Gene therapy for progressive muscular dystrophy using a new generation adenovirus vector and transposase

• Project/Area Number: 18590951
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Neurology
• Research Institution: Kumamoto University
• Principal Investigator: UCHINO Makoto Kumamoto University, Department of Neurology, Graduate School of Medical Sciences, Professor (20117336)
• Co-Investigator(Kenkyū-buntansha): MAEDA Yasushi Kumamoto University, Department of Neurology, Kumamoto University Hospital, assistant Professor (60346997)
• Project Period (FY): 2006 – 2007
• Project Status: Completed (Fiscal Year 2007)
• Budget Amount: ¥4,010,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥510,000); Fiscal Year 2007: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000); Fiscal Year 2006: ¥1,800,000 (Direct Cost: ¥1,800,000)
• Keywords: gene therapy / helper-dependent adenovirus vector / full-length dystrophin / Duchenne muscular dystrophy / mdx mouse / dko mouse / sleeping beautv-transoosone system / nNOS

Research Abstract

Duchenne muscular dystrophy(DMD) is a fatal progressive muscle wasting disease caused by defects in the dystrophin gene. No viral vector except the helper-dependent adenovirus vector(HDAdv) can package 14kb full-length dystrophin cDNA and HDAdv is considerably safer than old-generation adenovirus vectors due to the large-size deletion in its genome. We have generated HDAdv that carries myc-tagged murine full-length dystrophin cDNA(HDAdv-myc-mFLdys). We injected it into the multiple proximal muscles of 7-day-old utrophin/dystrophin double knockout mice (dko mice), which typically show symptoms quite similar to human DMD because the proximal muscles are organs affected in DMD patients. Eight weeks after injections, the transduced dystrophin was widely expressed and we found a significant reduction of centrally nucleated myofibers and the restoration of dystrophin associated proteins, p-dystroglycan (p-DG) and a-sarcoglycan (a-SG), as well as neuronal nitric oxide synthase. (nNOS). The injected dko mice also showed an increase in body weight, an improvement in motor performances, and prolonged lifespan. Using HDAdv, we could treat DMD model mice, even when the therapeutic gene was transferred into multiple skeletal muscles. Our results suggest that multiple intramuscular administrations of HDAdv carrying full-length dystrophin may reduce symptoms and compensate for lost functions in DMD patients.
For the long expression of target gene product, we tried to integrate the dystrophin gene into chromosome by using the sleeping beauty-transposone system and HDAdv. However, it revealed that the DNA construct of HDAdv is linear and it does not fit the circular DNA of the sleeping beauty-transposone system. So, further study is necessary to integrate the dystrophin gene into chromosome.

Research Products

• [Journal Article] Transduction of full-length dystrophin to multiple skeletal muscles improves motor performance and lifespan in utrophin/dystrophin double knockout mice (2008)
  • Author(s): Kawano R., Ishizaki M, Maeda Y., Uchida Y., Kimura E., Uchino M.
  • Journal Title: Mol Ther (in press)
  • NAID: 120002468583
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Mdx respiratory impairment following fibrosis of the diaphragm (2008)
  • Author(s): Ishizaki M, Suga T, Kimura E, Shiota T, Kawano R, Uchida U, Uchino K, Yamashita S, Maeda Y, Uchino M.
  • Journal Title: Neuromuscular disorders (in press)
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Regions downstream from the WW domain of dystrophin are impotant for binding to postsynaptic densities in the brain (2008)
  • Author(s): Sakamoto T, Arima T, Ishizaki M, Kawano R, Koide T, Uchida Y, Ymashita S, Kimura E, Hirano T, Maeda Y, Uchino M.
  • Journal Title: Neuromuscular disorders (in press)
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Transduction of full-length dystrophin to multiple skeletal muscles improves motor performance and lifespan in utrophin/ dystrophin double knockout mice. Mol Ther (2008)
  • Author(s): Kawano, R., Ishizaki, M., Maeda, Y., Uchida, Y., Kimura, E., Uchino, M.
  • Journal Title: (in press)
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Regions Downstream from the WW domain of dystrophin are important for binding to postsynaptic densities in the brain. Neuromuscul Disord (2008)
  • Author(s): Sakamoto, T., Arima, T., Ishizaki, M., Kawano, R., Koide, T., Uchida, Y., Yamashita, S., Kimura, E., Hirano, T., Maeda, Y., Uchino, M.
  • Journal Title: (in press)
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Mdx respiratory impairment following fibrosis of the diaphragm. Neuromuscul disord (2008)
  • Author(s): Ishizaki, M., Suga, T., Kimura, E., Shiota, T., Kawano, R., Uchida, U., Uchino, K., Yamashita, S., Maeda, Y., Uchino, M.
  • Journal Title: (in press)
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Transduction of full-length dystrophin to multiple skeletal muscles improves motor performance and lifespan in utrophin/dystrophin double knockout mice. (2008)
  • Author(s): Kawano R., Ishizaki M, Maeda Y., Uchida Y., Kimura E., Uchino M.
  • Journal Title: Mol Ther (in press)
  • NAID: 120002468583
  • Peer Reviewed
• [Presentation] Transduction of full-length dystrophin to multiple skeletal muscles improves motor performance and lifespan in utrophin/dystrophin double knockout mice (2007)
  • Author(s): Ishizaki M, Kawano R., Maeda Y., Kimura E., Uchino M.
  • Organizer: American Sosiety of Gene Terapy
  • Place of Presentation: Seattle
  • Year and Date: 2007-05-31
• [Presentation] Transduction of full-length dystrophin to multiple skeletal muscles improves motor performance and lifespan in utrophin/ dystrophin double knockout mice. (2007)
  • Author(s): Ishizaki, M., Kawano, R., Maeda, Y., Uchida, Y., Kimura, E., Uchino, M.
  • Organizer: American Society of Gene Therapy 10th annual meeting
  • Place of Presentation: Seattle, USA
  • Description: 「研究成果報告書概要(欧文)」より
• [Presentation] Restoration of physical performance in symptomatic utrophin/ dystrophin double knockout(U/D dko) mice by the helper-dependent adenovirus vector(HDAdv)-mediated full-length dystrophin expression. (2006)
  • Author(s): Kawano, R., Ishizaki, M., Uchida, Y., Kimura, E., Maeda, Y., Uchino, M.
  • Organizer: 11th European Society of Gene Therapy Annual meeting
  • Place of Presentation: Athens, Greece
  • Description: 「研究成果報告書概要(欧文)」より
• [Presentation] Effective repetitive dystrophin gene transfer into skeletal muscle of adult mdx mice using a helper-dependent adenovirus vector expressing the coxsackievirus and adenovirus receptor(CAR) and dystrophin (2005)
  • Author(s): Uchida, Y., Maeda, Y., Kimura, E., Uchino, M.
  • Organizer: American Society of Gene Therapy 9th annual meeting
  • Place of Presentation: Baltimore, USA
  • Description: 「研究成果報告書概要(欧文)」より