Gene suppressing therapy for Pelizaeus-Merzbacher disease using artificial microRNA
Heng Li, … , Takashi Okada, Ken Inoue
Heng Li, … , Takashi Okada, Ken Inoue
Published May 16, 2019
Citation Information: JCI Insight. 2019;4(10):e125052. https://doi.org/10.1172/jci.insight.125052.
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Research Article Neuroscience Therapeutics

Gene suppressing therapy for Pelizaeus-Merzbacher disease using artificial microRNA

Abstract

Copy number increase or decrease of certain dosage-sensitive genes may cause genetic diseases with distinct phenotypes, conceptually termed genomic disorders. The most common cause of Pelizaeus-Merzbacher disease (PMD), an X-linked hypomyelinating leukodystrophy, is genomic duplication encompassing the entire proteolipid protein 1 (PLP1) gene. Although the exact molecular and cellular mechanisms underlying PLP1 duplication, which causes severe hypomyelination in the central nervous system, remain largely elusive, PLP1 overexpression is likely the fundamental cause of this devastating disease. Here, we investigated if adeno-associated virus–mediated (AAV-mediated) gene-specific suppression may serve as a potential cure for PMD by correcting quantitative aberrations in gene products. We developed an oligodendrocyte-specific Plp1 gene suppression therapy using artificial microRNA under the control of human CNP promoter in a self-complementary AAV (scAAV) platform. A single direct brain injection achieved widespread oligodendrocyte-specific Plp1 suppression in the white matter of WT mice. AAV treatment in Plp1-transgenic mice, a PLP1 duplication model, ameliorated cytoplasmic accumulation of Plp1, preserved mature oligodendrocytes from degradation, restored myelin structure and gene expression, and improved survival and neurological phenotypes. Together, our results provide evidence that AAV-mediated gene suppression therapy can serve as a potential cure for PMD resulting from PLP1 duplication and possibly for other genomic disorders.

Authors

Heng Li, Hironori Okada, Sadafumi Suzuki, Kazuhisa Sakai, Hitomi Izumi, Yukiko Matsushima, Noritaka Ichinohe, Yu-ichi Goto, Takashi Okada, Ken Inoue

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Figure 2

Widespread oligodendrocyte-specific transgene expression in cerebral white matter following scAAV.CNP.Venus.miRneg vector injection.

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Widespread oligodendrocyte-specific transgene expression in cerebral whi...
One week prior to the analysis, scAAV.CNP.Venus.miRneg vector was injected into the right corpus striatum (CS) and internal capsule (IC) of P10 WT mice (n = 5). Representative low-magnification images in the corpus callosum (CC) (A) and IC (B), as well as high-magnification images in the CC (C), CS (D), and IC (E) demonstrate the extent of scAAV-derived reporter protein Venus expression. Cell nuclei were stained with DAPI (blue). (A and B figure inset) The schematic diagrams show the injection points (indicated by arrows) of the scAAV vector and the areas observed using a fluorescence microscope (indicated by boxes) in the CC (A inset), CS (A inset), and the IC (B inset). (F) Quantification of Venus expression areas in coronal brain sections taken from mice (n = 5) administered the scAAV vector. The x axis shows the relative distance from bregma and the arrows represent the injection points. (G–L) Immunostaining with cell-type-specific markers indicated that AAV-derived Venus expression rarely overlapped with NeuN-positive neurons (G), GFAP-positive astrocytes (H), or Iba1-positive microglia (I), but was mainly present in Gst-π–positive oligodendrocytes (J–L). (M) Quantification of the percentages of each cell type in Venus-positive cells (n = 5 mice, 4 sections per mouse). (N) Quantification of the percentage of Venus-positive oligodendrocytes relative to Gst-π–positive total mature oligodendrocytes in the center of AAV infection areas (n = 5 mice, 4 sections per mouse). Scale bars: 500 μm (A and B) and 50 μm (C–L). Statistical significance was determined using 1-way ANOVA with Bonferroni’s post hoc test and 2-tailed Student’s t test. ***P < 0.001.