Muscular Dystrophy Gene Therapy in Small Animal Models

  • Chunping Qiao
  • Xiao XiaoEmail author


Muscular dystrophies are inherited neuromuscular disorders characterized by progressive muscle loss and weakness. The morbidity and fatality associated with the diseases and a lack of effective treatment have prompted urgent search for novel therapeutics. Gene therapy is one of the frontiers. Currently, adeno-associated viral (AAV) vector-mediated gene transfer offers a powerful tool for muscular dystrophy gene therapy for both skeletal as well as cardiac muscles, by means of local, regional, and systemic deliveries. However, AAV has a packaging limit smaller than 5,000 nucleotides. Larger genes such as dystrophin will need to be truncated to functional miniature versions to be packaged in AAV particles. In this chapter, we will illustrate how gene therapy with AAV vectors is applied to small rodent muscular dystrophy models including those that mimic Duchenne muscular dystrophy (the dystrophin-deficient mdx mice), congenital muscular dystrophy (the laminin α2 knockout dy/dy mice), and limb-girdle muscular dystrophy (the delta-sarcoglycan deficient TO-2 hamsters). Challenges in larger animal studies and prospects for clinical trials in muscular dystrophies will be briefly discussed.


Muscular Dystrophy Duchenne Muscular Dystrophy Duchenne Muscular Dystrophy Dystrophin Gene Congenital Muscular Dystrophy 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



Most of the work was funded by NIH.


  1. Allamand, V., and Guicheney, P. (2002). Merosin-deficient congenital muscular dystrophy, autosomal recessive (MDC1A, MIM#156225, LAMA2 gene coding for alpha2 chain of laminin). Eur J Hum Genet 10, 91–94.CrossRefPubMedGoogle Scholar
  2. Athanasopoulos, T., Graham, I. R., Foster, H., and Dickson, G. (2004). Recombinant adeno-associated viral (rAAV) vectors as therapeutic tools for Duchenne muscular dystrophy (DMD). Gene Ther 11 Suppl 1, S109–121.CrossRefGoogle Scholar
  3. Berns, K. I., and Giraud, C. (1995). Adenovirus and adeno-associated virus as vectors for gene therapy. Ann N Y Acad Sci 772, 95–104.CrossRefPubMedGoogle Scholar
  4. Bezakova, G., and Ruegg, M. A. (2003). New insights into the roles of agrin. Nat Rev Mol Cell Biol 4, 295–308.CrossRefPubMedGoogle Scholar
  5. Blankinship, M. J., Gregorevic, P., and Chamberlain, J. S. (2006). Gene therapy strategies for duchenne muscular dystrophy utilizing recombinant adeno-associated virus vectors. Mol Ther 13, 241–249.CrossRefPubMedGoogle Scholar
  6. Chamberlain, J. S. (2002). Gene therapy of muscular dystrophy. Hum Mol Genet 11, 2355–2362.CrossRefPubMedGoogle Scholar
  7. Chao, H., Sun, L., Bruce, A., Xiao, X., and Walsh, C. E. (2002). Expression of human factor VIII by splicing between dimerized AAV vectors. Mol Ther 5, 716–722.CrossRefPubMedGoogle Scholar
  8. Cordier, L., Hack, A. A., Scott, M. O., Barton-Davis, E. R., Gao, G., Wilson, J. M., McNally, E. M., and Sweeney, H. L. (2000). Rescue of skeletal muscles of gamma-sarcoglycan-deficient mice with adeno-associated virus-mediated gene transfer. Mol Ther 1, 119–129.CrossRefPubMedGoogle Scholar
  9. Cox, G. A., Cole, N. M., Matsumura, K., Phelps, S. F., Hauschka, S. D., Campbell, K. P., Faulkner, J. A., and Chamberlain, J. S. (1993). Overexpression of dystrophin in transgenic mdx mice eliminates dystrophic symptoms without toxicity. Nature 364, 725–729.CrossRefPubMedGoogle Scholar
  10. Dressman, D., Araishi, K., Imamura, M., Sasaoka, T., Liu, L. A., Engvall, E., and Hoffman, E. P. (2002). Delivery of alpha- and beta-sarcoglycan by recombinant adeno-associated virus: efficient rescue of muscle, but differential toxicity. Hum Gene Ther 13, 1631–1646.CrossRefPubMedGoogle Scholar
  11. Duan, D., Yue, Y., Yan, Z., and Engelhardt, J. F. (2000). A new dual-vector approach to enhance recombinant adeno-associated virus-mediated gene expression through intermolecular cis activation. Nat Med 6, 595–598.CrossRefPubMedGoogle Scholar
  12. Emery, A. E. (2002). The muscular dystrophies. Lancet 359, 687–695.CrossRefPubMedGoogle Scholar
  13. England, S. B., Nicholson, L. V., Johnson, M. A., Forrest, S. M., Love, D. R., Zubrzycka-Gaarn, E. E., Bulman, D. E., Harris, J. B., and Davies, K. E. (1990). Very mild muscular dystrophy associated with the deletion of 46% of dystrophin. Nature 343, 180–182.CrossRefPubMedGoogle Scholar
  14. Fabb, S. A., Wells, D. J., Serpente, P., and Dickson, G. (2002). Adeno-associated virus vector gene transfer and sarcolemmal expression of a 144 kDa micro-dystrophin effectively restores the dystrophin-associated protein complex and inhibits myofibre degeneration in nude/mdx mice. Hum Mol Genet 11, 733–741.CrossRefPubMedGoogle Scholar
  15. Flotte, T. R. (2005). Adeno-associated virus-based gene therapy for inherited disorders. Pediatr Res 58, 1143–1147.CrossRefPubMedGoogle Scholar
  16. Gao, G. P., Alvira, M. R., Wang, L., Calcedo, R., Johnston, J., and Wilson, J. M. (2002). Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy. Proc Natl Acad Sci U S A 99, 11854–11859.CrossRefPubMedGoogle Scholar
  17. Grieger, J. C., and Samulski, R. J. (2005). Adeno-associated virus as a gene therapy vector: vector development, production and clinical applications. Adv Biochem Eng Biotechnol 99, 119–145.PubMedGoogle Scholar
  18. Hammermann, M., Brun, N., Klenin, K. V., May, R., Toth, K., and Langowski, J. (1998). Salt-dependent DNA superhelix diameter studied by small angle neutron scattering measurements and Monte Carlo simulations. Biophys J 75, 3057–3063.CrossRefPubMedGoogle Scholar
  19. Harper, S. Q., Hauser, M. A., DelloRusso, C., Duan, D., Crawford, R. W., Phelps, S. F., Harper, H. A., Robinson, A. S., Engelhardt, J. F., Brooks, S. V., and Chamberlain, J. S. (2002). Modular flexibility of dystrophin: implications for gene therapy of Duchenne muscular dystrophy. Nat Med 8, 253–261.CrossRefPubMedGoogle Scholar
  20. Hoffman, E. P. (1999). Counting muscular dystrophies in the post-molecular census. J Neurol Sci 164, 3–6.CrossRefPubMedGoogle Scholar
  21. Hoffman, E. P., Brown, R. H., Jr., and Kunkel, L. M. (1987). Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell 51, 919–928.CrossRefPubMedGoogle Scholar
  22. Jung, D., Duclos, F., Apostol, B., Straub, V., Lee, J. C., Allamand, V., Venzke, D. P., Sunada, Y., Moomaw, C. R., Leveille, C. J., et al. (1996). Characterization of delta-sarcoglycan, a novel component of the oligomeric sarcoglycan complex involved in limb-girdle muscular dystrophy. J Biol Chem 271, 32321–32329.CrossRefPubMedGoogle Scholar
  23. Kaemmerer, W. F., Reddy, R. G., Warlick, C. A., Hartung, S. D., McIvor, R. S., and Low, W. C. (2000). In vivo transduction of cerebellar Purkinje cells using adeno-associated virus vectors. Mol Ther 2, 446–457.CrossRefPubMedGoogle Scholar
  24. Kawada, T., Nakazawa, M., Nakauchi, S., Yamazaki, K., Shimamoto, R., Urabe, M., Nakata, J., Hemmi, C., Masui, F., Nakajima, T., et al. (2002). Rescue of hereditary form of dilated cardiomyopathy by rAAV-mediated somatic gene therapy: amelioration of morphological findings, sarcolemmal permeability, cardiac performances, and the prognosis of TO-2 hamsters. Proc Natl Acad Sci U S A 99, 901–906.CrossRefPubMedGoogle Scholar
  25. Kessler, P. D., Podsakoff, G. M., Chen, X., McQuiston, S. A., Colosi, P. C., Matelis, L. A., Kurtzman, G. J., and Byrne, B. J. (1996). Gene delivery to skeletal muscle results in sustained expression and systemic delivery of a therapeutic protein. Proc Natl Acad Sci U S A 93, 14082–14087.CrossRefPubMedGoogle Scholar
  26. Koenig, M., Hoffman, E. P., Bertelson, C. J., Monaco, A. P., Feener, C., and Kunkel, L. M. (1987). Complete cloning of the Duchenne muscular dystrophy (DMD) cDNA and preliminary genomic organization of the DMD gene in normal and affected individuals. Cell 50, 509–517.CrossRefPubMedGoogle Scholar
  27. Koenig, M., and Kunkel, L. M. (1990). Detailed analysis of the repeat domain of dystrophin reveals four potential hinge segments that may confer flexibility. J Biol Chem 265, 4560–4566.PubMedGoogle Scholar
  28. Li, J., Dressman, D., Tsao, Y. P., Sakamoto, A., Hoffman, E. P., and Xiao, X. (1999). rAAV vector-mediated sarcogylcan gene transfer in a hamster model for limb girdle muscular dystrophy. Gene Ther 6, 74–82.CrossRefPubMedGoogle Scholar
  29. Li, J., Wang, D., Qian, S., Chen, Z., Zhu, T., and Xiao, X. (2003). Efficient and long-term intracardiac gene transfer in delta-sarcoglycan-deficiency hamster by adeno-associated virus-2 vectors. Gene Ther 10, 1807–1813.CrossRefPubMedGoogle Scholar
  30. Lim, L. E., and Campbell, K. P. (1998). The sarcoglycan complex in limb-girdle muscular dystrophy. Curr Opin Neurol 11, 443–452.CrossRefPubMedGoogle Scholar
  31. Lu, H., Chen, L., Wang, J., Huack, B., Sarkar, R., Zhou, S., Xu, R., Ding, Q., Wang, X., Wang, H., and Xiao, W. (2008). Complete correction of hemophilia A with adeno-associated viral vectors containing a full-size expression cassette. Hum Gene Ther 19, 648–654.CrossRefPubMedGoogle Scholar
  32. McCabe, E. R., Towbin, J., Chamberlain, J., Baumbach, L., Witkowski, J., van Ommen, G. J., Koenig, M., Kunkel, L. M., and Seltzer, W. K. (1989). Complementary DNA probes for the Duchenne muscular dystrophy locus demonstrate a previously undetectable deletion in a patient with dystrophic myopathy, glycerol kinase deficiency, and congenital adrenal hypoplasia. J Clin Invest 83, 95–99.CrossRefPubMedGoogle Scholar
  33. McCarty, D. M., Monahan, P. E., and Samulski, R. J. (2001). Self-complementary recombinant adeno-associated virus (scAAV) vectors promote efficient transduction independently of DNA synthesis. Gene Ther 8, 1248–1254.CrossRefPubMedGoogle Scholar
  34. McCarty, D. M., Young, S. M., Jr., and Samulski, R. J. (2004). Integration of adeno-associated virus (AAV) and recombinant AAV vectors. Annu Rev Genet 38, 819-845.CrossRefPubMedGoogle Scholar
  35. Meinen, S., and Ruegg, M. A. (2006). Congenital muscular dystrophy: Mini-agrin delivers in mice. Gene Ther 13, 869–70.PubMedGoogle Scholar
  36. Moll, J., Barzaghi, P., Lin, S., Bezakova, G., Lochmuller, H., Engvall, E., Muller, U., and Ruegg, M. A. (2001). An agrin minigene rescues dystrophic symptoms in a mouse model for congenital muscular dystrophy. Nature 413, 302–307.CrossRefPubMedGoogle Scholar
  37. Muzyczka, N. (1992). Use of adeno-associated virus as a general transduction vector for mammalian cells. Curr Top Microbiol Immunol 158, 97–129.PubMedGoogle Scholar
  38. Pacak, C. A., Walter, G. A., Gaidosh, G., Bryant, N., Lewis, M. A., Germain, S., Mah, C. S., Campbell, K. P., and Byrne, B. J. (2007). Long-term skeletal muscle protection after gene transfer in a mouse model of LGMD-2D. Mol Ther 15, 1775–1781.CrossRefPubMedGoogle Scholar
  39. Piccolo, F., Roberds, S. L., Jeanpierre, M., Leturcq, F., Azibi, K., Beldjord, C., Carrie, A., Recan, D., Chaouch, M., Reghis, A., and et al. (1995). Primary adhalinopathy: a common cause of autosomal recessive muscular dystrophy of variable severity. Nat Genet 10, 243–245.CrossRefPubMedGoogle Scholar
  40. Qiao, C., Li, J., Zhu, T., Draviam, R., Watkins, S., Ye, X., Chen, C., Li, J., and Xiao, X. (2005). Amelioration of laminin-{alpha}2-deficient congenital muscular dystrophy by somatic gene transfer of miniagrin. Proc Natl Acad Sci U S A 102, 11999–2004.CrossRefPubMedGoogle Scholar
  41. Rafael, J. A., Cox, G. A., Corrado, K., Jung, D., Campbell, K. P., and Chamberlain, J. S. (1996). Forced expression of dystrophin deletion constructs reveals structure-function correlations. J Cell Biol 134, 93–102.CrossRefPubMedGoogle Scholar
  42. Ragot, T., Vincent, N., Chafey, P., Vigne, E., Gilgenkrantz, H., Couton, D., Cartaud, J., Briand, P., Kaplan, J. C., Perricaudet, M., and et al. (1993). Efficient adenovirus-mediated transfer of a human minidystrophin gene to skeletal muscle of mdx mice. Nature 361, 647–650.CrossRefPubMedGoogle Scholar
  43. Sakamoto, M., Yuasa, K., Yoshimura, M., Yokota, T., Ikemoto, T., Suzuki, M., Dickson, G., Miyagoe-Suzuki, Y., and Takeda, S. (2002). Micro-dystrophin cDNA ameliorates dystrophic phenotypes when introduced into mdx mice as a transgene. Biochem Biophys Res Commun 293, 1265–1272.CrossRefPubMedGoogle Scholar
  44. Sun, L., Li, J., and Xiao, X. (2000). Overcoming adeno-associated virus vector size limitation through viral DNA heterodimerization. Nat Med 6, 599–602.CrossRefPubMedGoogle Scholar
  45. Wang, B., Li, J., and Xiao, X. (2000). Adeno-associated virus vector carrying human minidystrophin genes effectively ameliorates muscular dystrophy in mdx mouse model. Proc Natl Acad Sci U S A 97, 13714–13719.CrossRefPubMedGoogle Scholar
  46. Wang, Z., Allen, J. M., Riddell, S. R., Gregorevic, P., Storb, R., Tapscott, S. J., Chamberlain, J. S., and Kuhr, C. S. (2007a). Immunity to adeno-associated virus-mediated gene transfer in a random-bred canine model of Duchenne muscular dystrophy. Hum Gene Ther 18, 18–26.CrossRefPubMedGoogle Scholar
  47. Wang, Z., Kuhr, C. S., Allen, J. M., Blankinship, M., Gregorevic, P., Chamberlain, J. S., Tapscott, S. J., and Storb, R. (2007b). Sustained AAV-mediated dystrophin expression in a canine model of Duchenne muscular dystrophy with a brief course of immunosuppression. Mol Ther 15, 1160–1166.PubMedGoogle Scholar
  48. Wang, Z., Ma, H. I., Li, J., Sun, L., Zhang, J., and Xiao, X. (2003). Rapid and highly efficient transduction by double-stranded adeno-associated virus vectors in vitro and in vivo. Gene Ther 10, 2105–2111.CrossRefPubMedGoogle Scholar
  49. Wang, Z., Zhu, T., Qiao, C., Zhou, L., Wang, B., Zhang, J., Chen, C., Li, J., and Xiao, X. (2005). Adeno-associated virus serotype 8 efficiently delivers genes to muscle and heart. Nat Biotechnol 23, 321–328.CrossRefPubMedGoogle Scholar
  50. Xiao, X., Li, J., and Samulski, R. J. (1996). Efficient long-term gene transfer into muscle tissue of immunocompetent mice by adeno-associated virus vector. J Virol 70, 8098–8108.PubMedGoogle Scholar
  51. Xiao, X., Li, J., Tsao, Y. P., Dressman, D., Hoffman, E. P., and Watchko, J. F. (2000). Full functional rescue of a complete muscle (TA) in dystrophic hamsters by adeno-associated virus vector-directed gene therapy. J Virol 74, 1436–1442.CrossRefPubMedGoogle Scholar
  52. Yoshida, K., Yoshimoto, M., Sasaki, K., Ohnishi, T., Ushiki, T., Hitomi, J., Yamamoto, S., and Sigeno, M. (1998). Fabrication of a new substrate for atomic force microscopic observation of DNA molecules from an ultrasmooth sapphire plate. Biophys J 74, 1654–1657.CrossRefPubMedGoogle Scholar
  53. Zhu, T., Zhou, L., Mori, S., Wang, Z., McTiernan, C. F., Qiao, C., Chen, C., Wang, D. W., Li, J., and Xiao, X. (2005). Sustained whole-body functional rescue in congestive heart failure and muscular dystrophy hamsters by systemic gene transfer. Circulation 112, 2650–2659.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Fred N. Eshelman Distinguished Professor of Gene Therapy, Division of Molecular PharmaceuticsUNC School of PharmacyChapel HillUSA

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