Targeted treatments for fragile X syndrome
© Springer Science+Business Media, LLC 2011
Received: 26 October 2010
Accepted: 24 January 2011
Published: 19 February 2011
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.
KeywordsFragile X syndrome Autism FMRP Metobotropic glutamate receptors GABA agonists Signal transduction Dendritic translation Synaptic plasticity
In the past decade, the study of the neurobiology and synaptic mechanisms in fragile X syndrome has emerged as a molecular doorway to future targeted treatments for autism and related developmental disorders. Fragile X syndrome (FXS) provides an excellent model for the translation of basic molecular neuroscience findings to clinical treatment because FXS is a genetically defined condition in which all affected individuals have a uniform single gene defect as the etiology for their condition, an increasing body of information is available regarding the mechanisms through which the genetic defect in FXS impacts molecular events in neurons and resultant synaptic plasticity, a sufficient number of individuals with FXS can be identified to carry out trials, and aspects of FXS model other more common neurodevelopmental conditions such as autistic spectrum disorders (ASDs), learning disability, and attention deficit/hyperactivity disorder (ADHD). Despite the positive aspects of FXS as a translational model, no real template exists for how to approach the translational process, and there are questions about the length of treatment required to impact the underlying disorder, the possibility of developmental windows beyond which treatment cannot be successful, dosing and trial design issues, and the problem of how to measure both short- and long-term outcomes, which are ongoing hurdles that must be addressed. Nonetheless, these questions will have to be addressed for targeted treatment of FXS or any neurodevelopmental disorder to be successful, and this paper will review the current state of the translational process for targeted treatment in FXS, including known targets, preclinical work with small molecule therapeutics aimed at those targets, active or completed clinical trials, and issues associated with trial design and outcome measures.
Genetics of fragile X syndrome
FXS is the most common known inherited cause of intellectual disability (ID), learning disability and ASD, with an estimated frequency of about 1/2,500–1/4,000 (Hagerman 2008; Turner et al. 1996). FXS results from a trinucleotide repeat (CGG) expansion mutation of >200 repeats (full mutation) in the promoter of the FMR1 (fragile X mental retardation 1) gene (Verkerk et al. 1991), which leads to methylation and transcriptional silencing of the FMR1 promoter with consequent loss or significant reduction of expression of the gene product, FMRP (fragile X mental retardation protein; Devys et al. 1993). FMRP is a multifunctional mRNA binding protein involved in the dendritic transport, localization, and translational regulation of several hundreds of mRNA ligands. Therefore, FMRP is thought to regulate translation at the dendrite in response to neural activation, thereby modulating synaptic plasticity and dendritic morphology (for reviews, see Bagni and Greenough 2005; Grossman et al. 2006; Bassell and Warren 2008).
Smaller FMR1 expansions with 55–200 repeats (normal is <45), termed the premutation, are not associated with FMR1 methylation or loss of FMRP expression, but do result in fragile X-associated tremor/ataxia syndrome (Berry-Kravis et al. 2007) or fragile X-associated primary ovarian insufficiency (Sullivan et al. 2005). These conditions occur through a presumed RNA toxicity mechanism due to elevated levels of CGG repeat-containing mRNA which accompany the mild reduction in translation of FMRP in the presence of the repeat expansion (Berry-Kravis et al. 2007). Some premutation carriers have been found to have subtle evidence of features that overlap those seen in FXS, including emotional problems such as anxiety, social deficits, obsessive thinking, and/or depression (Hessl et al. 2005; Cornish et al. 2005). A small subgroup of carriers with a larger premutation have mild cognitive disorders and features of FXS, presumed due to uncompensated reduction in translation, with a resultant deficit in FMRP (Tassone et al. 2000).
Because FMR1 is located on the X chromosome, females with a full mutation are more variably affected and, on average, more mildly affected than males due to the production of FMRP from the normal FMR1 allele on the non-mutated X chromosome. Severity of the cognitive impairment and behavioral phenotypes in females with FXS and the full mutation is inversely related to the activation ratio for the normal FMR1 allele and the level of FMRP (Loesch et al. 2002, 2004). Likewise, in males with a full mutation and mosaicism for an unmethylated allele, the severity of the cognitive disorder is related to the amount of unmethylated DNA and FMRP level (Loesch et al. 2002, 2004; Tassone et al. 1999).
Fragile X phenotype
Males with a completely methylated full mutation commonly display mild to moderate ID (Hagerman et al. 2009). Females with the full mutation typically present with learning disabilities, although approximately 25% have ID (de Vries et al. 1996). Physical features include macroorchidism (present in most adult males), and more variably present are prominent ears, macrocephaly, long face, high arched palate, and loose connective tissue leading to hyperextensible joints, flat feet, and soft skin (Hagerman et al. 2009). Medical problems commonly include frequent ear infections, mitral valve prolapse, and seizures (Hagerman et al. 2009). Males with FXS have characteristic behavioral features including hyperactivity, impulsivity, attention problems, anxiety, mood lability, and autistic features such as poor eye contact, shyness, self-talk, hand flapping, hand biting, hyperarousal to sensory stimuli, and substantial perseverative language and behavior (Hagerman et al. 2009; Berry-Kravis and Potanos 2004; Wang et al. 2010a). They demonstrate an enhanced sympathetic response to sensory stimuli, as measured by electrodermal responses (Miller et al. 1999), heart rate variability (Boccia and Roberts 2000), and pupillary responses (Farzin et al. 2009); abnormal sensory gating can be demonstrated in prepulse inhibition studies (Hessl et al. 2008a). Anxiety disorders are common in both males and females with FXS, including selective mutism, separation anxiety, social phobia, and specific phobias (Hagerman et al. 2009; de Vries et al. 1996; Sullivan et al. 2007), and there is often generalized anxiety with multiple specific areas of difficulty. Aggression occurs in at least 30–50% of males and is most commonly seen in adolescence (Hessl et al. 2008b).
Females with FXS exhibit more variable and less frequent involvement with respect to physical features and medical problems, but often have attention problems, impulsivity, and executive function deficits even when their IQ is in the normal range (Hagerman et al. 2009; de Vries et al. 1996; Berry-Kravis and Potanos 2004; Wang et al. 2010a). Shyness, selective mutism, specific phobias, social anxiety, and social deficits are common in females (Berry-Kravis and Potanos 2004; Wang et al. 2010a).
Molecular neurobiology and synaptic pathology in FXS
After transport to dendrites and spines, FMRP binds to mRNAs (including its own) and regulates their translation in response to neural activation (Weiler et al. 1997, 2004). Specifically, FMRP appears to regulate translation pathways activated by group 1 metabotropic glutamate receptors (mGluR1 and mGluR5; Huber et al. 2002; Antar et al. 2004; Aschrafi et al. 2005) and muscarinic (M1) acetylcholine receptors (Volk et al. 2007) and possibly has a more general function in the regulation of translational activation by multiple synaptic Gq-linked receptors, including dopamine D1 receptors (Wang et al. 2010b). Activation of these receptors transiently induces FMRP dephosphorylation (Narayanan et al. 2008), which results in the reduction of FMRP interaction with Argonaut (AGO2) and microRNAs in the RISC complex and increased interaction with Dicer (Cheever and Ceman 2009) and, hence, loss of interaction with bound mRNAs, thereby relieving the repressive effect of FMRP on the translation of these mRNA targets, leading to the production of a pulse of new protein (Qin et al. 2005). Some of the FMRP-regulated proteins, in particular STEP (Zhang et al. 2008) and Arc (Park et al. 2008), are likely responsible for AMPA receptor endocytosis and resultant maintenance of translation-dependent group 1 mGluR-induced long-term depression (LTD) in hippocampus, as well as other receptor-activated translation-dependent forms of LTD and long-term potentiation (LTP) throughout the brain.
In the fmr1 knockout mouse, FMRP is absent, and there is both constitutively elevated translation of FMRP target mRNAs (∼2-fold normal) and a loss of the translation “pulse” after mGluR stimulation (both due to loss of baseline translational repression when FMRP is not present). Thus, levels of synaptic proteins corresponding to a number of FMRP target mRNAs have been shown to be constitutively elevated in the fmr1 knockout mouse, including MAP1B, PSD95, CaMKII, STEP, PIKE, amyloid precursor protein (APP), Arc, PP2A, potassium channel Kv3.1b, and others (Darnell et al. 2001; Brown et al. 2001; Chen et al. 2003; Miyashiro et al. 2003; Park et al. 2008; Zhang et al. 2001; Muddashetty et al. 2007; Goebel-Goody et al. 2010; Westmark and Malter 2007; Shumbos et al. 2010). The levels of many of these proteins have also been shown to be unresponsive to group I mGluR activation. This dysregulated dendritic expression (either temporally, spatially, or quantitatively) of multiple FMRP mRNA ligands results in abnormal synaptic plasticity, including enhanced mGluR-activated hippocampal (Huber et al. 2002) and cerebellar (Koekkoek et al. 2005) LTD and impaired LTP in the hippocampus (Lauterborn et al. 2007), cortex (Li et al. 2002; Larson et al. 2005), and amygdala (Subrathan et al. 2010). Other expected consequences of excessive constitutive activation of mGluR-mediated dendritic protein synthesis are also found in the fmr1 knockout (KO) mouse, including reduction of synaptic AMPA receptors (Koekkoek et al. 2005; Lauterborn et al. 2007), abnormal epileptiform discharges (Chuang et al. 2005), and abnormal dendritic spine morphology (Grossman et al. 2006; Comery et al. 1997; Irwin et al. 2002; Nimchinsky et al. 2001).
Indeed, the cerebral cortex of adult fmr1 KO mice (Grossman et al. 2006; Chuang et al. 2005; Comery et al. 1997; Irwin et al. 2002; Nimchinsky et al. 2001) and autopsy specimens from individuals with FXS (Irwin et al. 2001) both show increased density of long, thin, tortuous postsynaptic dendritic spines which are considered “immature” and are normally seen in early neocortical development as well as reduction of mature, short mushroom-shaped spines. This pattern appears to represent a deficit in the “pruning” of unnecessary synaptic contacts, suggesting that FMRP is required for the important processes of synapse stabilization and pruning during synapse maturation. The morphological abnormalities and synaptic plasticity deficits found in the fmr1 knockout mouse are associated with numerous cognitive, behavioral, and electrophysiological phenotypes, including abnormal ocular dominance plasticity (Dolen et al. 2007), olfactory learning deficits (Larson et al. 2008), impaired memory formation (Ventura et al. 2004; Brennan et al. 2006), decreased motor learning (Koekkoek et al. 2005), increased open-field hyperactivity (Yan et al. 2004) and abnormal marble burying (Paylor 2008), abnormal social behaviors (Paylor 2008; Mines et al. 2010; Spencer et al. 2008), abnormal prepulse inhibition (PPI; Paylor et al. 2008; de Vrij et al. 2008), prolonged epileptiform bursts (Subrathan et al. 2010), neuronal network hyperexcitability (Gibson et al. 2008), audiogenic seizures (Yan et al. 2005), abnormal growth patterns (Dolen et al. 2007), and increased protein synthesis (Qin et al. 2005).
Findings in the Drosophila model of FXS, in which there is loss of dfmr1 (homolog of the FMR1 gene in the Drosophila genome) activity (Zhang et al. 2001) parallel those in the knockout mouse and include defects in circadian rhythms, synaptic branching, courtship behavior, and learning (Zhang et al. 2001; McBride et al. 2005; Dockendorff et al. 2002; Morales et al. 2002). Furthermore, many phenotypic features of FXS in humans resemble or overlap those seen in the mouse and fly models, including seizures, electrical excitability on EEGs, hypersensitivity to tactile stimuli, cognitive difficulty, strabismus, enhanced anxiety, coordination problems and loose stools, and are effects that have been proposed to occur in a setting of exaggeration of mGluR-mediated protein synthesis-dependent processes that would normally be inhibited by FMRP (Bear et al. 2004; Bear 2005).
The pathways through which group 1 mGluRs and other forms of neural stimulation activate translation show abnormal activity in the fmr1 knockout, suggesting the activity levels in these pathways themselves, are under the control of FMRP. Group 1 mGluRs regulate dendritic translation through several pathways (Waung and Huber 2009). One pathway involves the activation of phospholipase C (PL-C), with resultant production of IP3 and DAG from PIP2, leading to the activation of PKC and induction of the mitogen-activated protein kinase kinase pathway through the phosphorylation of MEK, extracellular signal-regulated kinase (ERK), and Mnk1 in sequence, leading to translational activation through eIF4E. The second pathway involves signaling through homer-linked PIKE/PI3K, with conversion of PIP2 to PIP3, which then recruits pleckstrin homology-containing kinases PDK1 and Akt to the membrane where they are phosphorylated, with subsequent activation of mTOR and phosphorylation of 4EBP leading to the release of eIF4E to form eukaryotic initiation factor complex 4F and activation of CAP-dependent translation. mTOR activation also results in the phosphorylation of p70S6 kinase, which activates TOP-dependent translation. The PI3K enhancer, PIKE, is a predicted mRNA target for FMRP and in fact is found to be elevated in the fmr1 knockout, with resultant downstream activation of Akt, mTOR, S6 kinase, and 4EBP (Sharma et al. 2010). A third pathway involves direct Homer interaction and phosphorylation of EF2 kinase, which inhibits translational elongation and affects translational regulation (Waung and Huber 2009). Furthermore, excessive dephosphorylation of GSK3ß to the inactive form of the enzyme (presumably due to higher levels of phosphatase PP2A, in the fmr1 knockout) occurs in fmr1 knockout mouse brain, is dependent on elevated signaling through group 1 mGluRs, and may mediate and/or result from disturbances of translational regulation in the absence of FMRP (Min et al. 2009). Thus, in the fmr1 knockout, translational activation due to the absence of direct FMRP-mediated ribosomal inhibition and stalling is compounded by an increase in the activity of translation-activating cascades because of increased basal levels of synthesis of FMRP-target proteins that participate in those cascades.
Additional complex changes in molecular events at the synapse occur in the absence of FMRP, in many cases because of increased basal activity of proteins representing FMRP target proteins, lack of appropriate sensitivity to synaptic activation, direct protein–protein interactions of FMRP, or other yet unknown mechanisms. Examples of identified synaptic disturbances include FMRP interaction with dRac, cAMP, and the potassium channel Slack. The small GTPase dRac is a ligand of the fly dFMR protein and activates p21-activated kinases (PAKs). It has been proposed that excessive translation of Rac in the absence of repression by FMRP might lead to the upregulation of PAK, with resultant excessive phosphorylation of ERK (also observed in the fmr1 knockout), and this might mediate some of the cellular effects of FMRP deficiency (Lee et al. 2003; Hayashi et al. 2007). PAK associated with PSD95 at the synaptic density (although not total PAK) has been shown to be increased in the fmr1 knockout, but PAK fails to activate with theta burst afferent stimulation (TBS) as it does in wild-type mice, leading to defects in actin polymer stabilization, spine morphology, and TBS-induced hippocampal LTP (Chen et al. 2010). Cells from the fmr1 knockout mouse and from individuals with fragile X show reduced cAMP production (Chen et al. 2010; Berry-Kravis and Hicar 1995; Kelley et al. 2007) which is dependent on FMRP levels (Berry-Kravis and Ciurlionis 1998). Likewise, adenylate cyclase activity modulates mGluR-mediated regulation of FMRP activity (Wang et al. 2008a). Although the mechanism through which FMRP regulates cAMP production is not known, FMRP is known to bind adenylate cyclase subunit mRNAs (Darnell et al. 2010). FMRP has recently been shown, for the first time, to directly interact with a membrane protein by binding the C-terminus of the sodium-activated potassium channel Slack and activating the channel (Brown et al. 2010). This suggests that FMRP has synaptic activities that are distinct from the role in translational regulation and thus may be very diverse.
Numerous receptor system alterations have been reported in the fmr1 knockout, including the following examples. Synaptic GABA system proteins show decreased levels of GAD and GABA-A receptor subunits α1,3 and 4, ß1 and 2, and δ1 and 2 in several studies (D’Hulst et al. 2006; Gantois et al. 2006; D’Hulst and Kooy 2007). In another report, GABA-A α1, ß2 and δ, and GAD were downregulated at different specific times during development in the knockout (Adusei et al. 2010), overall suggesting the potential for reduced inhibitory signaling and inhibitory/excitatory imbalance in FXS. There is additional evidence for dopamine D1 receptor signaling deficits in the fmr1 knockout brain (Wang et al. 2008b). New evidence has recently emerged demonstrating that group I mGluR-dependent protein synthesis-independent endocannabinoid (eCB) LTD is abnormal in the fmr1 knockout mouse, likely due to abnormal regulation of production and levels of endocannabinoid 2-AG at synapses (Zhang and Alger 2010; Maccarrone et al. 2010). Since eCB-producing neurons are widely distributed in the brain, this may be an important mechanism impacting synaptic function in FXS
Targets, therapeutics, and preclinical work
FXS targeted treatments in models and man
dfmr mutant fly
Fmr1 KO mouse
Humans with FXS
(1)a Block translational signaling pathway—external
mGluR5 inhibition (MPEP, fenobam, STX107, AFQ056, RO4917523); MPEP used in models except where markeda
Courtship behavior—immediate recall and short-term memory; mushroom body formation; odor-shock memory; survival on glutamate-containing food
Audiogenic seizuresa; Epileptiform bursts; open-field hyperactivity; dendritic spine morphologya; amygdala mEPSP frequency; prepulse inhibitiona; marble buryinga
Fenobam—phase IIa single-dose open-label trial—PPI improved, anxiety reduced; AFQ056—phase IIb trial completed with improvement in fully methylated patients, phase III trial being initiated; RO4917523—phase II trial in progress STX107—phase I completed
mGluR5 inhibition by genetic reduction of mGluR5 receptors
Audiogenic seizures; dendritic spine density; excessive protein synthesis; abnormal growth pattern; ocular dominance plasticity; inhibitory avoidance extinction
(1)b Block translational signaling pathway- internal
Lithium (inhibition of GSK3β and PI turnover)
Courtship behavior—immediate recall and short-term memory; mushroom body formation
Audiogenic seizures; open-field hyperactivity; dendritic spine morphology; learning and anxiety deficits in the elevated plus maze, elevated zero maze, passive avoidance; social interaction deficit with new mice and anxiety-related behaviors during social interaction
Open label trial—behavioral improvement, some adaptive skills and verbal memory improved; ERK biomarker normalized
GSK3β inhibition (AR-A014418 or SB-216763)
PAK inhibition by genetic reduction of PAK
Dendritic spine morphology; Cortical LTP deficits; open-field hyperactivity, repetitive behaviors, center field anxiety deficit; Fear conditioning
PI3K inhibition (LY294002)
Dendritic spine morphology; mTOR overactivity
ERK/MEK inhibition (SL327)
Audiogenic seizures; protein synthesis
(2) Inhibit activity of individual FMRP-regulated proteins
Inhibit MMP9 (minocycline)
Dendritic spine morphology; anxiety in elevated plus maze; exploratory behavior in Y maze
Improvement in behavior in small open-label trial
Inhibit APP/Aβ with antibody or by genetic reduction of APP
Audiogenic seizures; dendritic spine morphology; marble burying
Inhibit STEP by genetic reduction of STEP
AMPA receptor internalization; audiogenic seizures; open-field hyperactivity
(3) Activate surface AMPA receptors
Ampakines (CX516, CX614)
CX614 increases BDNF which reverses impairments in hippocampal TBS-LTP
CX516—phase II trial—no cognitive or behavioral effects overall—dose too low but may have helped subjects co-treated with antipsychotics
(4) Other synaptic receptors/proteins
GABA-B agonists (baclofen, R-baclofen)
Survival on glutamate-containing food; memory deficits
Audiogenic seizures; open-field hyperactivity; marble burying
R-baclofen phase II trial—improvement in overall function, social and language function in more socially impaired subject group
GABA-A agonists (ganaxolone)
Open label trial -behavioral and social improvement
NMDA antagonists (memantine, acamprosate)
Memantine—small open-label trial—no overall improvement
Acamprosate—open-label trial in 3 patients with improved language and socialization
Glutamate uptake inhibition (riluzole)
Riluzole—small open-label trial—no overall improvement, ERK biomarker normalized
Signaling from receptor to dendritic translational machinery
Treatment targets in this category can be divided into those that act extracellularly on receptors to decrease translational signaling and those that act intracellularly on the signaling cascade. Extracellular agents generally target a particular receptor and thus are more specific treatments whose effects are often limited to brain or even just specific subsets of neurons. Intracellular agents have the advantage of potentially normalizing signaling resulting from the activation of the pathway through any receptor, and thus may target the mechanism more generally than receptor-specific molecules. On the other hand, intracellular agents predominantly target proteins with substantial overlap in multiple cellular signaling pathways, including those for growth and specification and other hormonal responses; therefore, these agents will be prone to an increased likelihood of off-target effects as well as cellular and organ toxicity.
Potential extracellular agents considered thus far include group 1 mGluR receptor (particularly mGluR5) blockers and muscarinic (M1) acetyl choline receptor blockers. Of the group 1 mGluRs, mGluR1 receptors are present mainly in the cerebellum and hippocampus, while mGluR5 receptors are present throughout the brain except in the cerebellum (Bear et al. 2004; Bear 2005). The broader distribution of mGluR5 receptors, in conjunction with toxicity in the form of motor deficits observed in animal models treated with mGluR1 blockers, has pointed to mGluR5 receptors as the better initial target for the pharmacotherapy of FXS.
In cultured neural cells, knockdown of FMRP using antisense oligonucleotides directed at FMR1 mRNA results in increased AMPA receptor (GluR1) internalization, presumably due to mGluR5 pathway overactivity as 2-methyl-6-(phenylethynyl)-pyridine (MPEP), an mGluR5 blocker, is able to reduce AMPA internalization back to normal levels (Nakamoto et al. 2007). This finding suggests that synaptic cellular deficits due to the absence of FMRP can be reversed by the inhibition of mGluR5-mediated activity. In the dfmr mutant fly, treatment with MPEP has been shown to reverse impairments in naïve courtship behavior, immediate recall and short-term memory (McBride et al. 2005), mushroom body formation (McBride et al. 2005), odor-shock memory (Bolduc et al. 2008), and survival on glutamate-containing food (Chang et al. 2008). MPEP and other mGluR5 negative modulators (including fenobam, AFQ056, and STX107) have been shown to reverse multiple phenotypes including audiogenic seizures (Paylor 2008; de Vrij et al. 2008), epileptiform bursts (Chuang et al. 2005), open-field hyperactivity (de Vrij et al. 2008), dendritic spine shape (Spencer et al. 2008), prepulse inhibition (Spencer et al. 2008), and behavioral phenotypes including abnormal marble burying (Paylor 2008). Treatment with MPEP, however, only partially corrects synaptic plasticity deficits in the fmr1 knockout amygdala, with failure to rescue LTP deficits and excessive AMPA receptor internalization, but successful rescue of mEPSC frequency (Subrathan et al. 2010).
Additional support for the mGluR5 receptor as a treatment target in FXS has come from experiments in which fmr1 knockout mice also heterozygous for a null mutation in the gene coding for the mGluR5 receptor were generated, resulting in a 50% reduction in mGluR5 expression, in addition to loss of FMRP in these animals (Dolen et al. 2007). Numerous phenotypes were rescued by genetically reducing mGluR5 expression in these fmr1 KO mice, including abnormal ocular dominance plasticity, increased density of dendritic spines on cortical pyramidal neurons, increased basal protein synthesis in the hippocampus, exaggerated inhibitory avoidance extinction, audiogenic seizures, and accelerated body growth (Dolen et al. 2007). The accumulated data showing both pharmacological and genetic reversal of fmr1 knockout phenotypes have provided substantial support for the proposal of Bear et al. (Bear et al. 2004, 2008; Bear 2005) that the excessive mGluR5 signaling in the fmr1 knockout model could be at least partially responsible for many of the psychiatric and neurological symptoms, such as poor cognitive development, seizures, anxiety, movement disorders (stereotypic motor movements), and even accelerated body growth seen in FXS.
It has been proposed that mGluR1 negative modulators might also be beneficial for development in FXS, but little preclinical information is available regarding the effects of these molecules in the fmr1 knockout. These molecules are associated with substantial negative cognitive and motor effects in normal rats (Kolasiewicz et al. 2009; Steckler et al. 2005) and hence are not in clinical development; however, they remain possible targeted treatments in FXS due to the specific disease defect of excessive signaling through mGluR1 receptors.
These agents target the pathway for derepression of translation after mGluR activation and include lithium, PI3K inhibitors, GSK3β inhibitors, and PAK inhibitors, among others. Lithium may attenuate activation of the PL-C signaling pathway by inhibiting phosphatidyl inositol (PI) turnover (Berridge 1993) and clearly inhibits GSK3β activity (Min et al. 2009; Yuskaitis et al. 2010), which would decrease phosphorylation of ERK and multiple signaling molecules that regulate translation; all of these effects would theoretically lead to the reduction of translational activation. Lithium has been shown to improve defects in naïve courtship behavior, immediate recall, and short-term memory in dfxr mutant flies (McBride et al. 2005) and to reverse phenotypes including audiogenic seizures (Min et al. 2009), open-field hyperactivity (Min et al. 2009; Liu et al. 2010), deficits on a social interaction task (Mines et al. 2010; Liu et al. 2010), learning deficits (Yuskaitis et al. 2010; Liu et al. 2010), anxiety (Yuskaitis et al. 2010; Liu et al. 2010), novel object recognition (Venkitaramani et al. 2010), and dendritic spine shape (Liu et al. 2010) in the fmr1 knockout mouse model. Given that lithium treatment normalizes levels of activated ERK and GSK3β in the fmr1 knockout (Venkitaramani et al. 2010), it appears that the main effect of lithium in the fmr1 knockout is to reduce excessive GSK3β activity with resultant reduction in excessive ERK-mediated translation; however, lithium may also directly increase surface expression of AMPA receptors (Du et al. 2010) and reduce excess MAP1B activity (Owen and Gordon-Weeks 2003). Other GSK3β inhibitors such as SB-216763 can reverse these phenotypes as well (Min et al. 2009). Effects of GSK3β inhibitors are not additive with those of mGluR5 blockers, suggesting that excess GSK3β activity is due to excessive activity in mGluR5-activated pathways (Min et al. 2009).
It was noted that PAK-deficient mice had dendritic spine pathology opposite that observed in the fmr1 knockout, with excessive short stubby spines and reduced density of spines. Hence, in double-mutant fmr1 knockout/PAK-deficient mice (expressing a dominant negative PAK transgene in the forebrain), rescue of fmr1 knockout phenotypes, including spine morphology abnormalities, cortical LTP deficits, and behavioral abnormalities, was observed (Hayashi et al. 2007). It was proposed that because PAK plays a role in ERK activation, PAK deficiency may correct excessive ERK activation and signaling and thus partially correct translational activation and resultant phenotypes in the fmr1 knockout. This is consistent with the finding that reduction of ERK activation with SL-327 normalizes protein synthesis and blocks audiogenic seizures in the fmr1 knockout mouse (Osterweil et al. 2010), and small molecules that inhibit ERK have been proposed as possible targeted therapeutics in FXS on the basis of these findings.
Elevated PI3K activity observed in the fmr1 knockout can be reduced using PI3K antagonist LY294002, resulting in the reversal of dendritic spine phenotypes (Gross et al. 2010), normalization of aberrant mTOR activation, and restoration of sensitivity of mTOR to mGluR activation with 3,4-dihydroxyphenylglycine (DHPG; Sharma et al. 2010). Thus, agents acting in this pathway including PI3K, mTOR, and S6 kinase inhibitors may also be potential treatment targets.
Taken together, the above findings suggest that deficits in receptor-activated signaling pathways in the absence of FMRP are beginning to be sufficiently well understood as to generate many potential targets for eventual treatment of fragile X syndrome.
Activity of individual proteins regulated by FMRP
Treatment targets in this category would be key synaptic proteins (other than those in the translational signaling pathway discussed above) for most of which the mRNA is an FMRP cargo, and the protein has been shown to be excessively translated in the absence of FMRP, with resultant elevated basal levels of protein, typically also with loss of responsiveness of protein translation/levels to mGluR activation with DHPG. Even though only one misregulated protein would be targeted, the protein would have sufficient key synaptic activity that correction of levels would be at least partially helpful in reversing phenotypes. Examples of targets in this category that have been studied include matrix metalloprotein-9 (MMP9; Bilousova et al. 2009), APP (Westmark and Malter 2007), and STEP kinase (Goebel-Goody et al. 2010). Excessive activity of MMP9 was demonstrated in the hippocampus of the fmr1 knockout mouse relative to wild type, and minocycline, an antibiotic that inhibits MMP9, currently used predominantly for acne in teenagers, was found to normalize dendritic spine phenotypes in both in vivo and in cultured hippocampal neurons from the fmr1 knockout as well as to improve anxiety in the elevated plus maze and exploratory behavior in the Y maze (Bilousova et al. 2009). APP and metabolites have been shown to be elevated in the fmr1 knockout, and either pharmacological (with antibodies to APP/Aβ) or genetic (creation of heterozygous APP null, fmr1 knockout double mutant) reduction of APP results in at least partial reversal of audiogenic seizure, behavioral, and spine phenotypes (Westmark et al. 2010). STEP levels are increased in the fmr1 knockout; in fact, STEP may be a key LTD protein produced by translational activation, and which then is responsible for the internalization of AMPA receptors. STEP/fmr1 double knockout animals show rescue of open-field hyperactivity and audiogenic seizure phenotypes seen in the fmr1 knockout, and STEP inhibitors have been proposed as a potential targeted treatment (Park et al. 2008).
An example of a different type of target would be a protein regulated by FMRP directly rather than through control of translation of its mRNA. One such example would be Slack, a sodium-activated potassium channel subunit that is activated by FMRP through direct interaction of FMRP with the C-terminal of Slack (Brown et al. 2010). In the fmr1 knockout, current through this channel is significantly reduced relative to wild type, suggesting that activators of Slack-containing channels might be a potential therapeutic strategy (Brown et al. 2010), although no studies of the therapeutic effects of such agents have yet been conducted.
Surface AMPA receptors and activity
Agents in this category are AMPA activators or “ampakines” and could be utilized to increase deficient synaptic AMPA receptor activity by direct positive modulation of receptor activity in the presence of glutamate and/or inducing BDNF to increase the number of surface AMPA receptors, thus normalizing deficient LTP. The so-called ampakine CX614 has been shown to increase BDNF in the hippocampus of the fmr1 knockout and wild-type mouse. BDNF, itself, reverses hippocampal TBS-induced LTP deficits in the fmr1 knockout (Lauterborn et al. 2007). Behavioral effects of these treatments have not yet been measured in the fmr1 knockout.
Other receptors/proteins that regulate synaptic activity
The main group of agents in this category that have been thus far studied in FXS animal models are GABAergic agents aimed at correcting the reduced activity of GABA inhibitory systems and reestablishing a balance between inhibition and excitation in the brain. GABA-A agonists would be directed at compensating for GABA-A subunit deficiencies (D’Hulst et al. 2006; Gantois et al. 2006), while GABA-B agonists act presynaptically to block glutamate release and thus would tend to decrease group I mGluR activation and excessive signaling, although it is likely that GABA-B agonists have a complex mechanism involving postsynaptic effects as well (Sohn et al. 2007). In the dfmr mutant fly, a variety of GABA agonists rescue the lethality phenotype from glutamate-containing food, as well as memory deficits and neuropathological phenotypes (Chang et al. 2008). In the fmr1 knockout mouse, racemic baclofen, a GABA-B agonist, rescues the audiogenic seizure phenotype (Pacey et al. 2009). The R-baclofen enantiomer of baclofen appears to have substantially better potency than S-baclofen, and R-baclofen reverses several behavioral phenotypes, including marble burying and open-field hyperactivity as well as audiogenic seizures (Paylor 2008). The GABA-A agonist ganaxolone (Carter et al. 1997) reduces audiogenic seizures in the fmr1 knockout (Kooy 2010), but no other effects have yet been explored.
Additional receptor targets would theoretically include mGluR2/3 agonists or antagonists, through mechanisms having to do with the regulation of synaptic glutamate levels, and blockers of endocannabinoid degradation, which would increase defective mGluR-dependent endocannabinoid-mediated LTD observed in the fmr1 knockout (Zhang and Alger 2010; Maccarrone et al. 2010). Preclinical testing of these molecules in the knockout has not yet been accomplished.
Translation of mRNAs regulated by FMRP using antisense technology
This strategy for developing treatment targets was recently proposed when it was discovered that mir125a, a small RNA involved in translational silencing of specific messages, operates in conjunction with phosphorylated FMRP and AGO2 to bind to PSD-95 mRNA and block translation of PSD-95. DHPG and anti-mir125a increase the translation of PSD-95 by dissociating mir125a from PSD-95 mRNA. In the absence of FMRP, mir125a is reduced in inhibitory complexes in synaptoneurosomes and PSD-95 translation is increased and uncoupled from mGluR activation (Muddashetty et al. 2010). This has suggested that the delivery of specific siRNAs like mir125a that target FMRP cargos to fmr1 knockout dendrites might be a new therapeutic strategy that could compensate for the absence of FMRP by reducing (normalizing) the translation of their target messages (Muddashetty and Bassell 2009).
Clinical trial experience with targeted treatments in humans with FXS
Taken together, the preclinical data with agents acting in the above mechanistic categories of targeted treatment have been very promising. Although there is much less information on the impact of treatment of humans with FXS with these compounds, early phase clinical trials have been initiated and even completed for treatments in many of the categories (listed in Table 1).
Signaling from receptor to dendritic translational machinery
Several mGluR5 negative modulators are currently being developed for the treatment of FXS and are in clinical trials, but are not available for general use in humans. Fenobam, the first mGluR blocker used in FXS, was administered in a single oral dose to 12 adult males and females with FXS (Berry-Kravis et al. 2009). In this trial, an improvement in the general anxiety level of many of the participants was observed after the dose, although given that the trial was open-label, this could have been a placebo effect. A significant improvement in PPI was also seen, which would be far less likely to be due to placebo effect. There were no safety concerns. Concurrently, a phase II double-blind placebo-controlled, crossover design trial of AFQ056 (Novartis) with treatment of 30 adult males with FXS for 28 days each with AFQ and placebo was conducted in Europe (Jacquemont et al. 2011). Results of this trial suggested improvement in maladaptive behavior on the ABC-C, CGI-I, and the Repetitive Behavior Scale in males with FXS and full methylation of FMR1, an outcome which was adequate to support ongoing development of AFQ056 for the treatment of FXS, with a larger multinational double-blind placebo-controlled 3-month trial evaluating the effects of multiple doses of the medication started in Fall of 2010 (clinicaltrials.gov). STX107 (Seaside Therapeutics) is completing phase I trials in healthy individuals and is expected to be ready for treatment trials in FXS in 2011 (clinicaltrials.gov). A double-blind placebo-controlled dose finding phase II trial of RO4917523 in adult males and females with FXS is underway and should be completed by late 2011 (clinicaltrials.gov). No serious safety concerns have yet emerged in any of these studies, although only small populations of individuals with FXS have thus far been exposed to mGluR5 blockers. It has been proposed that mGluR1 negative modulators might also be beneficial for individuals with FXS, but these have not been studied.
Although a number of intracellular treatment targets have been proposed, in most cases, safe and available agents acting on these targets are not yet developed for use in humans. One exception is lithium, for which the preclinical findings in the dfmr mutant fly and fmr1 knockout mouse suggested promise of therapeutic benefit. Although lithium has been used for some time to treat mood instability and aggression in FXS (Hagerman et al. 2009; Berry-Kravis and Potanos 2004; Wang et al. 2010a), only anecdotal information on effectiveness was available prior to a pilot study initiated by Berry-Kravis et al. (2008a) to test the concept of inhibition of mGluR-activated translational signaling pathways as a treatment strategy for FXS by systematically exploring the effects of a short-term (2 months) treatment with lithium on a broad range of phenotypes, including behavior, cognition, and biophysical measures, in a small cohort of subjects with FXS. In addition, since ERK was shown to be misregulated in the fmr1 knockout and to have a reduced activation rate in lymphocytes from humans with FXS (Weng et al. 2008), ERK activation was explored as a potential biomarker for the effects of lithium on cellular signaling and more generally as a model for measuring changes in signaling during treatment with agents that may impact receptor-activated translational regulatory pathways. In this pilot open-label lithium trial in 15 patients with FXS, significant improvement in behavior was seen in on the Total Aberrant Behavior Checklist-Community Edition (ABC-C) Score, the Maladaptive Behavior subscore from the Vineland Adaptive Behavior Scale, a parent visual analog scale for target behaviors, and the CGI. Improvement in verbal memory on the RBANS List Learning task was also demonstrated in addition to normalization of abnormal ERK phosphorylation rates in lymphocytes (Berry-Kravis et al. 2008a). There were no major side effects, but polydipsia and polyuria were seen relatively frequently as expected, and there were a few subjects with abnormal thyroid measurements on lithium. These data suggested that further studies with a placebo-controlled trial would be indicated; however, such studies have not yet been carried out, partly due to concerns about chronic toxicity of lithium, but also related to hope that less toxic mechanism-based treatments will be available soon.
PAK inhibitors are being generated for possible use in humans with FXS (Drug Discovery: PAK Program), however these are still in preclinical development. PI3K inhibitors and GSK3β inhibitors have been proposed and used in the fmr1 knockout, but are not yet in preclinical development for FXS.
Activity of individual proteins regulated by FMRP
Of synaptic protein targets thought to be regulated by FMRP and overexpressed in the fmr1 knockout and, by extension, in FXS, only MMP9 has had a pharmaceutical inhibitor sufficiently well developed at present for human trials. Minocycline has been used in clinical trials to inhibit MMP9 in FXS. An open-label survey study suggested improvements in anxiety and language in patients with FXS treated in clinic with minocycline (Utari et al. 2010). An initial open-label trial of minocycline in 20 participants with FXS over age 12 showed behavioral improvements on the ABC-C, VAS, and CGI (Paribello et al. 2010). There were no major side effects, but two individuals had to stop treatment due to elevated antinuclear antibodies despite lack of signs of drug-induced lupus. These results have been the rationale for an in-progress double-blind placebo-controlled trial of minocycline in children and adolescents with FXS age 5 and up. The major detriment to using minocycline in children <12 has been the likely side effect of yellow/brown discoloration of the permanent teeth. Other side effects that can be seen at any age include GI symptoms such as vomiting and/or diarrhea (Utari et al. 2010), drug-induced lupus, and pseudotumor cerebri. The placebo-controlled trial should help delineate the true margin of benefit provided by minocycline to help with decisions, especially for younger children, about whether benefits of minocycline treatment exceed the impact of the tooth discoloration side effect.
Antibodies to reduce toxic Aβ metabolites of APP have already been piloted in human phase II trials in Alzheimer’s disease (Frisardi et al. 2010). Such antibodies could also be used to reduce overexpressed Aβ in the human FXS brain, if justified by additional preclinical data in the knockout mouse.
Surface AMPA receptors and activity
A single human trial has been completed with CX516 (Cortex Phamaceuticals), a direct AMPA receptor-positive modulator or “ampakine” known to increase LTP and raise BDNF kevels, thus potentially increasing surface AMPA receptors (Jourdi et al. 2009). This was a double-blind placebo-controlled trial of effects of CX516 on safety and cognitive and behavioral efficacy measures carried out in a cohort of 49 individuals with FXS (Berry-Kravis et al. 2006). Conceptually, it was thought the CX516 would help compensate or correct the AMPA receptor deficit resulting from mGluR pathway overactivity. Realistically, CX516 is a very weak ampakine, and thus, no improvements were seen except in the group of patients co-treated with an antipsychotic (known to potentiate ampakine activity). This suggests that a more potent ampakine molecule might show success in FXS; however, such molecules have not yet come to clinical trials.
Other receptors/proteins that regulate synaptic activity
Based on animal data discussed in the previous section, coupled with anecdotal clinical experience suggesting behavioral benefits from racemic baclofen and data from TMS studies demonstrating enhancement of cortical inhibition by racemic baclofen (McDonnell et al. 2007), a clinical trial of R-baclofen in humans with FXS has been conducted. R-baclofen is the enantiomer of racemic baclofen with more potent GABA-B agonist activity. The double-blind placebo-controlled crossover trial, conducted by Seaside Therapeutics, involved 4-week periods of placebo and active drug treatment for each subject. This trial showed benefit for R-baclofen over placebo in global preference for treatment period and clinician global impression, particularly evident in subgroups of FXS patients with autism, more severe irritable behavior, or more severe social deficits. In the group with more impairment in social behaviors, improvements in the ABC Social Withdrawal Scale and Vineland Play and Leisure Scale were also seen (Wang et al. 2010c). There were no significant safety issues, and many subjects are continuing treatment through an extension study to evaluate long-term benefits and toxicity. Further development of R-baclofen is planned with additional clinical trials.
Other agents acting at an array of receptors have undergone an exploratory study in groups with FXS. These include donazepil, an anticholinesterase which raises acetyl choline in the brain and is extensively utilized for the maintenance of cognitive function in Alzheimer’s disease. Donazepil showed promise for the treatment of behavior and social function in an open-label trial in participants with FXS and now is being studied in a larger placebo-controlled trial (clinicaltrials.gov). A small open-label study of memantine, an NMDA antagonist, in six individuals with FXS showed modest clinical benefit on a CGI in 4/6 patients, but no improvement on behavioral rating scales, and two patients developed significant irritability that limited treatment (Erickson et al. 2009). An open-label study of riluzole, a sodium channel blocker and glutamate uptake activator that indirectly decreases glutamate receptor activity, in five patients with FXS showed overall behavioral improvement in only one subject, although ERK activation rates normalized and there was a suggestion of improvement specifically in hyperactivity symptoms (Erickson et al. 2011). An anecdotal treatment experience has been reported with three adults with FXS treated with acamprosate, a drug approved for assisting with alcohol withdrawal that most likely interacts with multiple receptors but primarily may exert effects by acting as a mixed agonist/antagonist at NMDA receptors, activating GABA-A receptors and possibly inhibiting group I mGluRs. This case series observed improvement in language and behavior in all patients (Erickson et al. 2010a). One patient experienced limiting gastrointestinal side effects that are commonly seen with acamprosate. Although aripiprazole is a treatment directed primarily at behavior rather than molecular mechanism, it could theoretically be targeted to dopamine deficits thought to be present in FXS (Wang et al. 2008b), given its dopamine agonist activity at lower doses. Apipirazole has shown good success when used empirically in FXS clinic populations (Hagerman et al. 2009; Berry-Kravis and Potanos 2004), and resulted in improvement in ABC irritability, other ABC subscores and other behavioral rating scales in 15 individuals with FXS treated in a very recently completed open-label trial (Erickson et al. 2010b). Plans are underway to initiate a double-blind placebo-controlled trial of aripiprazole.
Translation of mRNAs regulated by FMRP using antisense technology
Treatments utilizing this strategy have just recently been initiated in preclinical animal models of FXS and are not in development yet for humans.
Trial design and hurdles to address
Although many neuronal targets for treating the underlying disorder in FXS have emerged, and early translational work has begun, there are still many uncertainties about how to optimally demonstrate treatment effects in a clinical trial setting. These issues surrounding clinical trial design have not yet been worked out, nor are there models for cognitive treatment trials for any neurodevelopmental disorders. FXS, in fact, serves as a good model to develop such designs, particularly because FXS is a single genetic disorder in which all affected individuals have the same basic cellular defect as the primary cause of their brain disorder, a mouse model is available, some information on the mechanisms of the synaptic function of FMRP in brain is known, and aspects of FXS model more common disorders with likely mechanistic overlap, including autistic spectrum disorders, ADHD, and non-verbal learning disability.
Trial design issues that need to be resolved for each targeted treatment trial in FXS include: (1) length of placebo treatment and use of crossover designs or open-label extensions to ensure everyone gets a chance at drug and increase recruitment; (2) lack of information on optimal dosing and whether to determine this through dose escalation or flexible dosing within or between subjects or multiple fixed-dose arms; (3) how to best detect side effects in cognitively impaired individuals who may not be able to report symptoms accurately; (4) the most appropriate age range to study treatment effects, balancing concerns about safety that dictate adult trials first, with the possibility that rapid movement to younger ages may be needed despite minimal effects in older individuals because much more significant results may be seen by treating the underlying disorder in young children who are not as far along in the process of brain wiring and development and are still in school; (5) lack of understanding the length of treatment needed to impact brain wiring and demonstrate measurable cognitive improvement; (6) drug formulation and how to best deliver drug to younger children and individuals with oromotor dyspraxia; (7) inclusion of females and mosaic individuals, and whether to analyze response separately in these groups, as individuals with FMRP present in a fraction of cells may have different dosing ranges and differing toxicities; (8) whether to allow baseline medications and the balance between the need to analyze treatment effects in the absence of medication interactions, problems with recruitment and patient discomfort if baseline medications have to be weaned, and the importance of demonstrating that new targeted treatments can actually improve symptoms even when treatment with the best available symptomatic regimen is already in place; (9) numbers of study visits and travel issues for a disorder in which subject numbers are limited and participants may need to come to trail sites from far away; and (10) the problem of lack of validated sensitive outcome measures for behavior or especially for cognition in FXS (or other developmental disorders) and lack of biomarkers known to correlate with functional improvement.
The design and evaluation of outcome measures for trials in FXS and neurodevelopmental disorders represents the most significant hurdle in trial design for targeted treatments. Choice of optimal outcome measures has been plagued by the need to test a broad ability range to prevent low- or high-functioning individuals from showing ceiling or floor effects, by problems with cooperation and variable performance, lack of knowledge about reproducibility of measures, and by the need to find measures that quantify core defects and correlate with quality of life and true functional improvement. Only a subset of outcome measures utilized in recent trials have turned out to be feasible and produce valid assessments (Berry-Kravis et al. 2006, 2008a, b, 2009). Only recently have investigators begun to develop templates for pre-trial feasibility, reproducibility, and validity assessment (Hessl et al. 2008a; Berry-Kravis et al. 2008b; Farzin et al. 2011; Knox and Berry-Kravis 2009). Choice of outcome measures must also balance the use of accepted behavioral measures which are generally caregiver rating scales (such as the ABC) with precedent for use in drug registration/FDA approval versus use of novel measures (Hessl et al. 2008a; Farzin et al. 2011; Knox and Berry-Kravis 2009) that are more quantitative and may objectively measure core phenotypes and electrophysiology (such as eye tracking or PPI), thus advancing treatment science, but have no precedent for registration and are difficult to use to predict a specific functional outcome. Outcome measure development for targeted treatment trials in FXS has been a sufficiently difficult problem that a series of NIH meetings have been convened specifically to address this issue, with participation from the FDA (NIH 2009). These meetings have resulted in recommendations about best choice of currently existing measures, validation needs for existing measures, optimal types of measures to be developed, and work that needs to be done to develop them. No one behavioral scale was felt to capture the range and character of behaviors typically problematic in individuals with FXS, and development and validation of a fragile X-specific behavioral scale has been suggested. Initial work in preparation for the development of such a scale has been done by collecting ABC ratings from multiple sites and looking at items most endorsed for affected individuals with FXS relative to age and gender. This early work has indicated that the factor structure and items incorporated into the ABC (which was developed for individuals with general cognitive impairment and has been used extensively in autism) needs to be modified for optimal validity in FXS (Hessl et al. 2010).
Several years ago, the Fragile X Clinical and Research Consortium (FXCRC) was created with the help of CDC funding to help ensure state-of-the-art care delivery to meet the needs of individuals with FXS across the country and facilitate large-scale national research efforts. This organization will also allow FXS Clinics across North America to collaborate in preparation for large multi-site clinical trials that will be necessary for FDA approval of targeted treatments.
Generalizing the FXS model to autism and other neurodevelopmental disorders
This work was supported in part by grants to Dr. Berry-Kravis from from the Spastic Paralysis and Allied Diseases of the Central Nervous System Research Foundation of The Illinois-Eastern Iowa District Kiwanis International, FRAXA Research Foundation, and the National Fragile X Foundation.
- Adusei DC, Pacey LK, Chen D, Hampson DR. Early developmental alterations in GABAergic protein expression in fragile X knockout mice. Neuropharmacology. 2010;59:167–71.PubMedGoogle Scholar
- Antar LN, Afroz R, Dictenberg JB, Carroll RC, Bassell GJ. Metabotropic glutamate receptor activation regulates fragile X mental retardation protein and FMR1 mRNA localization differentially in dendrites and at synapses. J Neurosci. 2004;24:2648–55.PubMedGoogle Scholar
- Aschrafi A, Cunningham BA, Edelman GM, Vanderklish PW. The fragile X mental retardation protein and group I metabotropic glutamate receptors regulate levels of mRNA granules in brain. Proc Natl Acad Sci USA. 2005;102:2180–5.PubMed CentralPubMedGoogle Scholar
- Awadalla P, Gauthier J, Myers RA, Casals F, Hamdan FF, Griffing AR, et al. Direct measure of the de novo mutation rate in autism and schizophrenia cohorts. Am J Hum Genet. 2010;87:316–24.PubMed CentralPubMedGoogle Scholar
- Bagni C, Greenough WT. From mRNP trafficking to spine dysmorphogenesis: the roots of fragile X syndrome. Nat Rev Neurosci. 2005;6:376–87.PubMedGoogle Scholar
- Bakker CE, Oostra BA. Understanding fragile X syndrome: insights from animal models. Cytogenet Genome Res. 2003;100:111–23.PubMedGoogle Scholar
- Bassell GJ, Warren ST. Fragile X syndrome: loss of local mRNA regulation alters synaptic development and function. Neuron. 2008;60:201–14.PubMed CentralPubMedGoogle Scholar
- Bear MF. Therapeutic implications of the mGluR theory of fragile X mental retardation. Genes Brain Behav. 2005;4:393–8.PubMedGoogle Scholar
- Bear MF, Huber KM, Warren ST. The mGluR theory of fragile X mental retardation. Trends Neurosci. 2004;27:370–7.PubMedGoogle Scholar
- Bear MF, Dolen G, Osterweil E, Nagarajan N. Fragile X: translation in action. Neuropsychopharmacology. 2008;33:84–7.PubMed CentralPubMedGoogle Scholar
- Berridge MJ. Inositol trisphosphate and calcium signalling. Nature. 1993;361:315–25.PubMedGoogle Scholar
- Berry-Kravis E, Ciurlionis R. Overexpression of fragile X gene (FMR-1) transcripts increases cAMP production in neural cells. J Neurosci Res. 1998;51:41–8.PubMedGoogle Scholar
- Berry-Kravis E, Hicar M. Reduced cyclic AMP production in fragile X syndrome: cytogenetic and molecular correlations. Pediatr Res. 1995;38:638–43.PubMedGoogle Scholar
- Berry-Kravis E, Potanos K. Pschopharmacology in fragile X syndrome—present and future. Ment Retard Dev Disabil Res Rev. 2004;10:42–8.PubMedGoogle Scholar
- Berry-Kravis E, Krause SE, Block SS, Guter S, Wuu J, Leurgans S, et al. Effect of CX516, an AMPA-modulating compound, on cognition and behavior in fragile X syndrome: a controlled trial. J Child Adolesc Psychopharm. 2006;16:525–40.Google Scholar
- Berry-Kravis E, Abrams L, Coffey S, Hall D, Greco C, Gane L, et al. Fragile X-associated tremor/ataxia syndrome (FXTAS): clinical features, genetics and testing guidelines. Mov Disord. 2007;22:2018–30.PubMedGoogle Scholar
- Berry-Kravis E, Sumis A, Hervey C, Nelson M, Porges SW, Weng N, et al. Open-label treatment trial of lithium to target the underlying defect in fragile X syndrome. J Dev Behav Pediatr. 2008a;29:293–302.PubMedGoogle Scholar
- Berry-Kravis E, Lara R, Kim O-K, Sumis A, Wuu J. Characterization of potential outcome measures for future clinical trials in fragile X syndrome. J Autism Dev Disord. 2008b;38:1751–7.PubMedGoogle Scholar
- Berry-Kravis E, Hessl D, Coffey S, Hervey C, Schneider A, Yuhas J, et al. A pilot open label, single dose trial of fenobam in adults with fragile X syndrome. J Med Genet. 2009;46:266–71.PubMed CentralPubMedGoogle Scholar
- Bilousova TV, Dansie L, Ngo M, Aye J, Charles JR, Ethell DW, et al. Minocycline promotes dendritic spine maturation and improves behavioural performance in the fragile X mouse model. J Med Genet. 2009;46:94–102.PubMedGoogle Scholar
- Blackwell E, Zhang X, Ceman S. Arginines of the RGG box regulate FMRP association with polyribosomes and mRNA. Hum Mol Genet. 2010;19:1314–23.PubMed CentralPubMedGoogle Scholar
- Boccia JL, Roberts JE. Behavior and autonomic nervous system function assessed via heart period measures: the case of hyperarousal in boys with fragile X syndrome. Behav Res Meth Instrum Comput. 2000;32:5–10.Google Scholar
- Bolduc FV, Bell K, Cox H, Broadie KS, Tully T. Excess protein synthesis in Drosophila fragile X mutants impairs long-term memory. Nat Neurosci. 2008;11(10):1143–5.PubMed CentralPubMedGoogle Scholar
- Brennan FX, Albeck DS, Paylor R. Fmr1 knockout mice are impaired in a leverpress escape/avoidance task. Genes Brain Behav. 2006;5:467–71.PubMedGoogle Scholar
- Brown V, Jin P, Ceman S, Darnell JC, O’Donnell WT, Tenenbaum SA, et al. Microarray identification of FMRP associated brain mRNAs and altered mRNA translational profiles in fragile X syndrome. Cell. 2001;107:477–87.PubMedGoogle Scholar
- Brown M, Kronengold J, Gazula V-R, Chen Y, Strumbos JG, Sigworth FJ, et al. Fragile X mental retardation protein controls gating of the sodium-activated potassium channel Slack. Nat Neurosci. 2010;13:819–21.PubMed CentralPubMedGoogle Scholar
- Carter RB, Wood PL, Wieland S, Hawkinson JE, Belelli D, Lambert JJ, et al. Characterization of the anticonvulsant properties of ganaxolone (CCD 1042; 3alpha-hydroxy-3beta-methyl-5alpha-pregnan-20-one), a selective, high-affinity, steroid modulator of the gamma-aminobutyric acid(A) receptor. J Pharmacol Exp Ther. 1997;280:1284–95.PubMedGoogle Scholar
- Ceman S, Brown V, Warren ST. Isolation of an FMRP-associated messenger ribonucleoprotein particle and identification of nucleolin and the fragile X-related proteins as components of the complex. Mol Cell Biol. 1999;19:7925–32.PubMed CentralPubMedGoogle Scholar
- Chang S, Bray SM, Li Z, Zarnescu DC, He C, Jin P, et al. Identification of small molecules rescuing fragile X syndrome phenotypes in Drosophila. Nat Chem Biol. 2008;4:256–63.PubMedGoogle Scholar
- Cheever A, Ceman S. Phosphorylation of FMRP inhibits association with Dicer. RNA. 2009;15:362–6.PubMed CentralPubMedGoogle Scholar
- Chen L, Yun SW, Seto J, Liu W, Toth M. The fragile X mental retardation protein binds and regulates a novel class of mRNAs containing U rich target sequences. Neuroscience. 2003;120:1005–17.PubMedGoogle Scholar
- Chen LY, Rex CS, Babayan AH, Kramar EA, Lynch G, Gall CM, et al. Physiological activation of synaptic Rac>PAK (p-21 activated kinase) signaling is defective in a mouse model of fragile X syndrome. J Neurosci. 2010;30:10977–84.PubMed CentralPubMedGoogle Scholar
- Chuang SC, Zhao W, Bauchwitz R, Yan Q, Bianchi R, Wong RK. Prolonged epileptiform discharges induced by altered group I metabotropic glutamate receptor-mediated synaptic responses in hippocampal slices of a fragile X mouse model. J Neurosci. 2005;25:8048–55.PubMedGoogle Scholar
- Comery TA, Harris JB, Willems PJ, Oostra BA, Irwin SA, Weiler IJ, et al. Abnormal dendritic spines in fragile X knockout mice: maturation and pruning deficits. Proc Natl Acad Sci USA. 1997;94:5401–4.PubMed CentralPubMedGoogle Scholar
- Cornish K, Kogan C, Turk J, Manly T, James N, Mills A, et al. The emerging fragile X premutation phenotype: evidence from the domain of social cognition. Brain Cogn. 2005;57:53–60.PubMedGoogle Scholar
- Darnell JC, Jensen KB, Jin P, Brown V, Warren ST, Darnell RB. Fragile X mental retardation protein targets G quartet mRNAs important for neuronal function. Cell. 2001;107:489–99.PubMedGoogle Scholar
- Darnell JC, van Dreische S, Zhang C, Mele A, Zang JB, Fak JJ, et al. HITS-CLIP identifies specific neuronal mRNA targets of translational repression by the fragile X mental retardation protein, FMRP. Keystone Symposia, Synapses: Formation, Function, and Misfunction, April 14, 2010. Snowbird, Utah.Google Scholar
- D’Hulst C, Kooy RF. The GABAA receptor: a novel target for treatment of fragile X? Trends Neurosci. 2007;30:425–31.PubMedGoogle Scholar
- D’Hulst C, De Geest N, Reeve SP, Van Dam D, De Deyn PP, Hassan BA, et al. Decreased expression of the GABAA receptor in fragile X syndrome. Brain Res. 2006;1121:238–45.PubMedGoogle Scholar
- de Vries BB, Wiegers AM, Smits AP, Mohkamsing S, Duivenvoorden HJ, Fryns JP, et al. Mental status of females with an FMR1 gene full mutation. Am J Hum Genet. 1996;58:1025–32.PubMed CentralPubMedGoogle Scholar
- de Vrij FM, Levenga J, van der Linde HC, Koekkoek SK, De Zeeuw CI, Nelson DL, et al. Rescue of behavioral phenotype and neuronal protrusion morphology in Fmr1 KO mice. Neurobiol Dis. 2008;31:127–32.PubMed CentralPubMedGoogle Scholar
- Devys D, Lutz Y, Rouyer N, Bellocq JP, Mandel JL. The FMR-1 protein is cytoplasmic, most abundant in neurons and appears normal in carriers of a fragile X premutation. Nat Genet. 1993;4:335–40.PubMedGoogle Scholar
- Dockendorff TC, Su HS, McBride SM, Yang Z, Choi CH, Siwicki KK, et al. Drosophila lacking dfmr1 activity show defects in circadian output and fail to maintain courtship interest. Neuron. 2002;34:973–84.PubMedGoogle Scholar
- Dolen G, Osterweil E, Shankaranarayana Rao BS, Smith GB, Auerbach BD, Chattarji S, et al. Correction of fragile X syndrome in mice. Neuron. 2007;56:955–62.PubMed CentralPubMedGoogle Scholar
- Drug Discovery: PAK Program. http://www.afraxis.com.
- Du J, Wei Y, Liu L, Wang Y, Khairova R, Blumenthal R, et al. A kinesin singnaling complex mediate the ability of GSK-3beta to affect mood-associated behaviors. Proc Natl Acad Sci USA. 2010;107:11573–8.PubMed CentralPubMedGoogle Scholar
- Erickson CA, Mullett JE, McDougle CJ. Open-label memantine in fragile X syndrome. J Autism Dev Disord. 2009;39:1629–35.PubMedGoogle Scholar
- Erickson CA, Mullett JE, McDougle CJ. Brief report: Acamprosate in fragile X syndrome. J Autism Dev Disord. 2010a;40:1412–6.Google Scholar
- Erickson CA, Stigler KA, Posey DJ, McDougle CJ. Aripiprazole in autism spectrum disorders and fragile X syndrome. Neurotherapeutics. 2010b;7:258–63.Google Scholar
- Erickson CA, Weng N, Weiler IJ, Greenough WT, Stigler KA, Wink LK, McDougle CJ. Open-label riluzole in fragile X syndrome. Brain Res. 2011. doi:10.1016/j.brainres.2010.10.108.
- Farzin F, Rivera SM, Hessl D. Brief report: Visual processing of faces in individuals with fragile X syndrome: an eye tracking study. J Autism Dev Disord. 2009;39:946–52.PubMed CentralPubMedGoogle Scholar
- Farzin F, Scaggs F, Hervey C, Hessl D, Berry-Kravis E. Feasibility, reliability and clinical validity of eye-tracking and pupillometry as an outcome measure for clinical trials in fragile X syndrome (FXS). J Autism Dev Disord. 2011 (in press).Google Scholar
- Feng Y, Absher D, Eberhart DE, Brown V, Malter HE, Warren ST. FMRP associates with polyribosomes as an mRNP, and the I304N mutation of severe fragile X syndrome abolishes this association. Mol Cell. 1997;1:109–18.PubMedGoogle Scholar
- Frisardi V, Solfrizzi V, Imbimbo PB, Capurso C, D’Introno A, Colacicco AM, et al. Towards disease-modifying treatment of Alzheimer’s disease: drugs targeting beta-amyloid. Curr Alzheimer Res. 2010;7(1):40–55.PubMedGoogle Scholar
- Gantois I, Vandesompele J, Speleman F, Reyniers E, D’Hooge R, Severijnen LA, et al. Expression profiling suggests underexpression of the GABA(A) receptor subunit delta in the fragile X knockout mouse model. Neurobiol Dis. 2006;21:346–57.PubMedGoogle Scholar
- Gibson JR, Bartley AF, Hays SA, Huber KM. Imbalance of neocortical excitation and inhibitionand altered UP states reflect network hyperexcitability in the mouse model of fragile X syndrome. J Neurophys. 2008;100:2615–26.Google Scholar
- Goebel-Goody SM, Wallis ED, Li DV, Hall V, Royston S, Baum M, et al. 2010. Loss of striatal-enriched protein tyrosine phosphatase (STEP) reverses deficits in a fragile x syndrome mouse model. Program No. 452.4. 2010 Neuroscience Meeting Planner. San Diego, CA: Society for Neuroscience (online).Google Scholar
- Gross C, Nakamoto M, Yao X, Chan C-B, Yim SY, Ye K, et al. Excess phosphoinositide 3-kinase subunit synthesis and activity as a novel therapeutic target in fragile X syndrome. J Neurosci. 2010;30:10624–38.PubMed CentralPubMedGoogle Scholar
- Grossman AW, Aldridge GM, Weiler IJ, Greenough WT. Local protein synthesis and spine morphogenesis: fragile X syndrome and beyond. J Neurosci. 2006;26:7151–5.PubMedGoogle Scholar
- Hagerman PJ. The fragile X prevalence paradox. J Med Genet. 2008;45:498–9.PubMed CentralPubMedGoogle Scholar
- Hagerman RJ, Berry-Kravis E, Ono MY, Tartaglia N, Lachiewicz A, Kronk B, et al. Advances in the treatment of fragile X syndrome. J Pediatr. 2009;123:378–90.Google Scholar
- Hayashi ML, Rao BS, Seo JS, Choi HS, Dolan BM, Choi SY, et al. Inhibition of p21-activated kinase rescues symptoms of fragile X syndrome in mice. Proc Natl Acad Sci USA. 2007;104:11489–94.PubMed CentralPubMedGoogle Scholar
- Hessl D, Tassone F, Loesch DZ, Berry-Kravis E, Leehey MA, Gane LW, et al. Abnormal elevation of FMR1 mRNA is associated with psychological symptoms in individuals with the fragile X premutation. Am J Med Genet B Neuropsychiatr Genet. 2005;139:115–21.Google Scholar
- Hessl D, Cirdeiro L, Yuhas J, Campball A, Ornitz E, Berry-Kravis E, et al. Prepulse inhibition in fragile X syndrome: feasibility, reliability, and implications for treatment. Am J Med Genet. 2008a;153B:545–53.Google Scholar
- Hessl D, Tassone F, Cordiero L, Koldewyn K, McCormick C, Green C, et al. Brief report: Aggression and stereotypic behavior in males with fragile X syndrome—moderating secondary genes in a “single gene” disorder. J Autism Dev Disord. 2008b;38:184–9.Google Scholar
- Hessl D, Maltas S, Hall S, Kaufmann W, Lachiewicz A, et al. 2010. Toward a fragile X behavior rating scale: the aberrant behavior checklist collaborative study. 12th International Fragile X Conference, 21 July 2010, Detroit, Michigan.Google Scholar
- Heulens I, Kooy F. Fragile X syndrome: from gene discovery to therapy. Front Biosci. 2011;16:1211–32.Google Scholar
- Huber KM, Gallagher SM, Warren ST, Bear MF. Altered synaptic plasticity in a mouse model of fragile X mental retardation. Proc Natl Acad Sci USA. 2002;99:7746–50.PubMed CentralPubMedGoogle Scholar
- Irwin SA, Patel B, Idupulapati M, Harris JB, Crisostomo RA, Larsen BP, et al. Abnormal dendritic spine characteristics in the temporal and visual cortices of patients with fragile-X syndrome: a quantitative examination. Am J Med Genet A. 2001;98:161–7.Google Scholar
- Irwin SA, Idupulapati M, Gilbert ME, Harris JB, Chakravarti AB, Rogers EJ, et al. Dendritic spine and dendritic field characteristics of later V pyramidal neurons in the visual cortex of fragile X-knockout mice. Am J Med Genet. 2002;111:140–6.PubMedGoogle Scholar
- Jacquemont S, Curie A, des Portes V, Berry-Kravis E, Hagerman RJ, Ramos F, et al. Epigenetic modification of the FMR1 gene in fragile X syndrome is associated with differential response to the mGluR5 antagonist AFQ056. Sci Transl Med. 2011;3:64ra1.PubMedGoogle Scholar
- Jourdi H, Hsu YT, Zhou M, Qin Q, Bi X, Baudry M. Positive AMPA receptor modulating rapidly stimulates BDNF release and increases dendritic mRNA translation. J Neurosci. 2009;29(8):8688–97.PubMed CentralPubMedGoogle Scholar
- Kelley DJ, Davidson RJ, Elliott JL, Lahvis GP, Yin JC, Bhattacharyya A. The cyclic AMP cascade is altered in fragile X nervous system. PLoS ONE. 2007;26:e931.Google Scholar
- Knox A, Berry-Kravis E. Feasibility and reproducibility of the Tests of Attentional Performance for Children (KiTAP) in fragile X syndrome (FXS). Ann Neurol. 2009;66:S108.Google Scholar
- Koekkoek SK, Yamaguchi K, Milojkovic BA, Dortland BR, Ruigrok TJ, Maex R, et al. Deletion of FMR1 in Purkinje cells enhances parallel fiber LTD, enlarges spines and attenuates cerebellar eyelid conditioning in fragile X syndrome. Neuron. 2005;47:339–52.PubMedGoogle Scholar
- Kolasiewicz W, Kuter K, Wardas J, Ossowska K. Role of the metabotropic glutamate receptor subtype 1 in the Harmaline-induced tremor in rats. J Neural Transm. 2009;116:1059–63.PubMedGoogle Scholar
- Kooy, F. 2010. The GABAA receptor as a potential target for therapy of the fragile X syndrome. The 12th International Fragile X Conference, 22 July 2010, Detroit, Michigan.Google Scholar
- Larson J, Jessen RE, Kim D, Fine AK, du Hoffman J. Age-dependent and selective impairment of long-term potentiation in the anterior piriform cortex of mice lacking the fragile X mental retardation protein. J Neurosci. 2005;25:9460–9.PubMedGoogle Scholar
- Larson J, Kim D, Patel RC, Floreani C. Olfactory discrimination learning in mice lacking the fragile X mental retardation protein. Neurobiol Learn Mem. 2008;90:90–102.PubMed CentralPubMedGoogle Scholar
- Lauterborn JC, Rex CS, Kramar E, Chen LY, Pandyarajan V, Lynch G, et al. Brain-derived neurotrophic factor rescues synaptic plasticity in a mouse model of fragile X syndrome. J Neurosci. 2007;27(40):10685–94.PubMedGoogle Scholar
- Lee A, Li W, Xu K, Bogert BA, Su K, Gao FB. Control of dendritic development by the Drosophila fragile X-related gene involves the small GTPase Rac1. Development. 2003;130:5543–52.PubMedGoogle Scholar
- Levenga J, de Vrig FMS, Oostra BA, Willamson R. Potential therapeutic interventions for fragile X syndrome. Trend Mol Med. 2010;16:516–27.Google Scholar
- Li J, Pelletier MR, Perez Velazquez JL, Carlen PL. Reduced cortical synaptic plasticity and GluR1 expression associated with fragile X mental retardation protein deficiency. Mol Cell Neurosci. 2002;19:138–51.PubMedGoogle Scholar
- Liu ZH, Chuang DM, Smith CB. Lithium ameliorates phenotypic deficits in a mouse model of fragile X syndrome. Neuropsychopharmacology. 2010;25:1–13.Google Scholar
- Loesch DZ, Huggins RM, Bui QM, Epstein JL, Taylor AK, Hagerman RJ. Effect of the deficits of fragile X mental retardation protein on cognitive status of fragile X males and females assessed by robust pedigree analysis. J Dev Behav Pediatr. 2002;23:416–23.PubMedGoogle Scholar
- Loesch DZ, Huggins RM, Hagerman RJ. Phenotypic variation and FMRP levels in fragile X. Ment Retard Dev Disabil Res Rev. 2004;10:31–41.PubMedGoogle Scholar
- Maccarrone M, Rossi S, Bari M, De Chiara V, Rapino C, Musella A, et al. Abnormal mGlu 5 receptor/endocannabinoid coupling in mice lacking FMRP and BC1 RNA. Neuropsychopharmacology. 2010;35:1500–9.PubMed CentralPubMedGoogle Scholar
- Marshall CR, Noor A, Vincent JB, Lionel AC, Feuk L, Skaug J, et al. Structural variation of chromosomes in autism spectrum disorder. Am J Hum Genet. 2008;82:477–88.PubMed CentralPubMedGoogle Scholar
- McBride SM, Choi CH, Wang Y, Liebelt D, Braunstein E, Ferreiro D, et al. Pharmacological rescue of synaptic plasticity, courtship behavior, and mushroom body defects in a model of fragile X syndrome. Neuron. 2005;45:753–64.PubMedGoogle Scholar
- McDonnell MN, Orekhov Y, Ziemann U. Suppression of LTP-like plasticity in human motor cortex by the GABAB receptor agonist baclofen. Exp Brain Res. 2007;180:181–6.PubMedGoogle Scholar
- Miller LJ, McIntosh DN, McGrath J, Shyu V, Lampe M, Taylor AK, et al. Electrodermal responses to sensory stimuli in individuals with fragile X syndrome: a preliminary report. Am J Med Genet. 1999;83:268–79.PubMedGoogle Scholar
- Min WW, Yuskaitis CJ, Yan Q, Sikorski C, Chen S, Jope RS, et al. Elevated glycogen synthase kinase-3 activity in fragile X mice: key metabolic regulator with evidence for treatment potential. Neuropharmacology. 2009;56:463–72.PubMed CentralPubMedGoogle Scholar
- Mines MA, Yuskaitis CJ, Beurak E, Jope RS. GSK3 influences social preference and anxiety-related behaviors during social interaction in a mouse model of fragile X syndrome and autism. PLoS ONE. 2010;5:e9706.PubMed CentralPubMedGoogle Scholar
- Miyashiro KY, Beckel-Mitchener A, Purk TP, Becker KG, Barret T, Liu L, et al. RNA cargoes associating with FMRP reveal deficits in cellular functioning in Fmr1 null mice. Neuron. 2003;37:417–31.PubMedGoogle Scholar
- Morales J, Hiesinger PR, Schroeder AJ, Kume K, Verstreken P, Jackson FR, et al. Drosophila fragile X protein, DFXR, regulates neuronal morphology and function in the brain. Neuron. 2002;34:961–72.PubMedGoogle Scholar
- Muddashetty R, Bassell GJ. A boost in microRNAs shapes up the synapse. EMBO J. 2009;28:617–8.PubMed CentralPubMedGoogle Scholar
- Muddashetty RS, Kelic S, Gross C, Xu M, Bassell GJ. Dysregulated metabotropic glutamate receptor-dependent translation of AMPA receptor and postsynaptic density-95 mRNAs at synapses in a mouse model of fragile X syndrome. J Neurosci. 2007;16(27):5338–48.Google Scholar
- Muddashetty RS, Nalavadi VC, Gross C, Yao X, Warren ST, Bassell GJ. Reversible inhibition of PSD-95 mRNA translation by miR-125a, FMRP phosphorylation and mGluR signaling. Program No. 548.2. 2010 Neuroscience Meeting Planner. San Diego, CA: Society for Neuroscience, 2010 (online).Google Scholar
- Nakamoto M, Nalavadi V, Epstein MP, Narayanan U, Bassell GJ, Warren ST. Fragile X mental retardation protein deficiency leads to excessive mGluR5-dependent internalization of AMPA receptors. Proc Natl Acad Sci USA. 2007;104:15537–42.PubMed CentralPubMedGoogle Scholar
- Narayanan U, Nalavadi V, Nakamoto M, Thomas G, Ceman S, Bassell GJ, et al. S6K1 phosphorylates and regulates fragile X mental retardation protein (FMRP) with the neuronal protein synthesis-dependent mammalian target of rapamycin (mTOR) signaling cascade. J Biol Chem. 2008;283:18478–82.PubMed CentralPubMedGoogle Scholar
- Nelson D. 2010. New models of FXS in the mouse to assess developmental effects of FMR1 Mutation. 12th International Fragile X Conference, 23 July 2010, Detroit, Michigan.Google Scholar
- NIH (2009). Outcome measures for clinical trials in children with fragile X syndrome—Meeting Summary November 5 and 6, 2009. Bethesda Marriott, Pooks Hill, Bethesda, MD. http://rarediseases.info.nih.gov/ScientificConferences.aspx?PageID=5&ID=989.
- Nimchinsky EA, Oberlander AM, Svoboda K. Abnormal development of dendritic spines in FMR1 knock-out mice. J Neurosci. 2001;21:5139–46.PubMedGoogle Scholar
- Osterweil EK, Krueger DD, Reinhold K, Bear MF. Hypersensitivity to mGluR5 and ERK1/2 leads to excessive protein synthesis in the hippocampus of a mouse model of fragile X syndrome. J Neurosci. 2010;30:15616–27.PubMed CentralPubMedGoogle Scholar
- Owen R, Gordon-Weeks PR. Inhibition of glycogen synthase kinase 3beta in sensory neurons in culture alters filopodia dynamics and microtubule distribution in growth cones. Mol Cell Neurosci. 2003;23:626–37.PubMedGoogle Scholar
- Pacey LK, Heximer SP, Hampson DR. Increased GABA(B) receptor-mediated signaling reduces the susceptibility of fragile X knockout mice to audiogenic seizures. Mol Pharmacol. 2009;76:18–24.PubMedGoogle Scholar
- Paribello C, Tao L, Folino A, Berry-Kravis E, Tranfaglia M, Ethell I, et al. Open-label add-on treatment trial of minocycline in fragile X syndrome. BMC Neurol. 2010;10:91.PubMed CentralPubMedGoogle Scholar
- Park S, Park JM, Kim S, Kim JA, Shepard JD, Smith-Hicks CL, et al. Elongation factor 2 and fragile X mental retardation protein control the dynamic translation of Arc/Arg3.1 essential for mGluR-LTD. Neuron. 2008;59:70–83.PubMed CentralPubMedGoogle Scholar
- Paylor R. 2008. Pharmacological modification of behavioral responses in the fragile X mouse model. 11th International Fragile X Conference, 25 July 2008, St. Louis, Missouri.Google Scholar
- Paylor R, Yuva-Paylor LA, Nelson DL, Spenser CM. Reversal of sensorimotor gating abnormalities in Fmr1 knockout mice carrying a human Fmr1 transgene. Behav Neurosci. 2008;122:1371–7.PubMedGoogle Scholar
- Pinto D, Pagnamenta AT, Klei L, Anney R, Merico D, Regan R, et al. Functional impact of global rare copy number variation in autism spectrum disorders. Nature. 2010;466:368–72.PubMed CentralPubMedGoogle Scholar
- Qin M, Kang J, Burlin TV, Jiang C, Smith CB. Postadolescent changes in regional cerebral protein synthesis: an in vivo study in the FMR1 null mouse. J Neurosci. 2005;25:5087–95.PubMedGoogle Scholar
- Sharma A, Hoeffer CA, Takayasu Y, Miyawaki T, McBride S, Klann E, et al. Dysregulation of mTOR signaling in fragile X syndrome. J Neurosci. 2010;30:694–702.PubMed CentralPubMedGoogle Scholar
- Shumbos JG, Brown M, Kronengold J, Polley DB, Kaczmerak LK. Fragile x mental retardation protein is required for rapid experience-dependent regulation of the potassium channel Kv3.1b. J Neurosci. 2010;30:10263–71.Google Scholar
- Sohn JW, Lee D, Cho H, Lim W, Hin HS, Lee SH, et al. Receptor-specific inhibition of GABAB-activated K+ currents by muscarinic and metabotropic glutamate receptors in immature rat hippocampus. J Physiol. 2007;580:411–22.PubMed CentralPubMedGoogle Scholar
- Spencer CM, Graham DF, Yuva-Paylor LA, Nelson DL, Paylor R. Social behavior in Fmr1 knockout mice carrying a human FMR1 transgene. Behav Neurosci. 2008;122:710–5.PubMedGoogle Scholar
- Steckler T, Oliveira AF, Van Dyck C, Van Craenendonck H, Mateus AM, Langloi X, et al. Metabotropic glutamate receptor 1 blockade impairs acquisition and retention in a spatial water maze task. Behav Brain Res. 2005;164:52.PubMedGoogle Scholar
- Subrathan A, Hoeffer CA, Wong H, Klann E, Chattarji S. Characterization and reversal of synaptic defects in the amygdale in a mouse model of fragile X syndrome. Proc Natl Acad Sci USA. 2010;107:11591–6.Google Scholar
- Sullivan AK, Marcus M, Epstein MP, Allen EG, Anido AE, Paquin JJ, et al. Association of FMR1 repeat size with ovarian dysfunction. Hum Reprod. 2005;20:402–12.PubMedGoogle Scholar
- Sullivan K, Hooper S, Hatton D. Behavioral equivalents of anxiety in children with fragile X syndrome: parent and teacher report. J Intellect Disabil Res. 2007;51:54–65.PubMedGoogle Scholar
- Tassone F, Hagerman RJ, Ikle DN, Dyer PN, Lampe N, Willemson R, et al. FMRP expression as a potential prognostic indicator in fragile X syndrome. Am J Med Genet. 1999;84:250–61.PubMedGoogle Scholar
- Tassone F, Hagerman RJ, Taylor AK, Mills JB, Harris SW, Gane LW, et al. Clinical involvement and protein expression in individuals with the FMR1 premutation. Am J Med Genet. 2000;91:144–52.PubMedGoogle Scholar
- Turner G, Webb T, Wake S, Robinson H. Prevalence of fragile X syndrome. Am J Med Genet. 1996;64:196–7.PubMedGoogle Scholar
- Utari A, Chonchaiya W, Rivera SM, Schneider A, Hagerman RJ, Faradz SM, et al. Side effects of minocycline treatment in patients with fragile X syndrome and exploration of outcome measures. Am J Intellect Dev Disabil. 2010;115(5):433–43.PubMed CentralPubMedGoogle Scholar
- Venkitaramani DV, Aldridge GM, Kao D-I, Morris DR, Weiler I, Greenough WT. 2010. Rescue of fragile X syndrome metabolic and behavioral phenotypes by lithium. Program No. 757.12. 2010 Neuroscience Meeting Planner. San Diego, CA: Society for Neuroscience (online).Google Scholar
- Ventura R, Pascucci T, Catania MV, Musumeci SA, Puglisi-Allegra S. Object recognition impairment in Fmr1 knockout mice is reversed by amphetamine: involvement of dopamine in the medial prefrontal cortex. Behav Pharmacol. 2004;15:433–42.PubMedGoogle Scholar
- Verkerk AJ, Pieretti M, Sutcliffe JS, Fu YH, Kuhl DP, Pizzuti A, et al. Identification of a gene (FMR-1) containing a CGG repeat coincident with a breakpoint cluster region exhibiting length variation in fragile X syndrome. Cell. 1991;65:905–14.PubMedGoogle Scholar
- Volk LJ, Pfeiffer BE, Gibson JR, Huber KM. Multiple Gq-coupled receptors converge on a common protein synthesis-dependent long-term depression that is affected in fragile X syndrome mental retardation. J Neurosci. 2007;27:11624–34.PubMedGoogle Scholar
- Wang H, Wu LJ, Zhang F, Zhou M. Roles of calcium-stimulated adenylyl cyclase and calmodulin-depebdent protein kinase IV in the regulation of FMRP by group I metabotropic glutamate receptors. JNeurosci. 2008a;28:4385–97.Google Scholar
- Wang H, Wu LJ, Kim SS, Lee FJ, Gong B, Toyoda H. FMRP acts as a key messenger for dopamine modulation in the forebrain. Neuron. 2008b;59:634–47.PubMedGoogle Scholar
- Wang LW, Berry-Kravis E, Hagerman RJ. Fragile X: leading the way for targeted treatments in autism. Neurotherapeutics. 2010a;7:264–74.PubMed CentralPubMedGoogle Scholar
- Wang H, Kim SS, Zhuo M. Roles of fragile X mental retardation protein in dopaminergic stimulation-induced synapse-associated protein synthesis and subsequent α-amino-3-hydroxyl-5-methyl-4-isoxazole-4-propionate (AMPA) receptor internalization. J Biol Chem. 2010b;285:21888–901.PubMed CentralPubMedGoogle Scholar
- Wang L, Hagerman R, Rathmell B, Wang P, Berry-Kravis E. 2010c. Arbaclofen treatment is associated with global behavioral improvement in fragile X syndrome (FXS): results of a randomized, controlled phase 2 trial. 2010 Annual Meeting of the Society for Developmental and Behavioral Pediatrics, Sept 12, 2010, Boston, Massachusetts.Google Scholar
- Waung MW, Huber KM. Protein translation in synaptic plasticity: mGluR-LTD, fragile X. Curr Opin Neurobiol. 2009;19:319–26.PubMed CentralPubMedGoogle Scholar
- Weiler IJ, Irwin SA, Klintsova AY, Spencer CM, Brazelton AD, Miyashiro K, et al. Fragile X mental retardation protein is translated near synapses in response to neurotransmitter activation. Proc Natl Acad Sci USA. 1997;94:5395–400.PubMed CentralPubMedGoogle Scholar
- Weiler IJ, Spangler CC, Klintsova AY, Grossman AW, Kim SH, Bertaina-Anglade V, et al. Fragile X mental retardation protein is necessary for neurotransmitter-activated protein translation at synapses. Proc Natl Acad Sci USA. 2004;101:17329–30.Google Scholar
- Weng N, Weiler IJ, Sumis A, Berry-Kravis E, Greenough WT. Early-phase ERK activation as a biomarker for metabolic status in fragile X syndrome. Am J Med Genet. 2008;147B:1253–7.PubMedGoogle Scholar
- Westmark CJ, Malter JS. FMRP mediates mgluR5-dependent translation of amyloid precursor protein. PLoS Biol. 2007;5:e52.PubMed CentralPubMedGoogle Scholar
- Westmark CJ, Westmark PR, Hays SA, Hervey CM, Salamat MS, Stein KM, et al. 2010. Reversal of fragile X phenotypes by manipulation of APP/Ab levels. Society for Neuroscience 17 November 2010, San Diego, California.Google Scholar
- Yan QJ, Asafo-Adjei PK, Arnold HM, Brown RE, Bauchwitz RP. A phenotypic and molecular characterization of the fmr1-tm1Cgr fragile X mouse. Genes Brain Behav. 2004;3:337–59.PubMedGoogle Scholar
- Yan QJ, Rammal M, Tranfaglia M, Bauchwitz RP. Suppression of two major fragile X syndrome mouse model phenotypes by the mGluR5 antagonist MPEP. Neuropharmacology. 2005;49:1053–66.PubMedGoogle Scholar
- Yuskaitis CJ, Mines MA, King MK, Sweatt JD, Miller CA, Jope RS. Lithium ameliorates altered glycogen synthase kinase-3 and behavior in a mouse model of fragile X syndrome. Biochem Pharmacol. 2010;79:632–46.PubMed CentralPubMedGoogle Scholar
- Zalfa F, Giorgi M, Primerano B, Moro A, Di Penta A, Reis S, et al. The fragile X syndrome protein FMRP associates with BC1 RNA and regulates the translation of specific mRNAs at synapses. Cell. 2003;112:317–27.PubMedGoogle Scholar
- Zhang L, Alger BE. Enhanced endocannabinoid signaling elevates neuronal excitability in fragile X syndrome. J Neurosci. 2010;30:5724–9.PubMed CentralPubMedGoogle Scholar
- Zhang YQ, Bailey AM, Matthies HJ, Renden RB, Smith MA, Speese SD, et al. Drosophila fragile X-related gene regulates the MAP1B homolog Futsch to control synaptic structure and function. Cell. 2001;107:591–603.PubMedGoogle Scholar
- Zhang Y, Venkitaramani DV, Gladding CM, Zhang Y, Kurup P, Molnar E, et al. The tyrosine phosphatase STEP mediates AMPA receptor endocytosis after metabotropic glutamate receptor stimulation. J Neurosci. 2008;28:10561–6.PubMed CentralPubMedGoogle Scholar