermore, attempts to induce stem cell division to render them susceptible to retrovirus transduction may irreversibly alter their phenotype essentially losing the capacity for self renewal.

    Development of vectors based on the family of retroviruses called lentiviruses represented a major advance in overcoming this limitation. Lentiviruses are pathogens capable of infecting non-dividing cells; the most notorious is HIV.

    Investigators at the Salk Institute developed replication-defective vectors based on HIV, which demonstrated efficient stable transduction in nondividing cell targets.15

    The first clinical application of lentiviral vectors for a genetic disease deployed an ex vivo bone marrow strategy in a lethal pediatric neurologic disease called adrenoleukodystrophy (ALD). The difference between ALD and the previous work with SCID is that there is no selective advantage of genetically corrected HSCs in ALD as was the case with SCID.

    Successful reconstitution of the ALD patient with corrected cells would require very high transduction efficiency ex vivo and some form of ablation of the recipient’s marrow to make space for the corrected cells. The pilot human study was a dramatic scientific and clinical success.16 Patients were repopulated with a high frequency of transduced cells in virtually all hematopoietic lineages that stabilized their disease.

    The other technological advance was in the development of second-generation AAV vectors for in vivo gene therapy. We hypothesized that natural variation in the AAV capsid that would occur during natural infections may translate into improved vector performance, albeit in a stochastic manner. We sought to identify AAV capsids from viruses that circulate in primates as infectious agents.

    The problem was identifying primates with active AAV infections to obtain samples for virus recovery since the known AAV serotypes were isolated from laboratory stocks of adenoviruses; these AAVs could have been passengers from the initial adenovirus isolations or come from tissue culture components such as bovine serum.

    Serologic studies in primates, however, showed antibodies against some of the known AAVs suggesting they circulate in these populations. Based on previous in vitro work we speculated that AAV may remain as a latent genome following resolution of the infection as is the case with the Herpes simplex virus.

    We tested this hypothesis by analyzing DNA from primate sources using PCR with primers to conserved regions of the known AAV capsids. The results were startling—latent AAV genomes are widely disseminated through many tissues from a wide array of primates, including macaques, great apes, and humans.17,18,19 Sequencing of these genomes demonstrated a marvelous diversity of structure.

    Vectors were created from many of these novel capsids and shown in animal models to be substantially improved over vectors based on the previously known AAVs 1–6. For example, AAV8 shows substantially higher transduction in a number of highly relevant target cells such as liver, muscle, and photoreceptors of the retina.

    A clinical trial of liver-directed gene transfer with AAV8 in subjects with hemophilia showed partial correction of the clotting defect that has replaced the need for some protein replacement and has been stable.20

    AAV9 is a capsid that is capable of transferring its genome across vascular and blood brain barriers and has been shown in preclinical models to treat a variety of cardiac and neurological disorders.21

    Another advantage of the novel AAV isolates we recently discovered, such as AAV8 and AAV9 noted above, is that they present diminished problems of host immunity. They show much lower levels of pre-existing immunity in humans which, if present, can diminish efficacy and are less likely to activate destructive T-cell responses.

  • Next Frontier: Commercialization

    The technical challenges of gene therapy have been overcome with over 30 years of investment in basic and translational research. Success in pilot human experiments has been demonstrated with many exciting new applications to emerge.

    One of the biggest challenges will be to develop business models to encourage the participation of the biotechnology industry in the development of commercial gene therapy products.

    Biotechnology did indeed play a role in the initial development of gene therapy in the 1990s. This occurred in a very different investment climate where promise was as valuable as progress and liquidity could be achieved through IPOs.

    These early companies eventually failed with little subsequent investment due to the fact that the field fell out of favor and venture capital investment in the post-dotcom era changed.

    However, the positive clinical trials and the upswing in public support have led to a re-engagement of the biopharmaceutical industry. Virtually every major venture capital firm and biopharmaceutical company is in the process of re-examining the value of gene therapy products. Concerns that emerge are not related to technical feasibility but rather to uncertainty about the regulatory process and concerns about the business model.

    The fact is that there is no approved gene therapy product in Europe or the U.S.—although this is likely to change in the near future. Challenges regarding the business of gene therapy relate to the point that many of these products will confer long-term effectiveness after a single administration of vector. As one person stated, “How do you price a cure?”

    Will the payers be willing to provide sufficient reimbursement for a single curative treatment that is necessary to justify the investment of development? This is further complicated by the fact that many early models of gene therapy involved orphan diseases.

    I believe that reimbursement will be adequate so long as the treatment addresses an unmet need in a way that dramatically improves the quality of life of those afflicted with the disease. The field of gene therapy is now ready to achieve this goal.

  • References


    1 Muul, L.M., et al., Persistence and expression of the adenosine deaminase gene for 12 years and immune reaction to gene transfer components: long-term results of the first clinical gene therapy trial. Blood, 2003. 101(7): p. 2563-9.

    2 Crystal, R.G., et al., Administration of an adenovirus containing the human CFTR cDNA to the respiratory tract of individuals with cystic fibrosis. Nat Genet, 1994. 8(1): p. 42-51.

    3 Zabner, J., et al., Adenovirus-mediated gene transfer transiently corrects the chloride transport defect in nasal epithelia of patients with cystic fibrosis. Cell, 1993. 75(2): p. 207-16.

    4 Zuckerman, J.B., et al., A phase I study of adenovirus-mediated transfer of the human cystic fibrosis transmembrane conductance regulator gene to a lung segment of individuals with cystic fibrosis. Hum Gene Ther, 1999. 10(18): p. 2973-85.

    5 Yang, Y., et al., Cellular and humoral immune responses to viral antigens create barriers to lung-directed gene therapy with recombinant adenoviruses. J Virol, 1995. 69(4): p. 2004-15.

    6 Berns, K.I., Parvoviridae: The Viruses and Their Replication, in Fields Virology 1996.

    7 Moss, R.B., et al., Repeated aerosolized AAV-CFTR for treatment of cystic fibrosis: a randomized placebo-controlled phase 2B trial. Hum Gene Ther, 2007. 18(8): p. 726-32.

    8 Manno, C.S., et al., Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response. Nat Med, 2006. 12(3): p. 342-7.

    9 Hacein-Bey-Abina, S., et al., Sustained correction of X-linked severe combined immunodeficiency by ex vivo gene therapy. N Engl J Med, 2002. 346(16): p. 1185-93.

    10 Hacein-Bey-Abina, S., et al., LMO2-associated clonal T cell proliferation in two patients after gene therapy for SCID-X1. Science, 2003. 302(5644): p. 415-9.

    11 Acland, G.M., et al., Gene therapy restores vision in a canine model of childhood blindness. Nat Genet, 2001. 28(1): p. 92-5.

    12 Maguire, A.M., et al., Safety and efficacy of gene transfer for Leber’s congenital amaurosis. N Engl J Med, 2008. 358(21): p. 2240-8.

    13 Bainbridge, J.W., et al., Effect of gene therapy on visual function in Leber’s congenital amaurosis. N Engl J Med, 2008. 358(21): p. 2231-9.

    14 Cideciyan, A.V., et al., Human RPE65 gene therapy for Leber congenital amaurosis: persistence of early visual improvements and safety at 1 year. Hum Gene Ther, 2009. 20(9): p. 999-1004.

    15 Naldini, L., et al., In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science, 1996. 272(5259): p. 263-7.

    16 Cartier, N., et al., Hematopoietic stem cell gene therapy with a lentiviral vector in X-linked adrenoleukodystrophy. Science, 2009. 326(5954): p. 818-23.

    17 Gao, G.P., et al., Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy. Proc Natl Acad Sci U S A, 2002. 99(18): p. 11854-9.

    18 Gao, G., et al., Clades of Adeno-associated viruses are widely disseminated in human tissues. J Virol, 2004. 78(12): p. 6381-8.

    19 Gao, G., et al., Adeno-associated viruses undergo substantial evolution in primates during natural infections. Proc Natl Acad Sci U S A, 2003. 100(10): p. 6081-6.

    20 Ponder, K.P., Hemophilia gene therapy: a Holy Grail found. Mol Ther, 2011. 19(3): p. 427-8.

    21 Forsayeth, J.R. and K.S. Bankiewicz, AAV9: Over the Fence and Into the Woods. Mol Ther, 2011. 19(6): p. 1006-7.

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