Targeted treatments for fragile X syndrome

Fragile X syndrome (FXS) is the most common identifiable genetic cause of intellectual disability and autistic spectrum disorders (ASD), with up to 50% of males and some females with FXS meeting criteria for ASD. Autistic features are present in a very high percent of individuals with FXS, even those who do not meet full criteria for ASD. Recent major advances have been made in the understanding of the neurobiology and functions of FMRP, the FMR1 (fragile X mental retardation 1) gene product, which is absent or reduced in FXS, largely based on work in the fmr1 knockout mouse model. FXS has emerged as a disorder of synaptic plasticity associated with abnormalities of long-term depression and long-term potentiation and immature dendritic spine architecture, related to the dysregulation of dendritic translation typically activated by group I mGluR and other receptors. This work has led to efforts to develop treatments for FXS with neuroactive molecules targeted to the dysregulated translational pathway. These agents have been shown to rescue molecular, spine, and behavioral phenotypes in the FXS mouse model at multiple stages of development. Clinical trials are underway to translate findings in animal models of FXS to humans, raising complex issues about trial design and outcome measures to assess cognitive change that might be associated with treatment. Genes known to be causes of ASD interact with the translational pathway defective in FXS, and it has been hypothesized that there will be substantial overlap in molecular pathways and mechanisms of synaptic dysfunction between FXS and ASD. Therefore, targeted treatments developed for FXS may also target subgroups of ASD, and clinical trials in FXS may serve as a model for the development of clinical trial strategies for ASD and other cognitive disorders

FMR1 premutation and full mutation molecular mechanisms related to autism

Fragile X syndrome (FXS) is caused by an expanded CGG repeat (>200 repeats) in the 5′ un-translated portion of the fragile X mental retardation 1 gene (FMR1) leading to a deficiency or absence of the FMR1 protein (FMRP). FMRP is an RNA-binding protein that regulates the translation of a number of other genes that are important for synaptic development and plasticity. Furthermore, many of these genes, when mutated, have been linked to autism in the general population, which may explain the high comorbidity that exists between FXS and autism spectrum disorders (ASD). Additionally, premutation repeat expansions (55 to 200 CGG repeats) may also give rise to ASD through a different molecular mechanism that involves a direct toxic effect of FMR1 mRNA. It is believed that RNA toxicity underlies much of the premutation-related involvement, including developmental concerns like autism, as well as neurodegenerative issues with aging such as the fragile X-associated tremor ataxia syndrome (FXTAS). RNA toxicity can also lead to mitochondrial dysfunction, which is common in older premutation carriers both with and without FXTAS. Many of the problems with cellular dysregulation in both premutation and full mutation neurons also parallel the cellular abnormalities that have been documented in idiopathic autism. Research regarding dysregulation of neurotransmitter systems caused by the lack of FMRP in FXS, including metabotropic glutamate receptor 1/5 (mGluR1/5) pathway and GABA pathways, has led to new targeted treatments for FXS. Preliminary evidence suggests that these new targeted treatments will also be beneficial in non-fragile X forms of autism

Fragile x syndrome and autism: from disease model to therapeutic targets

Autism is an umbrella diagnosis with several different etiologies. Fragile X syndrome (FXS), one of the first identified and leading causes of autism, has been modeled in mice using molecular genetic manipulation. These Fmr1 knockout mice have recently been used to identify a new putative therapeutic target, the metabotropic glutamate receptor 5 (mGluR5), for the treatment of FXS. Moreover, mGluR5 signaling cascades interact with a number of synaptic proteins, many of which have been implicated in autism, raising the possibility that therapeutic targets identified for FXS may have efficacy in treating multiple other causes of autism

Transcriptome Analysis of Synaptoneurosomes Identifies Neuroplasticity Genes Overexpressed in Incipient Alzheimer's Disease

In Alzheimer's disease (AD), early deficits in learning and memory are a consequence of synaptic modification induced by toxic beta-amyloid oligomers (oAβ). To identify immediate molecular targets downstream of oAβ binding, we prepared synaptoneurosomes from prefrontal cortex of control and incipient AD (IAD) patients, and isolated mRNAs for comparison of gene expression. This novel approach concentrates synaptic mRNA, thereby increasing the ratio of synaptic to somal mRNA and allowing discrimination of expression changes in synaptically localized genes. In IAD patients, global measures of cognition declined with increasing levels of dimeric Aβ (dAβ). These patients also showed increased expression of neuroplasticity related genes, many encoding 3′UTR consensus sequences that regulate translation in the synapse. An increase in mRNA encoding the GluR2 subunit of the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) was paralleled by elevated expression of the corresponding protein in IAD. These results imply a functional impact on synaptic transmission as GluR2, if inserted, maintains the receptors in a low conductance state. Some overexpressed genes may induce early deficits in cognition and others compensatory mechanisms, providing targets for intervention to moderate the response to dAβ

Dysregulated metabotropic glutamate receptor-dependent translation of AMPA receptor and postsynaptic density-95 mRNAs at synapses in a mouse model of fragile X syndrome

Fragile X syndrome, a common form of inherited mental retardation, is caused by the loss of fragile X mental retardation protein (FMRP), an mRNA binding protein that is hypothesized to regulate local mRNA translation in dendrites downstream of gp1 metabotropic glutamate receptors (mGluRs). However, specific FMRP-associated mRNAs that localize to dendrites in vivo and show altered mGluR-dependent translation at synapses of Fmr1 knock-out mice are unknown so far. Using fluorescence in situ hybridization, we discovered that GluR1/2 and postsynaptic density-95 (PSD-95) mRNAs are localized to dendrites of cortical and hippocampal neurons in vivo. Quantitative analyses of their dendritic mRNA levels in cultured neurons and synaptoneurosomes did not detect differences between wild-type and Fmr1 knock-out (KO) mice. In contrast, PSD-95, GluR1/2, and calcium/calmodulin-dependent kinase IIα (CaMKIIα) mRNA levels in actively translating polyribosomes were dysregulated in synaptoneurosomes from Fmr1 knock-out mice in response to mGluR activation. [35S]methionine incorporation into newly synthesized proteins similarly revealed impaired stimulus-induced protein synthesis of CaMKIIα and PSD-95 in synaptoneurosomes from Fmr1 KO mice. Quantitative analysis of mRNA levels in FMRP-specific immunoprecipitations from synaptoneurosomes demonstrated the association of FMRP with CaMKIIα, PSD-95, and GluR1/2 mRNAs. These findings suggest a novel mechanism whereby FMRP regulates the local synthesis AMPA receptor (AMPAR) subunits, PSD-95, and CaMKIIα downstream of mGluR-activation. Dysregulation of local translation of AMPAR and associated factors at synapses may impair control of the molecular composition of the postsynaptic density and consequently alter synaptic transmission, causing impairments of neuronal plasticity observed in Fmr1 knock-out mice and fragile X syndrome

Generation of two iPSC lines with either a heterozygous V717I or a heterozygous KM670/671NL mutation in the APP gene

Alzheimer's disease (AD) is the most common form of dementia, affecting millions of people worldwide. Mutations in the genes PSEN1, PSEN2 or APP are known to cause familial forms of AD with an early age of onset. In this study, specific pathogenic mutations in the APP gene were introduced into an iPSC line from a healthy individual by the use of CRISPR-Cas9. The study resulted in the generation of two new cell lines, one carrying the V717I APP mutation and one with the KM670/671NL APP mutation

Generation of a set of isogenic, gene-edited iPSC lines homozygous for all main APOE variants and an APOE knock-out line

Alzheimer's disease (AD) is the most frequent neurodegenerative disease amongst the elderly. The SNPs rs429358 and rs7412 in the APOE gene are the most common risk factor for sporadic AD, and there are three different alleles commonly referred to as APOE-ε2, APOE-ε3 and APOE-ε4. Induced pluripotent stem cells (iPSCs) hold great promise to model AD as such cells can be differentiated in vitro to the required cell type. Here we report the use of CRISPR/Cas9 technology employed on iPSCs from a healthy individual with an APOE-ε3/ε4 genotype to obtain isogenic APOE-ε2/ε2, APOE-ε3/ε3, APOE-ε4/ε4 lines as well as an APOE-knock-out line