title: Pmp22 mutant allele-specific siRNA alleviates demyelinating neuropathic phenotype in vivo.
abstract: Charcot-Marie-Tooth disease (CMT) is a genetic disorder that can be caused by aberrations in >80 genes. CMT has heterogeneous modes of inheritance, including autosomal dominant, autosomal recessive, X-linked dominant, and X-linked recessive. Over 95% of cases are dominantly inherited. In this study, we investigated whether regulation of a mutant allele by an allele-specific small interfering RNA (siRNA) can alleviate the demyelinating neuropathic phenotype of CMT. We designed 19 different allele-specific siRNAs for Trembler J (Tr-J) mice harboring a naturally occurring mutation (Leu16Pro) in Pmp22. Using a luciferase assay, we identified an siRNA that specifically and selectively reduced the expression level of the mutant allele and reversed the low viability of Schwann cells caused by mutant Pmp22 over-expression in vitro. The in vivo efficacy of the allele-specific siRNA was assessed by its intraperitoneal injection to postnatal day 6 of Tr-J mice. Administration of the allele-specific siRNA to Tr-J mice significantly enhanced motor function and muscle volume, as assessed by the rotarod test and magnetic resonance imaging analysis, respectively. Increases in motor nerve conduction velocity and compound muscle action potentials were also observed in the treated mice. In addition, myelination, as evidenced by toluidine blue staining and electron microscopy, was augmented in the sciatic nerves of the mice after allele-specific siRNA treatment. After validating suppression of the Pmp22 mutant allele at the mRNA level in the Schwann cells of Tr-J mice, we observed increased expression levels of myelinating proteins such as myelin basic protein and myelin protein zero. These data indicate that selective suppression of the Pmp22 mutant allele by non-viral delivery of siRNA alleviates the demyelinating neuropathic phenotypes of CMT in vivo, implicating allele-specific siRNA treatment as a potent therapeutic strategy for dominantly inherited peripheral neuropathies.
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea; Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, Republic of Korea.
authors.forename: Ji-Su
authors.initials: JS
authors.lastname: Lee
authors.affiliation: Stem Cell & Regenerative Medicine Institute, Samsung Medical Center, Seoul, Republic of Korea; Samsung Biomedical Research Institute, Samsung Advanced Institute of Technology, Samsung Electronics Co., Ltd., Seoul, Republic of Korea.
authors.forename: Eun Hyuk
authors.initials: EH
authors.lastname: Chang
authors.affiliation: Samsung Biomedical Research Institute, Samsung Advanced Institute of Technology, Samsung Electronics Co., Ltd., Seoul, Republic of Korea.
authors.forename: Ok Jae
authors.initials: OJ
authors.lastname: Koo
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea.
authors.forename: Dong Hwan
authors.initials: DH
authors.lastname: Jwa
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea.
authors.forename: Won Min
authors.initials: WM
authors.lastname: Mo
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea; Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, Republic of Korea.
authors.forename: Geon
authors.initials: G
authors.lastname: Kwak
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea.
authors.forename: Hyo Won
authors.initials: HW
authors.lastname: Moon
authors.affiliation: Department of Physiology, College of Medicine, Dong-A University, Busan, Republic of Korea.
authors.forename: Hwan Tae
authors.initials: HT
authors.lastname: Park
authors.affiliation: Stem Cell & Regenerative Medicine Institute, Samsung Medical Center, Seoul, Republic of Korea; Department of Neurology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea. Electronic address: youngbinhong@gmail.com.
authors.forename: Young Bin
authors.initials: YB
authors.lastname: Hong
authors.affiliation: Neuroscience Center, Samsung Medical Center, Seoul, Republic of Korea; Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, Republic of Korea; Department of Neurology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea. Electronic address: bochoi77@hanmail.net.
authors.forename: Byung-Ok
authors.initials: BO
authors.lastname: Choi
mesh_terms: D000483:Alleles; D000818:Animals; D002607:Charcot-Marie-Tooth Disease / Q000235:genetics* / Q000473:pathology; D003711:Demyelinating Diseases / Q000235:genetics* / Q000473:pathology; D008822:Mice, Transgenic; D009154:Mutation / Q000235:genetics*; D009185:Myelin Proteins / Q000235:genetics*; D010641:Phenotype; D034741:RNA, Small Interfering / Q000235:genetics*; D012583:Schwann Cells / Q000378:metabolism; D012584:Sciatic Nerve / Q000378:metabolism
chemical_list: D009185:Myelin Proteins; C489545:Pmp22 protein, mouse; D034741:RNA, Small Interfering
keywords: Charcot-Marie-Tooth disease; Demyelination; Gene therapy; Non-viral delivery; PMP22