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Myth Unmethylated: Novel Therapies for Methylation-Related Neurodevelopmental Disorders

Shivani Mishra
Department of Medical Genetics, Sanjay Gandhi Post graduate Institute of Medical Sciences, Lucknow, India
Email: drshivani2005@gmail.com

1 Reactivation of FMR1 by CRISPR/Cas9 (Xie et al., 2016)

Fragile X syndrome (FXS) is a CGG-repeat disorder of the FMR1 gene caused by epigenetic gene silencing. In the presence of the elongated CGG repeat, epigenetic modifying drugs result in only transient FMR1 reactivation. CRISPR/Cas9 genome editing was used to excise the expanded CGG-repeat in both somatic cell hybrids containing the human fragile X chromosome and human FXS iPS cells. Transcriptional reactivation was observed in approximately 67% of the CRISPR cut hybrid colonies and in 20% of isolated human FXS iPSC colonies. The reactivated cells produced fragile X mental retardation protein (FMRP) and exhibited a decrease in DNA methylation at the FMR1 locus.

2 Transcriptional reactivation of the FMR1 gene (Tabolacci et al., 2016)

In Fragile X syndrome (FXS), CGG expansion and subsequent DNA methylation of the promoter region, is accompanied by additional epigenetic histone modifications that result in a block of transcription and absence of the fragile X mental retardation protein (FMRP). In vitro treatment of FXS lymphoblastoid cell lines with the demethylating agent 5-azadeoxycytidine for 7 days resulted in transcriptional reactivation of the FMR1 gene and FMRP production, demonstrating that DNA methylation is key to FMR1 inactivation. These observations demonstrate that a therapeutic approach to FXS based on the pharmacological reactivation of the FMR1 gene is conceptually worthy of being pursued further.

3 Systemic delivery of MECP2 rescues behavioral and cellular deficits in female mouse models of Rett syndrome (Garg et al., 2013)

Rett syndrome is a severe X-linked neurodevelopmental disorder that is primarily caused by mutations in the methyl CpG binding protein 2 (MECP2) gene. Systemic administration of self-complementary Adeno-associated virus-9 (AAV9), bearing MeCP2 cDNA under control of a fragment of its own promoter (scAAV9/MeCP2), was demonstrated to be capable of significantly stabilizing or reversing symptoms in female mice with Rett syndrome. This increased MeCP2 level to 65% from 50% and resulted in improvement in motor function, tremors, seizures and hind limb clasping. Smaller body size of neurons was restored to normal. However, it could not rectify breathing deficits in the Rett-affected mice. This study has shown the first potential gene therapy for females afflicted with Rett syndrome.

4 Reduction of a long non-coding RNA in Angelman syndrome (Meng et al., 2015)

Angelman syndrome is caused by maternal deficiency of the imprinted gene UBE3A, encoding an E3 ubiquitin ligase. All patients carry at least one copy of paternal UBE3A, which is intact but silenced by a nuclear-localized long non-coding RNA, UBE3A antisense transcript (UBE3A-ATS). Murine Ube3a-ATS reduction by either transcription termination or topoisomerase I inhibition has been shown to increase paternal Ube3a expression. Antisense oligonucleotides (ASOs) treatment achieved specific reduction of Ube3a-ATS and sustained unsilencing of paternal Ube3a in neurons in vitro and in vivo. Partial restoration of UBE3A protein in an Angelman syndrome mouse model ameliorated some cognitive deficits associated with the disease, although additional studies of phenotypic correction are needed.


1.    Garg SK, et al. Systemic delivery of MeCP2 rescues behavioral and cellular deficits in female mouse models of Rett syndrome. J Neurosci 2013; 33: 13612-13620.

2.    Meng L, et al. Towards a therapy for Angelman syndrome by targeting a long non-coding RNA. Nature 2015; 518: 409-412.

3.    Tabolacci E, et al. Transcriptional Reactivation of the FMR1 Gene. A Possible Approach to the Treatment of the Fragile X Syndrome. Genes (Basel) 2016; 7: pii E49.

4.    Xie N, et al. Reactivation of FMR1 by CRISPR/Cas9-mediated deletion of the expanded CGG-repeat of the fragile X chromosome. PLoS ONE 2016; 11: e0165499.

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