Adeno-associated virus vector-mediated gene transfer into dystrophin-deficient skeletal muscles evokes enhanced immune response against the transgene product

Yuasa, Katsutoshi; Sakamoto, Miki; Miyagoe-Suzuki, Yuko; Tanouchi, Aki; Yamamoto, Hiroshi; Li, J; Chamberlain, Jeffrey S.; Xiao, Xianmin; Takeda, Shin’ichi

doi:10.1038/sj.gt.3301829

Cited by 124 publications

(96 citation statements)

References 38 publications

(59 reference statements)

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“…The endogenous expression of the LacZ gene in these transgenic mice avoided interference factors, such as an immune response to the AAV viral vector in C57/ BL10 28 and mdx mice. 7 The 10-day-old dMCK-LacZ transgenic mice were strongly positive for b-galactosidase expression in all the posterior skeletal muscles, while the negative control littermates were clearly negative for X-gal staining (Figure 4a). High levels of b-galactosidase were also observed in the skeletal muscles of the forelimbs and the face of dMCK-LacZ transgenic mice (Figure 4b), but not in the negative control littermates (Figure 4c).…”

Section: Examination Of the Dmck Promoter In Transgenic Mice With Lacmentioning

confidence: 98%

“…A main concern in the development of DMD gene therapy vectors is that widespread targeted gene expression in tissues such as the liver and lung will result in overall toxicity and/or the initiation of a host immune response against tissues expressing the transgene or gene vector. [6][7][8] The viral promoters, such as human cytomegalovirus (CMV) immediate-early gene promoter, achieve high levels of nonspecific gene expression; in contrast, the use of the muscle creatine kinase (MCK) promoter is specific to muscle tissue and can avoid the potentially harmful effects of ectopic transgene expression. [9][10][11][12] However, the MCK promoter is less active than viral promoters, and its large size (6.5 kb) makes it incompatible with adeno-associated viral (AAV) vectors, a noteworthy gene delivery vehicle with a 4.5-kb capacity for the treatment of muscular dystrophies and other degenerative disorders.…”

Section: Instructionmentioning

confidence: 99%

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Construction and analysis of compact muscle-specific promoters for AAV vectors

et al. 2008

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Adeno-associated viral (AAV) vectors have been broadly used for gene transfer in vivo for various applications. However, AAV precludes the use of most of the original large-sized tissue-specific promoters for expression of transgenes. Efforts are made to develop highly compact, active and yet tissue-specific promoters for use in AAV vectors. In this study, we further abbreviated the muscle creatine kinase (MCK) promoter by ligating a double or triple tandem of MCK enhancer (206-bp) to its 87-bp basal promoter, generating the dMCK (509-bp) and tMCK (720-bp) promoters. The dMCK promoter is shorter but stronger than some previously developed MCK-based promoters such as the enh358MCK (584-bp) and CK6 (589-bp) in vitro in C2C12 myotubes and in vivo in skeletal muscles. The tMCK promoter is the strongest that we tested here, more active than the promiscuous cytomegalovirus (CMV) promoter. Furthermore, both the dMCK and tMCK promoters are essentially inactive in nonmuscle cell lines as well as in the mouse liver (4200-fold weaker than the CMV promoter).The dMCK promoter was further tested in a few lines of transgenic mice. Expression of LacZ or minidystrophin gene was detected in skeletal muscles throughout the body, but was weak in the diaphragm, and undetectable in the heart and other tissues. Similar to other miniature MCK promoters, the dMCK promoter also shows preference for fast-twitch myofibers. As a result, we further examined a short, synthetic muscle promoter C5-12 (312-bp). It is active in both skeletal and cardiac muscles but lacks apparent preference on myofiber types. Combination of a MCK enhancer to promoter C5-12 has increased its strength in muscle by two-to threefold. The above-mentioned compact muscle-specific promoters are well suited for AAV vectors in muscle-directed gene therapy studies.

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Section: Examination Of the Dmck Promoter In Transgenic Mice With Lacmentioning

confidence: 98%

Section: Instructionmentioning

confidence: 99%

Construction and analysis of compact muscle-specific promoters for AAV vectors

et al. 2008

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“…12,37 The proviral vector plasmid pAAV hDCS2, an AAV9 helper plasmid pAAV2/9 (a gift from Dr James M Wilson), 38 and an adenoviral helper plasmid pAdeno 39 were transfected into HEK293 cells at 60% confluence in a 10-tray Cell Factory (Nunc, Thermo Fisher, Roskilde Site, Denmark), and incubated for 48 h at 37 1C and 5% CO 2 with use of the active gassing technique. 40 The rAAV particles were purified by the dual ion-exchange procedures with the highperformance membrane adsorbers.…”

Section: Construction Of Proviral Plasmid and Recombinant Aav Vector mentioning

confidence: 99%

Improvement of cardiac fibrosis in dystrophic mice by rAAV9-mediated microdystrophin transduction

et al. 2011

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Duchenne muscular dystrophy (DMD) is the most common form of the progressive muscular dystrophies characterized by defects of the dystrophin gene. Although primarily characterized by degeneration of the limb muscles, cardiomyopathy is a major cause of death. Therefore, the development of curative modalities such as gene therapy is imperative. We evaluated the cardiomyopathic features of mdx mice to observe improvements in response to intravenous administration of recombinant adeno-associated virus (AAV) type 9 encoding microdystrophin. The myocardium was extensively transduced with microdystrophin to significantly prevent the development of fibrosis, and expression persisted for the duration of the study. Intraventricular conduction patterns, such as the QRS complex duration and S/R ratio in electrocardiography, were also corrected, indicating that the transduced microdystrophin has a protective effect on the dystrophin-deficient myocardium. Furthermore, BNP and ANP levels were reduced to normal, suggesting the absence of cardiac dysfunction. In aged mice, prevention of ectopic beats as well as echocardiographic amelioration was also demonstrated with improved exercise performance. These findings indicate that AAV-mediated cardiac transduction with microdystrophin might be a promising therapeutic strategy for the treatment of dystrophin-deficient cardiomyopathy.

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“…As administration of an equivalent dose of rAAV6 empty capsids with VEGF did not induce mononuclear cell infiltrates (Fig. 4d, e), we hypothesized that the inflammatory reaction observed with CMV-lacZ in wild-type mice results from the constitutive widespread expression of a bacterial protein (β-gal) 18 . Indeed, administration of 1 × 10 12 vector genomes of rAAV6-CMV-lacZ and VEGF to transgenic mice that express bacterial β-gal within the intestinal villi 19 , did not cause mononuclear cell accumulation (Fig.…”

Section: Raav6 Vectors Potently Transduce the Myocardium In Vivomentioning

confidence: 99%

Systemic delivery of genes to striated muscles using adeno-associated viral vectors

Gregorevic

Blankinship

Allen

et al. 2004

Nat Med

573

503

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A major obstacle limiting gene therapy for diseases of the heart and skeletal muscles is an inability to deliver genes systemically to muscles of an adult organism. Systemic gene transfer to striated muscles is hampered by the vascular endothelium, which represents a barrier to distribution of vectors via the circulation. Here we show the first evidence of widespread transduction of both cardiac and skeletal muscles in an adult mammal, after a single intravenous administration of recombinant adenoassociated virus pseudotype 6 vectors. The inclusion of vascular endothelium growth factor/vascular permeability factor, to achieve acute permeabilization of the peripheral microvasculature, enhanced tissue transduction at lower vector doses. This technique enabled widespread muscle-specific expression of a functional micro-dystrophin in the skeletal muscles of dystrophin-deficient mdx mice, which model Duchenne muscular dystrophy. We propose that these methods may be applicable for systemic delivery of a wide variety of genes to the striated muscles of adult mammals.Human mortality and quality of life are significantly affected by diseases of the striated musculature. Genetic treatments that are being developed for conditions such as heart disease, aging-associated muscle wasting and the muscular dystrophies have been limited by an inability to achieve widespread and efficient gene transfer to the heart and dispersed skeletal muscles of an adult organism 1-4 . For example, anesthesia, invasive surgery and hazardous cofactors are required to transduce varying fractions of the cardiomyocyte population efficiently 1,5 . Similarly, the transfer of genes to the muscles of individual limbs using various vectors requires either direct injection of individual muscles, or complex surgical procedures performed under anesthesia to distribute vectors via the circulation 2-4,6-10 . Here we describe a simple and highly efficient method to transfer genes systemically to the cardiac and skeletal muscles of adult mammals. This approach uses intravenous administration of recombinant adeno-associated virus pseudotype 6 (rAAV6-pseudotyped vectors) 11 , which are extremely effective at transducing skeletal muscles after intramuscular injection 12 .Correspondence should be addressed to J.S.C. (jsc5@u.washington.edu).. 4 These authors contributed equally to this work.Note: Supplementary information is available on the Nature Medicine website. COMPETING INTERESTS STATEMENTThe authors declare competing financial interests (see the Nature Medicine website for details). NIH Public Access RESULTS Systemic transduction of skeletal muscles by rAAV6First, we examined the potential for systemic gene transfer after intravenous administration of rAAV6 vectors at the whole-body level in young adult (6-8 wk) C57Bl/10J mice (Fig. 1). The muscles of mice examined 11 days after administration through the tail vein of ~2 × 10 11 vector genomes of rAAV6 vector containing a CMV-lacZ expression cassette, did not show obvious exogenous β-galactosidase (...

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Adeno-associated virus vector-mediated gene transfer into dystrophin-deficient skeletal muscles evokes enhanced immune response against the transgene product

Cited by 124 publications

References 38 publications

Construction and analysis of compact muscle-specific promoters for AAV vectors

Construction and analysis of compact muscle-specific promoters for AAV vectors

Improvement of cardiac fibrosis in dystrophic mice by rAAV9-mediated microdystrophin transduction

Systemic delivery of genes to striated muscles using adeno-associated viral vectors

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