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  • Artificial miRNAs Reduce Human Mutant Huntingtin Throughout the Striatum in a Transgenic Sheep Model of Huntington's Disease.

Artificial miRNAs Reduce Human Mutant Huntingtin Throughout the Striatum in a Transgenic Sheep Model of Huntington's Disease.

Human gene therapy (2017-12-07)
Edith L Pfister, Natalie DiNardo, Erica Mondo, Florie Borel, Faith Conroy, Cara Fraser, Gwladys Gernoux, Xin Han, Danjing Hu, Emily Johnson, Lori Kennington, PengPeng Liu, Suzanne J Reid, Ellen Sapp, Petr Vodicka, Tim Kuchel, A Jennifer Morton, David Howland, Richard Moser, Miguel Sena-Esteves, Guangping Gao, Christian Mueller, Marian DiFiglia, Neil Aronin
要旨

Huntington's disease (HD) is a fatal neurodegenerative disease caused by a genetic expansion of the CAG repeat region in the huntingtin (HTT) gene. Studies in HD mouse models have shown that artificial miRNAs can reduce mutant HTT, but evidence for their effectiveness and safety in larger animals is lacking. HD transgenic sheep express the full-length human HTT with 73 CAG repeats. AAV9 was used to deliver unilaterally to HD sheep striatum an artificial miRNA targeting exon 48 of the human HTT mRNA under control of two alternative promoters: U6 or CβA. The treatment reduced human mutant (m) HTT mRNA and protein 50-80% in the striatum at 1 and 6 months post injection. Silencing was detectable in both the caudate and putamen. Levels of endogenous sheep HTT protein were not affected. There was no significant loss of neurons labeled by DARPP32 or NeuN at 6 months after treatment, and Iba1-positive microglia were detected at control levels. It is concluded that safe and effective silencing of human mHTT protein can be achieved and sustained in a large-animal brain by direct delivery of an AAV carrying an artificial miRNA.

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Sigma-Aldrich
ウシ血清アルブミン ウシ血清由来, heat shock fraction, protease free, essentially globulin free, pH 7, ≥98%
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抗NeuN抗体、クローンA60, clone A60, Chemicon®, from mouse
Sigma-Aldrich
抗ハンチンチンタンパク質抗体、a.a. 181-810、クローン 1HU-4C8, ascites fluid, clone 1HU-4C8, Chemicon®
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モノクローナル抗アクチン マウス宿主抗体, clone AC-40, ascites fluid
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抗スペクトリンアルファ鎖(非赤血球)抗体、クローンAA6, clone AA6, Chemicon®, from mouse