記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：CELL PRESS  

The retina is an ideal target for gene therapy because of its easy accessibility and limited immunological response. We previously reported that intravitreally injected adeno-associated virus (AAV) vector transduced the inner retina with high efficiency in a rodent model. In large animals, however, the efficiency of retinal transduction was low, because the vitreous and internal limiting membrane (ILM) acted as barriers to transduction. To overcome these barriers in cynomolgus monkeys, we performed vitrectomy (VIT) and ILM peeling before AAV vector injection. Following intravitreal injection of 50 pi triple-mutated self-complementary AAV serotype 2 vector encoding EGFP, transduction efficiency was analyzed. Little expression of GFP was detected in the control and VIT groups, but in the VIT+ILM group, strong GFP expression was detected within the peeled ILM area. To detect potential adverse effects, we monitored the retinas using color fundus photography, optical coherence tomography, and electroretinography. No serious side effects associated with the pretreatment were observed. These results indicate that surgical ILM peeling before AAV vector administration would be safe and useful for efficient transduction of the nonhuman primate retina and provide therapeutic benefits for the treatment of retinal diseases.

DOI： 10.1016/j.ymthe.2016.10.008

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

Metachromatic leukodystrophy (MLD) is a lysosomal storage disease caused by a functional deficiency in human arylsulfatase A (hASA). We recently reported that ependymal cells and the choroid plexus are selectively transduced by intracerebroventricular (ICV) injection of adeno-associated virus serotype 1 (AAV1) vector and serve as a biological reservoir for the secretion of lysosomal enzymes into the cerebrospinal fluid (CSF). In the present study, we examined the feasibility of this AAV-mediated gene therapy to treat MLD model mice. Preliminary experiments showed that the hASA level in the CSF after ICV injection of self-complementary (sc) AAV1 was much higher than in mice injected with single-stranded AAV1 or scAAV9. However, when 18-week-old MLD mice were treated with ICV injection of scAAV1, the concentration of hASA in the CSF gradually decreased and was not detectable at 12 weeks after injection, probably due to the development of anti-hASA antibodies. As a result, the sulfatide levels in brain tissues of treated MLD mice were only slightly reduced compared with those of untreated MLD mice. These results suggest that this approach is potentially promising for treating MLD, but that controlling the immune response appears to be crucial for long-term expression of therapeutic proteins in the CSF.

DOI： 10.1038/srep13104

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

Enzyme replacement via the cerebrospinal fluid (CSF) has been shown to ameliorate neurological symptoms in model animals with neuropathic metabolic disorders. Gene therapy via the CSF offers a means to achieve a long-term sustainable supply of therapeutic proteins within the central nervous system (CNS) by setting up a continuous source of transgenic products. In the present study, a serotype 1 adeno-associated virus (AAV1) vector was injected into a lateral cerebral ventricle in adult mice to transduce the gene encoding human lysosomal enzyme arylsulfatase A (hASA) into the cells of the CNS. Widespread transduction and stable expression of hASA in the choroid plexus and ependymal cells was observed throughout the ventricles for more than 1 year after vector injection. Although humoral immunity to hASA developed after 6 weeks, which diminished the hASA levels detected in CSF from AAV1-injected mice, hASA levels in CSF were maintained for at least 12 weeks when the mice were tolerized to hASA prior of vector injection. Our results suggest that the cells lining the ventricles could potentially serve as a biological reservoir for long-term continuous secretion of lysosomal enzymes into the CSF following intracerebroventricular injection of an AAV1 vector.

DOI： 10.1038/srep05506

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記述言語：英語   出版者・発行元：MEDICAL ASSOC NIPPON MEDICAL SCH  

A variety of gene transfer strategies have been developed to treat inherited, degenerative, and acquired diseases. Among the different vector systems developed so far, recombinant adeno-associated viral (AAV) vectors have shown notable benefits, including prolonged gene expression, transduction of both dividing and nondividing cells, and a lack of pathogenicity caused by wild-type infections. Thanks to these features, the use of AAV vectors as a gene transfer tool has increased dramatically during the past several years, and several recent clinical trials have used AAV vectors. However, AAV vectors are more complicated to produce than are other viral vectors. With steady advances toward clinical application, much effort has been made to isolate novel AAV serotypes and to develop methods for their efficient, scalable, and versatile production and purification. Here we review state of the art methods for AAV vector production and purification, which we have refined in our laboratory. (J Nippon Med Sch 2012; 79: 394-402)

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記述言語：日本語   出版者・発行元：日本医科大学医学会  

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記述言語：日本語   出版者・発行元：眼科臨床紀要会  

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記述言語：日本語   出版者・発行元：(公財)日本眼科学会  

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記述言語：日本語   出版者・発行元：(公財)日本眼科学会  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：CELL PRESS  

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開催年月日： 2022年7月

記述言語：英語   会議種別：口頭発表（一般）  

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開催年月日： 2022年7月

記述言語：英語   会議種別：口頭発表（一般）  

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開催年月日： 2021年12月

会議種別：ポスター発表  

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開催年月日： 2021年9月

記述言語：英語   会議種別：口頭発表（一般）  

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開催年月日： 2020年12月

記述言語：英語   会議種別：ポスター発表  

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