Validation of the Observer-Reported Communication Ability (ORCA) measure for individuals with Rett syndrome.

PURPOSE: The study goal was to validate the Observer-Reported Communication Ability (ORCA) measure for use with females with Rett Syndrome (RTT). METHODS: Qualitative interviews, including concept elicitation and cognitive interviewing methods, were conducted with 19 caregivers of individuals with RTT ages 2 and older. A quantitative study was then conducted in 279 caregivers to evaluate construct validity and reliability. RESULTS: After minor modifications were made, the modified ORCA measure was well understood and captured key communication concepts. Quantitative data showed evidence for reliable scores (α = 0.90, test-retest intraclass correlation = 0.88), minimal floor and no ceiling effects, and strong correlation with the Communication and Symbolic Behaviors Scale (r = 0.73). CONCLUSIONS: This study provided initial support that the modified ORCA measure is an acceptable caregiver-reported measure of communication ability for females with RTT. Future work should include evaluation of longitudinal validity of the measure and its associations with clinician- and performance-based measures in diverse samples.

Saliva MicroRNA Differentiates Children With Autism From Peers With Typical and Atypical Development.

OBJECTIVE: Clinical diagnosis of autism spectrum disorder (ASD) relies on time-consuming subjective assessments. The primary purpose of this study was to investigate the utility of salivary microRNAs for differentiating children with ASD from peers with typical development (TD) and non-autism developmental delay (DD). The secondary purpose was to explore microRNA patterns among ASD phenotypes. METHOD: This multicenter, prospective, case-control study enrolled 443 children (2-6 years old). ASD diagnoses were based on DSM-5 criteria. Children with ASD or DD were assessed with the Autism Diagnostic Observation Schedule II and Vineland Adaptive Behavior Scales II. MicroRNAs were measured with high-throughput sequencing. Differential expression of microRNAs was compared among the ASD (n = 187), TD (n = 125), and DD (n = 69) groups in the training set (n = 381). Multivariate logistic regression defined a panel of microRNAs that differentiated children with ASD and those without ASD. The algorithm was tested in a prospectively collected naïve set of 62 samples (ASD, n = 37; TD, n = 8; DD, n = 17). Relations between microRNA levels and ASD phenotypes were explored. RESULT: Fourteen microRNAs displayed differential expression (false discovery rate < 0.05) among ASD, TD, and DD groups. A panel of 4 microRNAs (controlling for medical/demographic covariates) best differentiated children with ASD from children without ASD in training (area under the curve = 0.725) and validation (area under the curve = 0.694) sets. Eight microRNAs were associated (R > 0.25, false discovery rate < 0.05) with social affect, and 10 microRNAs were associated with restricted/repetitive behavior. CONCLUSION: Salivary microRNAs are "altered" in children with ASD and associated with levels of ASD behaviors. Salivary microRNA collection is noninvasive, identifying ASD-status with moderate accuracy. A multi-"omic" approach using additional RNA families could improve accuracy, leading to clinical application. CLINICAL TRIAL REGISTRATION INFORMATION: A Salivary miRNA Diagnostic Test for Autism; https://clinicaltrials.gov/; NCT02832557.

The levo enantiomer of amphetamine increases memory consolidation and gene expression in the hippocampus without producing locomotor stimulation.

Dextro-amphetamine enhances memory and other cognitive functions in animals and humans. The use of d-amphetamine as a memory enhancer, however, is limited by a robust stimulatory side-effect profile caused by release of dopamine. The levo enantiomer of amphetamine has been shown to be considerably less effective as a dopamine releaser and less potent in producing the stimulatory effects characteristic of d-amphetamine. In order to determine whether l-amphetamine and the structurally related compound, l-methamphetamine, retain cognitive-enhancing effects despite their lack of stimulatory activity, we administered the compounds to rats prior to activity monitoring experiments, and in different animals, immediately after training on inhibitory avoidance and object recognition tasks. Results demonstrated that l-amphetamine and l-methamphetamine did not increase locomotion and stereotypies beyond control levels, but did produce significant memory enhancement. In addition, l-amphetamine and l-methamphetamine alleviated scopolamine-induced amnesia in the inhibitory avoidance task. In all cases, these compounds produced an effect comparable to that of d-amphetamine, but required only one quarter of the d-amphetamine dose to produce the same effect size. We also found that l-amphetamine modulates learning-induced changes in hippocampal Arc/Arg3.1 protein synthesis that correlate with memory consolidation. These results suggest that l-amphetamine and l-methamphetamine are potent memory enhancers in rats and may ultimately be useful for treating memory disorders in humans.

Validation of a Salivary RNA Test for Childhood Autism Spectrum Disorder.

Background: The diagnosis of autism spectrum disorder (ASD) relies on behavioral assessment. Efforts to define biomarkers of ASD have not resulted in an objective, reliable test. Studies of RNA levels in ASD have demonstrated potential utility, but have been limited by a focus on single RNA types, small sample sizes, and lack of developmental delay controls. We hypothesized that a saliva-based poly-"omic" RNA panel could objectively distinguish children with ASD from their neurotypical peers and children with non-ASD developmental delay. Methods: This multi-center cross-sectional study included 456 children, ages 19-83 months. Children were either neurotypical (n = 134) or had a diagnosis of ASD (n = 238), or non-ASD developmental delay (n = 84). Comprehensive human and microbial RNA abundance was measured in the saliva of all participants using unbiased next generation sequencing. Prior to analysis, the sample was randomly divided into a training set (82% of subjects) and an independent validation test set (18% of subjects). The training set was used to develop an RNA-based algorithm that distinguished ASD and non-ASD children. The validation set was not used in model development (feature selection or training) but served only to validate empirical accuracy. Results: In the training set (n = 372; mean age 51 months; 75% male; 51% ASD), a set of 32 RNA features (controlled for demographic and medical characteristics), identified ASD status with a cross-validated area under the curve (AUC) of 0.87 (95% CI: 0.86-0.88). In the completely separate validation test set (n = 84; mean age 50 months; 85% male; 60% ASD), the algorithm maintained an AUC of 0.88 (82% sensitivity and 88% specificity). Notably, the RNA features were implicated in physiologic processes related to ASD (axon guidance, neurotrophic signaling). Conclusion: Salivary poly-omic RNA measurement represents a novel, non-invasive approach that can accurately identify children with ASD. This technology could improve the specificity of referrals for ASD evaluation or provide objective support for ASD diagnoses.

Drug development for neurodevelopmental disorders: lessons learned from fragile X syndrome.

Neurodevelopmental disorders such as fragile X syndrome (FXS) result in lifelong cognitive and behavioural deficits and represent a major public health burden. FXS is the most frequent monogenic form of intellectual disability and autism, and the underlying pathophysiology linked to its causal gene, FMR1, has been the focus of intense research. Key alterations in synaptic function thought to underlie this neurodevelopmental disorder have been characterized and rescued in animal models of FXS using genetic and pharmacological approaches. These robust preclinical findings have led to the implementation of the most comprehensive drug development programme undertaken thus far for a genetically defined neurodevelopmental disorder, including phase IIb trials of metabotropic glutamate receptor 5 (mGluR5) antagonists and a phase III trial of a GABAB receptor agonist. However, none of the trials has been able to unambiguously demonstrate efficacy, and they have also highlighted the extent of the knowledge gaps in drug development for FXS and other neurodevelopmental disorders. In this Review, we examine potential issues in the previous studies and future directions for preclinical and clinical trials. FXS is at the forefront of efforts to develop drugs for neurodevelopmental disorders, and lessons learned in the process will also be important for such disorders.

Arbaclofen in Children and Adolescents with Autism Spectrum Disorder: A Randomized, Controlled, Phase 2 Trial.

Several lines of emerging data point to an imbalance between neuronal excitation and inhibition in at least a subgroup of individuals with autism spectrum disorder (ASD), including in those with fragile X syndrome (FXS), one of the most common genetic syndromes within ASD. In animal models of FXS and of ASD, GABA-B agonists have improved both brain and behavioral phenotypes, including social behavior. A phase 2 randomized, placebo-controlled, crossover trial found that the GABA-B agonist arbaclofen improved social avoidance symptoms in FXS. A pilot open-label trial of arbaclofen suggested similar benefits in ASD. We therefore evaluated arbaclofen in a randomized, placebo-controlled, phase 2 study of 150 participants, aged 5-21 years, with ASD. No difference from placebo was detected on the primary outcome measure, the parent-rated Aberrant Behavior Checklist Social Withdrawal/Lethargy subscale. However, a specified secondary analysis found improvement on the clinician-rated Clinical Global Impression of Severity. An exploratory post hoc analysis of participants with a consistent rater across the trial revealed greater improvement in the Vineland Adaptive Behavior Scales II socialization domain in participants receiving arbaclofen. Affect lability (11%) and sedation (9%) were the most common adverse events. In this exploratory study, secondary analyses suggest that arbaclofen may have the potential to improve symptoms in some children with ASD, but further study will be needed to replicate and extend these initial findings.

Arbaclofen in fragile X syndrome: results of phase 3 trials.

BACKGROUND: Arbaclofen improved multiple abnormal phenotypes in animal models of fragile X syndrome (FXS) and showed promising results in a phase 2 clinical study. The objective of the study is to determine safety and efficacy of arbaclofen for social avoidance in FXS. METHODS: Two phase 3 placebo-controlled trials were conducted, a flexible dose trial in subjects age 12-50 (209FX301, adolescent/adult study) and a fixed dose trial in subjects age 5-11 (209FX302, child study). The primary endpoint for both trials was the Social Avoidance subscale of the Aberrant Behavior Checklist-Community Edition, FXS-specific (ABC-CFX). Secondary outcomes included other ABC-CFX subscale scores, Clinical Global Impression-Improvement (CGI-I), Clinical Global Impression-Severity (CGI-S), and Vineland Adaptive Behavior Scales, Second Edition (Vineland-II) Socialization domain score. RESULTS: A total 119 of 125 randomized subjects completed the adolescent/adult study (n = 57 arbaclofen, 62 placebo) and 159/172 completed the child study (arbaclofen 5 BID n = 38; 10 BID n = 39; 10 TID n = 38; placebo n = 44). There were no serious adverse events (AEs); the most common AEs included somatic (headache, vomiting, nausea), neurobehavioral (irritability/agitation, anxiety, hyperactivity), decreased appetite, and infectious conditions, many of which were also common on placebo. In the combined studies, there were 13 discontinuations (n = 12 arbaclofen, 1 placebo) due to AEs (all neurobehavioral). The adolescent/adult study did not show benefit for arbaclofen over placebo for any measure. In the child study, the highest dose group showed benefit over placebo on the ABC-CFX Irritability subscale (p = 0.03) and Parenting Stress Index (PSI, p = 0.03) and trends toward benefit on the ABC-CFX Social Avoidance and Hyperactivity subscales (both p < 0.1) and CGI-I (p = 0.119). Effect size in the highest dose group was similar to effect sizes for FDA-approved serotonin reuptake inhibitors (SSRIs). CONCLUSIONS: Arbaclofen did not meet the primary outcome of improved social avoidance in FXS in either study. Data from secondary measures in the child study suggests younger patients may derive benefit, but additional studies with a larger cohort on higher doses would be required to confirm this finding. The reported studies illustrate the challenges but represent a significant step forward in translating targeted treatments from preclinical models to clinical trials in humans with FXS.

Alvimopan: an oral, peripherally acting, mu-opioid receptor antagonist for the treatment of opioid-induced bowel dysfunction--a 21-day treatment-randomized clinical trial.

Alvimopan has been shown to reverse the inhibitory effect of opioids on gastrointestinal transit without affecting analgesia. We evaluated oral alvimopan, 0.5 or 1 mg, versus placebo, once daily for 21 days, in 168 patients with opioid-induced bowel dysfunction (OBD) who were receiving chronic opioid therapy (minimum, 1 month) for nonmalignant pain (n = 148) or opioid dependence (n = 20). The primary outcome was the proportion of patients having at least one bowel movement (BM) within 8 hours of study drug on each day during the 21-day treatment period. Averaged over the 21-day treatment period, 54%, 43%, and 29% of patients had a BM within 8 hours after alvimopan 1 mg, 0.5 mg, or placebo, respectively (P < .001). Secondary outcomes of median times to first BM were 3, 7, and 21 hours after initial doses of 1 mg, 0.5 mg, and placebo, respectively (P < .001; 1 mg vs placebo). Weekly BMs and overall patient satisfaction were increased after the 1-mg dose (P < .001 at weeks 1 and 2 vs placebo, and P = .046, respectively). Treatment-emergent adverse events were primarily bowel-related, occurred during the first week of treatment, and were of mild to moderate severity. Alvimopan was generally well tolerated and did not antagonize opioid analgesia. Patients treated with chronic opioid therapy often experience opioid-induced bowel dysfunction as a result of undesirable effects on peripheral opioid receptors located in the gastrointestinal tract. Alvimopan, a novel peripheral opioid mu-receptor antagonist, has demonstrated significant efficacy for the management of opioid-induced bowel dysfunction without compromise of centrally mediated opioid-induced analgesia.

STX209 (arbaclofen) for autism spectrum disorders: an 8-week open-label study.

STX209 (arbaclofen), a selective GABA-B agonist, is hypothesized to modulate the balance of excitatory to inhibitory neurotransmission, and has shown preliminary evidence of benefit in fragile X syndrome. We evaluated its safety, tolerability, and efficacy in non-syndromic autism spectrum disorders, in an 8-week open-label trial enrolling 32 children and adolescents with either Autistic Disorder or Pervasive Developmental Disorder-Not Otherwise Specified, and a score ≥17 on the Aberrant Behavior Checklist (ABC)-Irritability subscale. STX209 was generally well-tolerated. The most common adverse events were agitation and irritability, which typically resolved without dose changes, and were often felt to represent spontaneous variation in underlying symptoms. Improvements were observed on several outcome measures in this exploratory trial, including the ABC-Irritability (the primary endpoint) and the Lethargy/Social Withdrawal subscales, the Social Responsiveness Scale, the CY-BOCS-PDD, and clinical global impression scales. Placebo-controlled study of STX209 is warranted.

Effects of STX209 (arbaclofen) on neurobehavioral function in children and adults with fragile X syndrome: a randomized, controlled, phase 2 trial.

Research on animal models of fragile X syndrome suggests that STX209, a γ-aminobutyric acid type B (GABA(B)) agonist, might improve neurobehavioral function in affected patients. We evaluated whether STX209 improves behavioral symptoms of fragile X syndrome in a randomized, double-blind, placebo-controlled crossover study in 63 subjects (55 male), ages 6 to 39 years, with a full mutation in the FMR1 gene (>200 CGG triplet repeats). We found no difference from placebo on the primary endpoint, the Aberrant Behavior Checklist-Irritability (ABC-I) subscale. In the other analyses specified in the protocol, improvement was seen on the visual analog scale ratings of parent-nominated problem behaviors, with positive trends on multiple global measures. Post hoc analysis with the ABC-Social Avoidance scale, a newly validated scale for the assessment of fragile X syndrome, showed a significant beneficial treatment effect in the full study population. A post hoc subgroup of 27 subjects with more severe social impairment showed improvements on the Vineland II-Socialization raw score, on the ABC-Social Avoidance scale, and on all global measures. STX209 was well tolerated, with 8% incidences of sedation and of headache as the most frequent side effects. In this exploratory study, STX209 did not show a benefit on irritability in fragile X syndrome. Nonetheless, our results suggest that GABA(B) agonists have potential to improve social function and behavior in patients with fragile X syndrome.

Reversal of disease-related pathologies in the fragile X mouse model by selective activation of GABAB receptors with arbaclofen.

Fragile X syndrome (FXS), the most common inherited cause of intellectual disability and autism, results from the transcriptional silencing of FMR1 and loss of the mRNA translational repressor protein fragile X mental retardation protein (FMRP). Patients with FXS exhibit changes in neuronal dendritic spine morphology, a pathology associated with altered synaptic function. Studies in the mouse model of fragile X have shown that loss of FMRP causes excessive synaptic protein synthesis, which results in synaptic dysfunction and altered spine morphology. We tested whether the pharmacologic activation of the γ-aminobutyric acid type B (GABA(B)) receptor could correct or reverse these phenotypes in Fmr1-knockout mice. Basal protein synthesis, which is elevated in the hippocampus of Fmr1-knockout mice, was corrected by the in vitro application of the selective GABA(B) receptor agonist STX209 (arbaclofen, R-baclofen). STX209 also reduced to wild-type values the elevated AMPA receptor internalization in Fmr1-knockout cultured neurons, a known functional consequence of increased protein synthesis. Acute administration of STX209 in vivo, at doses that modify behavior, decreased mRNA translation in the cortex of Fmr1-knockout mice. Finally, the chronic administration of STX209 in juvenile mice corrected the increased spine density in Fmr1-knockout mice without affecting spine density in wild-type mice. Thus, activation of the GABA(B) receptor with STX209 corrected synaptic abnormalities considered central to fragile X pathophysiology, a finding that suggests that STX209 may be a potentially effective therapy to treat the core symptoms of FXS.

Modulation of behavioral phenotypes by a muscarinic M1 antagonist in a mouse model of fragile X syndrome.

RATIONALE: Muscarinic acetylcholine receptors (mAChR) are G protein-coupled receptors, widely expressed in the CNS. Electrophysiological and molecular studies have provided evidence for overactive M1 receptor signaling in the fragile X knockout (Fmr1 KO) mouse model, suggesting the involvement of the M1 receptors in fragile X syndrome. Overactive signaling through the M1 receptor has been hypothesized to contribute to the phenotypes seen in fragile X mice. OBJECTIVE: We investigated the modulation of behavioral responses in the Fmr1 KO animals by reducing the activity through the muscarinic M1 receptor using the pharmacological agent dicyclomine, an M1 antagonist. METHODS: The behavioral assays used to investigate the pharmacological effects include marble burying (perseverative behavior), open-field exploration (activity), passive avoidance (learning and memory), prepulse inhibition (sensorimotor gating), and audiogenic seizures. RESULTS: Data from the marble-burying assay suggests that treatment with dicyclomine results in a decrease in the number of marbles buried in the wild-type and in the KO animals. To examine the possibility of drug-induced sedation, overall activity was measured in an open-field chamber. Dicyclomine only increases activity at a dose of 20 mg/kg in the wild-type mice but did not affect exploration in the KO animals. Lastly, we observed that dicyclomine causes a significant decrease in the percentage of audiogenic seizures in the Fmr1 KO animals. CONCLUSION: Our findings suggest that pharmacologically reducing the activity through the mAChR M1 alters select behavioral responses in the Fmr1 KO mice.

Mechanism-based approaches to treating fragile X.

Fragile X is the leading inherited cause of mental retardation and autism. Recent advances in our mechanistic understanding of the disease have led to the identification of the metabotropic glutamate receptor (mGluR) as a therapeutic target for the disease. These studies have revealed that core defects in multiple animal models can be corrected by down regulation of mGluR5 signaling. Although it remains to be seen if mGluR5 antagonists or related approaches will succeed in humans with fragile X, the progress in fragile X stands as a strong testament to the power of applying knowledge of basic neurobiology to understand pathophysiology in a genetically validated model of human psychiatric disease. These breakthroughs and several of the resulting drug development efforts are reviewed.