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Human cell line models, including the neuronal precursor line LUHMES, are important for investigating developmental transcriptional dynamics within imprinted regions, particularly the 15q11-q13 Angelman (AS) and Prader-Willi (PWS) syndrome locus. AS results from loss of maternal UBE3A in neurons, where the paternal allele is silenced by a convergent antisense transcript UBE3A-ATS, a lncRNA that terminates at PWAR1 in non-neurons. qRT-PCR analysis confirmed the exclusive and progressive increase in UBE3A-ATS in differentiating LUHMES neurons, validating their use for studying UBE3A silencing. Genome-wide transcriptome analyses revealed changes to 11 834 genes during neuronal differentiation, including the upregulation of most genes within the 15q11-q13 locus. To identify dynamic changes in chromatin loops linked to transcriptional activity, we performed a HiChIP validated by 4C, which identified two neuron-specific CTCF loops between MAGEL2-SNRPN and PWAR1-UBE3A. To determine if allele-specific differentially methylated regions (DMR) may be associated with CTCF loop anchors, whole genome long-read nanopore sequencing was performed. We identified a paternally hypomethylated DMR near the SNRPN upstream loop anchor exclusive to neurons and a paternally hypermethylated DMR near the PWAR1 CTCF anchor exclusive to undifferentiated cells, consistent with increases in neuronal transcription. Additionally, DMRs near CTCF loop anchors were observed in both cell types, indicative of allele-specific differences in chromatin loops regulating imprinted transcription. These results provide an integrated view of the 15q11-q13 epigenetic landscape during LUHMES neuronal differentiation, underscoring the complex interplay of transcription, chromatin looping, and DNA methylation. They also provide insights for future therapeutic approaches for AS and PWS.

BackgroundOxytocin and vasopressin systems are altered in Prader Willi syndrome (PWS). However, investigations into endogenous oxytocin and vasopressin levels as well as clinical trials evaluating the effect of exogenous oxytocin on PWS symptoms have had mixed results. It is also unknown whether endogenous oxytocin and vasopressin levels are associated with certain PWS behaviours.MethodWe compared plasma oxytocin and vasopressin and saliva oxytocin levels in 30 adolescents and adults with PWS to 30 typically developing age-matched controls. We also compared neuropeptide levels between gender and genetic subtypes within the PWS cohort and examined the relationship between neuropeptide levels and PWS behaviours.ResultsWhile we did not measure a group difference in plasma or saliva oxytocin levels, plasma vasopressin was significantly lower in individuals with PWS compared to controls. Within the PWS cohort, saliva oxytocin levels were higher in females compared to males and individuals with the mUPD compared to the deletion genetic subtype. We also found the neuropeptides correlated with different PWS behaviours for males and females and for genetic subtypes. For the deletion group, higher plasma and saliva oxytocin levels were related to fewer behaviour problems. For the mUPD group, higher plasma vasopressin levels were related to more behaviour problems.ConclusionThese findings support existing evidence of a vasopressin system defect in PWS and for the first time identify potential differences in the oxytocin and vasopressin systems across PWS genetic subtypes.

Prader-Willi syndrome (PWS) is a complex, genetic, neurodevelopmental disorder. PWS has three molecular genetic classes. The most common defect is due to a paternal 15q11-q13 deletion observed in about 60% of individuals. This is followed by maternal disomy 15 (both 15 s from the mother), found in approximately 35% of cases. the remaining individuals have a defect of the imprinting center that controls the activity of imprinted genes on chromosome 15. Mild cognitive impairment and behavior problems in PWS include self-injury, anxiety, compulsions, and outbursts in childhood, impacted by genetic subtypes. Food seeking and hyperphagia can lead to morbid obesity and contribute to diabetes and cardiovascular or orthopedic problems. The control of hyperphagia and improving food-related behaviors are the most important unmet needs in PWS and could be addressed with the development of a new therapeutic agent, as currently no approved therapeutics exist for PWS treatment. The status of clinical trials with existing results for the management of obesity and hyperphagia in PWS will be discussed in this review, including treatments such as beloranib, setmelanotide, a diazoxide choline controlled-release tablet (DCCR), an unacylated ghrelin analogue, oxytocin and related compounds, glucagon-like peptide 1 receptor agonists, surgical intervention, and transcranial direct-current stimulation.

BackgroundPrader-Willi syndrome (PWS) is an imprinting disorder caused by the absence of paternal expressed genes in the Prader-Willi critical region (PWCR) on chromosome 15q11.2-q13. Three molecular mechanisms have been known to cause PWS, including a deletion in the PWCR, uniparental disomy 15 and imprinting defects.ResultsWe report the first case of PWS associated with a single-nucleotide SNRPN variant in a 10-year-old girl presenting with clinical features consistent with PWS, including infantile hypotonia and feeding difficulty, developmental delay with cognitive impairment, excessive eating with central obesity, sleep disturbances, skin picking and related behaviour issues. Whole-exome sequencing revealed a de novo mosaic nonsense variant of the SNRPN gene (c.73C>T, p.R25X) in 10% of DNA isolated from buccal cells and 19% of DNA from patient-derived lymphoblast cells. DNA methylation study did not detect an abnormal methylation pattern in the SNRPN locus. Parental origin studies showed a paternal source of an intronic single-nucleotide polymorphism within the locus in proximity to the SNRPN variant.ConclusionsThis is the first report that provides evidence of a de novo point mutation of paternal origin in SNRPN as a new disease-causing mechanism for PWS. This finding suggests that gene sequencing should be considered as part of the diagnostic workup in patients with clinical suspicion of PWS.

BackgroundAdequate sleep is important for proper neurodevelopment and positive health outcomes. Sleep disturbances are more prevalent in children with genetically determined neurodevelopmental syndromes compared with typically developing counterparts. We characterize sleep behavior in Rett (RTT), Angelman (AS), and Prader-Willi (PWS) syndromes to identify effective approaches for treating sleep problems in these populations. We compared sleep-related symptoms across individuals with these different syndromes with each other, and with typically developing controls.MethodsChildren were recruited from the Rare Diseases Clinical Research Network consortium registries; unaffected siblings were enrolled as related controls. For each participant, a parent completed multiple sleep questionnaires including Pediatric Sleep Questionnaire (Sleep-Disordered Breathing), Children's Sleep Habits Questionnaire (CSHQ), and Pediatric Daytime Sleepiness Scale.ResultsSleep data were analyzed from 714 participants, aged two to 18 years. Young children with AS had more reported sleep problems than children with RTT or PWS. Older children with RTT had more reported daytime sleepiness than those with AS or PWS. Finally, all individuals with RTT had more evidence of sleep-disordered breathing when compared with individuals with PWS. Notably, typically developing siblings were also reported to have sleep problems, except for sleep-related breathing disturbances, which were associated with each of the genetic syndromes.ConclusionsIndividuals with RTT, AS, and PWS frequently experience sleep problems, including sleep-disordered breathing. Screening for sleep problems in individuals with these and other neurogenetic disorders should be included in clinical assessment and managements. These data may also be useful in developing treatment strategies and in clinical trials.

Prader-Willi syndrome (PWS) is a complex genetic disorder with three molecular classes but clinical ascertainment is based on distinctive features. The prevalence of dysmorphic features was studied in 355 PWS participants (61% deletion, 36% maternal disomy [UPD], and 3% imprinting defects) from the National Institute of Health PWS Rare Diseases Clinical Research Network. The effect of growth hormone (GH) treatment on growth and dysmorphic features was compared. Among participants, upslanting palpebral fissures were seen in 23%; strabismus in 42%; abnormal dentition in 32%; small hands in 63% and small feet in 70%; hypopigmentation in 30%; striae in 32% and skin picking in 26%. Compared to those with UPD, participants with deletions were found to be heavier (p = 0.002), had smaller head circumference (HC) (p = 0.009), higher incidence of a flat occiput (p = 0.005); low-anterior hairline (p = 0.04); abnormal dentition (p = 0.009); abdominal striae (p = 0.045), nail abnormalities (p = 0.050), and fair-haired (p

Prader-Willi syndrome (PWS) affects 1/15,000-1/30,000 live births and is characterized by lack of expression of paternally inherited genes on 15q11.2-15q13 caused by paternal deletions, maternal uniparental disomy (UPD), or imprinting defects. Affected individuals have distinct physical features, and growth hormone (GH) deficiency occurs in some individuals with PWS. The aim of this study is to test the hypotheses that (a) individuals with deletions and UPD have different physical and dysmorphic features, (b) individuals treated with GH have different physical and dysmorphic features than those not treated, and (c) GH treatment effects are different for individuals with UPD in comparison to those with deletions. Study participants included 30 individuals with deletions or UPD, who did or did not have GH treatment. Participants' molecular abnormalities were determined by molecular and cytogenetic analysis. Clinical data were obtained by a single dysmorphologist. Individuals with deletions were found to be heavier (p = .001), taller (p = .031), with smaller head circumferences (p = .042) and were more likely to have fair skin and hair than their family members (p = .031, .049, respectively) compared to UPD patients. Females with deletions more commonly had hypoplastic labia minora (p = .009) and clitoris (.030) in comparison to those with UPD. Individuals who received GH in both deletion and UPD groups were taller (p = .004), had larger hands (p = .011) and feet (p = .006) and a trend for a larger head circumference (p = .103). Interestingly, the GH-treated group also had a lower rate of strabismus (esotropia [p = .017] and exotropia [p = .039]). This study showed statistically significant correlations between phenotype and molecular subtypes and also between phenotype and GH treatment.

Prader-Willi syndrome (PWS) is a complex multisystemic condition caused by a lack of paternal expression of imprinted genes from the 15q11.2-q13 region. Limited literature exists on the association between molecular classes, growth hormone use, and the prevalence of psychiatric phenotypes in PWS. In this study, we analyzed nine psychiatric phenotypes (depressed mood, anxiety, skin picking, nail picking, compulsive counting, compulsive ordering, plays with strings, visual hallucinations, and delusions) recognized in PWS and investigated associations with growth hormone treatment (GHT), deletions (DEL) and uniparental disomy (UPD) in a cohort of 172 individuals with PWS who met the criteria for analysis. Associations were explored using Pearson chi-square tests and univariable and multivariable logistic regression analyses to control for confounding exposures. This observational study of the largest dataset of patients with PWS to date suggested the following genetic subtype and phenotype correlations in psychiatric behaviors: (1) skin picking was more frequent in those with DEL vs. UPD; (2) anxiety was more common in those with UPD vs. DEL; and (3) an increased frequency of anxiety was noted in the UPD group treated with GHT compared to the DEL group. No other significant associations were found between the genetic subtype or GHT including for depressed mood, nail picking, compulsive counting, compulsive ordering, playing with strings, and visual hallucinations. Further studies will be required before any conclusions can be reached.

Prader-Willi syndrome (PWS) is an imprinting genetic disorder characterized by lack of expression of genes on the paternal chromosome 15q11-q13 region. Growth hormone (GH) replacement positively influences stature and body composition in PWS. Our hypothesis was that early diagnosis delays onset of obesity in PWS. We studied 352 subjects with PWS, recruited from the NIH Rare Disease Clinical Research Network, to determine if age at diagnosis, ethnicity, gender, and PWS molecular class influenced the age they first become heavy, as determined by their primary care providers, and the age they first developed an increased appetite and began seeking food. The median ages that children with PWS became heavy were 10 years, 6 years and 4 years for age at diagnosis < 1 year, between 1 and 3 years, and greater than 3 years of age, respectively. The age of diagnosis and ethnicity were significant factors influencing when PWS children first became heavy (p < 0.01), however gender and the PWS molecular class had no influence. Early diagnosis delayed the onset of becoming heavy in individuals with PWS, permitting early GH and other treatment, thus reducing the risk of obesity-associated co-morbidities. Non-white individuals had an earlier onset of becoming heavy.

Prader-Willi syndrome (PWS) is an imprinted neurodevelopmental disease caused by a loss of paternal genes on chromosome 15q11-q13. It is characterized by cognitive impairments, developmental delay, sleep abnormalities, and hyperphagia often leading to obesity. Clinical research has shown that a lack of expression of SNORD116, a paternally expressed imprinted gene cluster that encodes multiple copies of a small nucleolar RNA (snoRNA) in both humans and mice, is most likely responsible for many PWS symptoms seen in humans. The majority of previous research using PWS preclinical models focused on characterization of the hyperphagic and metabolic phenotypes. However, a crucial understudied clinical phenotype is cognitive impairments and thus we investigated the learning and memory abilities using a model of PWS, with a heterozygous deletion in Snord116. We utilized the novel object recognition task, which doesn't require external motivation, or exhaustive swim training. Automated findings were further confirmed with manual scoring by a highly trained blinded investigator. We discovered deficits in Snord116+/- mutant mice in the novel object recognition, location memory and tone cue fear conditioning assays when compared to age-, sex- matched, littermate control Snord116+/+ mice. Further, we confirmed that despite physical neo-natal developmental delays, Snord116+/- mice had normal exploratory and motor abilities. These results show that the Snord116+/- deletion murine model is a valuable preclinical model for investigating learning and memory impairments in individuals with PWS without common confounding phenotypes.

Prader-Willi syndrome (PWS) is a genomic imprinting disorder characterized by infantile hypotonia with a poor suck and failure to thrive, hypogenitalism/hypogonadism, behavior and cognitive problems, hormone deficiencies, hyperphagia, and obesity. The Stanford Binet and Wechsler (WAIS-R; WISC-III) intelligence (IQ) tests were administered on 103 individuals with PWS from two separate cohorts [University of California, Irvine (UCI) (N = 56) and Vanderbilt University (N = 47)] and clinical information obtained including growth hormone (GH) treatment, PWS molecular classes, weight and height. Significantly higher IQ scores (p

Prader-Willi syndrome (PWS) is generally due to sporadic paternal deletions of the chromosome 15q11-q13 region followed by maternal disomy 15. Advanced maternal age is more commonly seen in those with maternal disomy 15. Environmental factors (e.g., drug use, occupational chemical exposure, infectious agents, and irradiation) could account for chromosome changes. Previous evidence of differences in male and female gametogenesis could suggest an environmental role in the causation of the paternal 15q11-q13 deletion seen in PWS. Certain occupations such as hydrocarbon-exposing occupations (e.g., landscaping, farming, and painting) and viral exposure (e.g., human coronavirus 229E causing upper respiratory infections in adults with an incorporation site in the human genome at chromosome 15q11) can be seasonal in nature and contribute to chromosome damage. To assess, we reviewed birth seasonality data in a large cohort of individuals with PWS recruited nationally (N = 355) but no significant differences were seen by month between those with the 15q11-q13 deletion compared with maternal disomy 15 when analyzing quarterly seasonal patterns. Although early evidence supported birth seasonality differences in PWS, a larger number of individuals in our recent study using advanced genetic testing methods did not find this observation.

BackgroundPrader-Willi syndrome (PWS) is due to errors in genomic imprinting. PWS is recognised as the most common known genetic cause of life-threatening obesity. This report summarises the frequency and further characterises the PWS molecular classes and maternal age effects.MethodsHigh-resolution microarrays, comprehensive chromosome 15 genotyping and methylation-specific multiplex ligation probe amplification were used to describe and further characterise molecular classes of maternal disomy 15 (UPD15) considering maternal age.ResultsWe summarised genetic data from 510 individuals with PWS and 303 (60%) had the 15q11-q13 deletion; 185 (36%) with UPD15 and 22 (4%) with imprinting defects. We further characterised UPD15 findings into subclasses based on the presence (size, location) or absence of loss of heterozygosity (LOH). Additionally, significantly older mothers (mean age=32.5 years vs 27.7 years) were found in the UPD15 group (n=145) compared with the deletion subtype (n=200).ConclusionsWe report on molecular classes in PWS using advanced genomic technology in the largest cohort to date. LOH patterns in UPD15 may impact the risk of having a second genetic condition if the mother carries a recessive mutant allele in the isodisomic region on chromosome 15. The risk of UPD15 may also increase with maternal age.

Prader-Willi syndrome (PWS) is a multi-system disorder resulting from a lack of paternal gene expression in the 15q11.2-q13 region. Using databases compiled through response questionnaires completed by families known to the Prader-Willi Syndrome Association (USA), this study tested the hypothesis that PWS genetic subtype, BMI, age of diagnosis, clinical symptoms, and growth hormone treatment differ among deceased and living individuals with PWS. Categorical and continuous variables were compared using chi-square and two-group t tests, respectively. Deceased individuals had higher rates of clinical features, including increased weight concerns, heart problems, sleep apnea, other respiratory complications, diabetes, osteoporosis, high pain tolerance, and severe skin picking, when compared to living individuals. Meanwhile, living individuals had higher rates of growth hormone use and early puberty. Obesity and subsequent consequences are the primary contributors to increased mortality in PWS. Additional emphasis on areas to decrease mortality is needed.

Prader-Willi syndrome (PWS), is a complex genetic disease affecting 1/15,000 individuals, characterized by lack of expression of genes on the paternal chromosome 15q11-q13 region. Clinical features include central hypotonia, poor suck, learning and behavior problems, growth hormone deficiency with short stature, hyperphagia, and morbid obesity. Despite significant advances in genetic testing, the mean age for diagnosis in PWS continues to lag behind. Our goal was to perform a pilot feasibility study to confirm the diagnosis utilizing different genetic technologies in a cohort of 34 individuals with genetically confirmed PWS and 16 healthy controls from blood samples spotted and stored on newborn screening (NBS) filter paper cards. DNA was isolated from NBS cards, and PWS testing performed using DNA methylation-specific PCR (mPCR) and the methylation specific-multiplex ligation dependent probe amplification (MS-MLPA) chromosome 15 probe kit followed by DNA fragment analysis for methylation and copy number status. DNA extraction was successful in 30 of 34 PWS patients and 16 controls. PWS methylation testing was able to correctly identify all PWS patients and MS-MLPA was able to differentiate between 15q11-q13 deletion and non-deletion status and correctly identify deletion subtype (i.e., larger Type I or smaller Type II). mPCR can be used to diagnose PWS and MS-MLPA testing to determine both methylation status as well as the type of deletion or non-deletion status from DNA extracted from NBS filter paper. We propose that PWS testing in newborns is possible and could be included in the Recommended Uniform Screening Panel after establishing a validated cost-effective method.

INTRODUCTION:Prader-Willi syndrome (PWS) is a complex genetic condition characterized by hyperphagia, hypotonia, low muscle mass, excess body fat, developmental delays, intellectual disability, behavioral problems, and growth hormone deficiency. This study evaluated the safety and efficacy of orally administered Diazoxide Choline Controlled-Release Tablets (DCCR) in subjects with PWS. METHOD:This was a single-center, Phase II study and included a 10-week Open-Label Treatment Period during which subjects were dose escalated, followed by a 4-week Double-Blind, Placebo-Controlled Treatment Period. RESULTS:Five female and eight male overweight or obese, adolescent and adult subjects with genetically-confirmed PWS with an average age of 15.5±2.9 years were enrolled in the study. There was a statistically significant reduction in hyperphagia at the end of the Open-Label Treatment Period (-4.32, n = 11, p = 0.006). The onset of effect on hyperphagia was rapid and greater reductions in hyperphagia were seen in subjects with moderate to severe Baseline hyperphagia (-5.50, n = 6, p = 0.03), in subjects treated with the highest dose (-6.25, n = 4, p = 0.08), and in subjects with moderate to severe Baseline hyperphagia treated with the highest dose (-7.83, n = 3, p = 0.09). DCCR treatment resulted in a reduction in the number of subjects displaying aggressive behaviors (-57.1%, n = 10, p = 0.01), clinically-relevant reductions in fat mass (-1.58 kg, n = 11, p = 0.02) and increases in lean body mass (2.26 kg, n = 11, p = 0.003). There was a corresponding decrease in waist circumference, and trends for improvements in lipids and insulin resistance. The most common adverse events were peripheral edema and transient increases in glucose. Many of the adverse events were common medical complications of PWS and diazoxide. CONCLUSION:DCCR treatment appears to address various unmet needs associated with PWS, including hyperphagia and aggressive behaviors in this proof-of-concept study. If the results were replicated in a larger scale study, DCCR may be a preferred therapeutic option for patients with PWS.

Prader-Willi syndrome (PWS) is an imprinting genetic disorder characterized by lack of expression of genes on the paternal chromosome 15q11-q13 region. Growth hormone (GH) replacement positively influences stature and body composition in PWS. Our hypothesis was that early diagnosis delays onset of obesity in PWS. We studied 352 subjects with PWS, recruited from the NIH Rare Disease Clinical Research Network, to determine if age at diagnosis, ethnicity, gender, and PWS molecular class influenced the age they first become heavy, as determined by their primary care providers, and the age they first developed an increased appetite and began seeking food. The median ages that children with PWS became heavy were 10 years, 6 years and 4 years for age at diagnosis < 1 year, between 1 and 3 years, and greater than 3 years of age, respectively. The age of diagnosis and ethnicity were significant factors influencing when PWS children first became heavy (p < 0.01), however gender and the PWS molecular class had no influence. Early diagnosis delayed the onset of becoming heavy in individuals with PWS, permitting early GH and other treatment, thus reducing the risk of obesity-associated co-morbidities. Non-white individuals had an earlier onset of becoming heavy.

Prader-Willi syndrome (PWS), an imprinted neurodevelopmental disorder characterized by metabolic, sleep and neuropsychiatric features, is caused by the loss of paternal SNORD116, containing only non-coding RNAs (ncRNAs). The primary SNORD116 transcript is processed into small nucleolar RNAs (snoRNAs), which localize to nucleoli, and their spliced host gene 116HG, which is retained at its site of transcription. While functional complementation of the SNORD116 ncRNAs is a desirable goal for treating PWS, the mechanistic requirements of SNORD116 RNA processing are poorly understood. Here we developed and tested a novel transgenic mouse which ubiquitously expresses Snord116 on both a wild-type and a Snord116 paternal deletion (Snord116+/-) background. Interestingly, while the Snord116 transgene was ubiquitously expressed in multiple tissues, splicing of the transgene and production of snoRNAs was limited to brain tissues. Knockdown of Rbfox3, encoding neuron-specific splicing factor neuronal nuclei (NeuN) in Snord116+/--derived neurons, reduced splicing of the transgene in neurons. RNA fluorescence in situ hybridization for 116HG revealed a single significantly larger signal in transgenic mice, demonstrating colocalization of transgenic and endogenous 116HG RNAs. Similarly, significantly increased snoRNA levels were detected in transgenic neuronal nucleoli, indicating that transgenic Snord116 snoRNAs were effectively processed and localized. In contrast, neither transgenic 116HG nor snoRNAs were detectable in either non-neuronal tissues or Snord116+/- neurons. Together, these results demonstrate that exogenous expression and neuron-specific splicing of the Snord116 locus are insufficient to rescue the genetic deficiency of Snord116 paternal deletion. Elucidating the mechanisms regulating Snord116 processing and localization is essential to develop effective gene replacement therapies for PWS.

INTRODUCTION:Prader-Willi syndrome (PWS) is a complex genetic disorder associated with three different genetic subtypes: deletion of the paternal copy of 15q11-q13, maternal UPD for chromosome 15 and imprinting defect. Patients are typically diagnosed because of neonatal hypotonia, dysmorphism and feeding difficulties; however, data on the prenatal features of PWS are limited. OBJECTIVE:The aim of the study was to identify and compare frequencies of prenatal and neonatal clinical features of PWS among the three genetic subtypes. METHODS:Data from 355 patients with PWS from the Rare Diseases Clinical Research Network PWS registry were used to analyse multiple maternal and neonatal factors collected during an 8-year multisite study. RESULTS:Among our cohort of 355 patients with PWS (61% deletion, 36% UPD and 3% imprinting defect) 54% were born by caesarean section, 26% were born prematurely and 34% with a low birth weight (frequencies 32%, 9.6% and 8.1%, respectively, in the general population). Fetal movements were reported as decreased in 72%. All babies were hypotonic, and 99% had feeding difficulties. Low Apgar scores (

Prader-Willi syndrome (PWS) is caused by a deficiency of imprinted genes in the 15q11-q13 region and is characterized by prenatal onset of hypotonia, poor feeding, childhood-onset obesity, hyperphagia, short stature, facial dysmorphism, intellectual disability, and behavioral problems. We studied perinatal factors in a cohort of 64 people with PWS resulting from paternal deletion of 15q11-q13 and maternal uniparental disomy (UPD) for chromosome 15. We recruited 34 individuals with deletion and 30 with UPD. We compared the frequency of multiple prenatal and neonatal factors with the general population as well as between the two genetic subtypes. Of the 64 individuals with PWS, fetal movements were decreased in 82.8%, 31.7% were born prematurely, 42.1% by Cesarean section, and 35.9% required oxytocin induction. Apgar scores were low in 34.6%, 96.8% had feeding difficulty, 50% needed tube feeding, and 6.2% subsequently had gastrostomy tube placement. On comparing findings in the deletion versus the UPD groups, we did not find many significant differences. We, however, found a higher maternal age, and also later age at diagnosis in the UPD versus the deletion group. PWS subjects have higher rates of perinatal complications, especially Cesarean section rate, hypotonia, and low Apgar scores compared to the general population. We did not find many differences between the genetic subtypes, except for later age of diagnosis of the UPD 15 group suggesting a milder phenotype. We also found that the mothers in the UPD were older, supporting the hypothesis that UPD results from nondisjunction associated trisomy rescue.

Rhythmic oscillations of physiological processes depend on integrating the circadian clock and diurnal environment. DNA methylation is epigenetically responsive to daily rhythms, as a subset of CpG dinucleotides in brain exhibit diurnal rhythmic methylation. Here, we show a major genetic effect on rhythmic methylation in a mouse Snord116 deletion model of the imprinted disorder Prader-Willi syndrome (PWS). More than 23,000 diurnally rhythmic CpGs are identified in wild-type cortex, with nearly all lost or phase-shifted in PWS. Circadian dysregulation of a second imprinted Snord cluster at the Temple/Kagami-Ogata syndrome locus is observed at the level of methylation, transcription, and chromatin, providing mechanistic evidence of cross-talk. Genes identified by diurnal epigenetic changes in PWS mice overlapped rhythmic and PWS-specific genes in human brain and are enriched for PWS-relevant phenotypes and pathways. These results support the proposed evolutionary relationship between imprinting and sleep, and suggest possible chronotherapy in the treatment of PWS and related disorders.

We describe the National Institutes of Health rare disease consortium for Prader-Willi syndrome (PWS) developed to address concerns regarding medical care, diagnosis, growth and development, awareness, and natural history. PWS results from errors in genomic imprinting leading to loss of paternally expressed genes due to 15q11-q13 deletion, maternal disomy 15 or imprinting defects. The 8 year study was conducted at four national sites on individuals with genetically confirmed PWS and early-onset morbid obesity (EMO) with data accumulated to gain a better understanding of the natural history, cause and treatment of PWS. Enrollment of 355 subjects with PWS and 36 subjects with EMO began in September 2006 with study completion in July 2014. Clinical, genetic, cognitive, behavior, and natural history data were systematically collected along with PWS genetic subtypes, pregnancy and birth history, mortality, obesity, and cognitive status with study details as important endpoints in both subject groups. Of the 355 individuals with PWS, 217 (61%) had the 15q11-q13 deletion, 127 (36%) had maternal disomy 15, and 11 (3%) had imprinting defects. Six deaths were reported in our PWS cohort with 598 cumulative years of study exposure and one death in the EMO group with 42 years of exposure. To our knowledge, this description of a longitudinal study in PWS represents the largest and most comprehensive cohort useful for investigators in planning comparable studies in other rare disorders. Ongoing studies utilizing this database should have a direct impact on care and services, diagnosis, treatment, genotype-phenotype correlations, and clinical outcomes in PWS.

BackgroundIndividuals with fragile X syndrome (FXS) have both behavioral and medical comorbidities and the latter include obesity in approximately 30% and the Prader-Willi Phenotype (PWP) characterized by severe hyperphagia and morbid obesity in less than 10%. Metformin is a drug used in individuals with type 2 diabetes, obesity or impaired glucose tolerance and it has a strong safety profile in children and adults. Recently published studies in the Drosophila model and the knock out mouse model of FXS treated with metformin demonstrate the rescue of multiple phenotypes of FXS.Materials and methodsWe present 7 cases of individuals with FXS who have been treated with metformin clinically. One case with type 2 diabetes, 3 cases with the PWP, 2 adults with obesity and/or behavioral problems and, a young child with FXS. These individuals were clinically treated with metformin and monitored for behavioral changes with the Aberrant Behavior Checklist and metabolic changes with a fasting glucose and HgbA1c.ResultsWe found consistent improvements in irritability, social responsiveness, hyperactivity, and social avoidance, in addition to comments from the family regarding improvements in language and conversational skills. No significant side-effects were noted and most patients with obesity lost weight.ConclusionWe recommend a controlled trial of metformin in those with FXS. Metformin appears to be an effective treatment of obesity including those with the PWP in FXS. Our study suggests that metformin may also be a targeted treatment for improving behavior and language in children and adults with FXS.

AimsThere are no treatments for the extreme hyperphagia and obesity in Prader-Willi syndrome (PWS). The bestPWS clinical trial assessed the efficacy, safety and tolerability of the methionine aminopeptidase 2 (MetAP2) inhibitor, beloranib.Materials and methodsParticipants with PWS (12-65 years old) were randomly assigned (1:1:1) to biweekly placebo, 1.8 mg beloranib or 2.4 mg beloranib injection for 26 weeks at 15 US sites. Co-primary endpoints were the changes in hyperphagia [measured by Hyperphagia Questionnaire for Clinical Trials (HQ-CT); possible score 0-36] and weight by intention-to-treat. ClinicalTrials.gov registration: NCT02179151.ResultsOne-hundred and seven participants were included in the intention-to-treat analysis: placebo (n = 34); 1.8 mg beloranib (n = 36); or 2.4 mg beloranib (n = 37). Improvement (reduction) in HQ-CT total score was greater in the 1.8 mg (mean difference -6.3, 95% CI -9.6 to -3.0; P = .0003) and 2.4 mg beloranib groups (-7.0, 95% CI -10.5 to -3.6; P = .0001) vs placebo. Compared with placebo, weight change was greater with 1.8 mg (mean difference - 8.2%, 95% CI -10.8 to -5.6; P < .0001) and 2.4 mg beloranib (-9.5%, 95% CI -12.1 to -6.8; P < .0001). Injection site bruising was the most frequent adverse event with beloranib. Dosing was stopped early due to an imbalance in venous thrombotic events in beloranib-treated participants (2 fatal events of pulmonary embolism and 2 events of deep vein thrombosis) compared with placebo.ConclusionsMetAP2 inhibition with beloranib produced statistically significant and clinically meaningful improvements in hyperphagia-related behaviours and weight loss in participants with PWS. Although investigation of beloranib has ceased, inhibition of MetAP2 is a novel mechanism for treating hyperphagia and obesity.

Prader-Willi syndrome (PWS) is a rare, complex multisystem genetic disorder which includes hypothalamic dysfunction, hyperphagia, cognitive and behavioral problems, increased anxiety, and compulsive behaviors. Individuals with PWS have a deficit of oxytocin producing neurons in the paraventricular nucleus of the hypothalamus. Oxytocin plays a role in regulation of feeding behaviors, social interactions, and emotional reactivity, which are all issues that significantly affect the quality of life for individuals with this syndrome. We performed a double-blind, placebo-controlled, crossover study in 24 children with PWS at three academic institutions using 5 days of intranasal oxytocin (IN-OT) or 5 days of intranasal placebo spray, followed by a 4 week washout period, and then patients returned for 5 days of treatment with the alternate source. Questionnaires, including the Aberrant Behavior Checklist, Social Responsiveness Scale, Repetitive Behavior Scale - Revised, and the Hyperphagia Questionnaire, as well as Clinical Global Impression scales were administered. Blood testing for sodium, potassium, and glucose levels on days 2, 4, and 6, and a 24 hr diet recall. All scales factor improvement from Day 3 to Day 6 favored oxytocin over placebo. No single factor showed a statistically significant difference (P

The purpose of the current study was to develop syndrome-specific standardized growth curves for growth hormone-treated Prader-Willi syndrome (PWS) individuals aged 0 to 18 years. Anthropometric growth-related measures were obtained on 171 subjects with PWS who were treated with growth hormone for at least 40% of their lifespan. They had no history of scoliosis. PWS standardized growth curves were developed for 7 percentile ranges using the LMS method for weight, height, head circumference, weight/length, and BMI along with normative 3rd, 50th, and 97th percentiles plotted using control data from the literature and growth databases. Percentiles were plotted on growth charts for comparison purposes. Growth hormone treatment appears to normalize stature and markedly improves weight in PWS compared with standardized curves for non-growth hormone-treated PWS individuals. Growth chart implications and recommended usage are discussed.

Prader-Willi syndrome (PWS) is a complex, multisystem genetic disorder characterized by endocrine, neurologic, and behavioral abnormalities. We report the first case of an unbalanced de novo reciprocal translocation of chromosomes 15 and 19, 45,XY,-15,der(19)t(15;19)(q12;p13.3), resulting in monosomy for the PWS critical chromosome region. Our patient had several typical features of PWS including infantile hypotonia, a poor suck and feeding difficulties, tantrums, skin picking, compulsions, small hands and feet, and food seeking, but not hypopigmentation, a micropenis, cryptorchidism or obesity as common findings seen in PWS at the time of examination at 6 years of age. He had seizures noted from 1 to 3 years of age and marked cognitive delay. High-resolution SNP microarray analysis identified an atypical PWS type I deletion in chromosome 15 involving the proximal breakpoint BP1. The deletion extended beyond the GABRB3 gene but was proximal to the usual distal breakpoint (BP3) within the 15q11q13 region, and GABRA5, GABRG3, and OCA2 genes were intact. No deletion of band 19p13.3 was detected; therefore, the patient was not at an increased risk of tumors from the Peutz-Jeghers syndrome associated with a deletion of the STK11 gene.

ObjectiveThe goal of this study was to generate and report standardized growth curves for weight, height, head circumference, and BMI for non-growth hormone-treated white male and female US subjects with Prader-Willi syndrome (PWS) between 3 and 18 years of age and develop standardized growth charts.MethodsAnthropometric measures (N = 133) were obtained according to standard methods from 120 non-growth hormone-treated white subjects (63 males and 57 females) with PWS between 3 and 18 years of age. Standardized growth curves were developed for the third, 10th, 25th, 50th, 75th, 90th, and 97th percentiles by using the LMS method for weight, height, head circumference, and BMI for PWS subjects along with the normative third, 50th, and 97th percentiles from national and international growth data. The LMS smoothing procedure summarized the distribution of the anthropometric variables at each age using three parameters: power of the Box-Cox transformation λ (L), median μ (M) and coefficient of variation δ (S).ResultsWeight, height, head circumference, and BMI standardized growth charts representing 7 percentile ranges were developed from 120 non-growth hormone-treated white male and female US subjects with PWS (age range: 3-18 years) and normative third, 50th, and 97th percentiles from national and international data.ConclusionsWe encourage the use of syndrome-specific growth standards to examine and evaluate subjects with PWS when monitoring growth patterns and determining nutritional and obesity status. These variables can be influenced by culture, individual medical care, diet intervention, and physical activity plans.

UnlabelledAbstract Background: Currently, there is limited information on the effects of growth hormone and of the different genetic subtypes on bone mineral density (BMD) in Prader-Willi syndrome (PWS).MethodsWe evaluated BMD in 79 individuals with the common subtypes of PWS (48 with deletion and 27 with UPD) and the effect of growth hormone treatment (n=46) vs. no growth hormone treatment.ResultsForty-four percent of the individuals studied had whole body, hip, or spine BMD

PURPOSE: Prader-Willi syndrome is an imprinting disorder characterized by typical facial, physical, and cognitive/behavioral features, resulting from lack of paternally expressed genes on chromosome 15q11.2-q13. Studies have suggested an increased risk of other imprinting disorders in children conceived by assisted reproductive techniques. This study was designed to determine the association between assisted reproductive technology and Prader-Willi syndrome. METHODS: Data on individuals with Prader-Willi syndrome were collected from three distinct sources and the proportion of assisted reproductive technology births analyzed. RESULTS: The proportions of assisted reproductive technology births in the Prader-Willi Syndrome Association (USA), Rare Diseases Clinical Research Network, and University of California, Irvine Medical Center populations were 1.0% (18/1,736), 1.0% (1/98), and 2.0% (1/50), respectively (overall 1.1%; population frequency for the United States was 1.0%). Of note, 2.4% (45/1,898) of participants were co-twins (11 born after assisted reproductive technology procedures); US twin frequency is 1.6% (P = 0.007). The proportion of individuals with maternal disomy 15/imprinting defects born after assisted reproductive technology was higher than that in the total sample, 55.6% (10/18) and 34.5% (431/1,250), respectively. CONCLUSION: This study found no association between assisted reproductive technology and Prader-Willi syndrome. There was an increased frequency of twinning. The number of individuals with maternal disomy 15/imprinting defect was nearly double in the assisted reproductive technology group as compared with the total Prader-Willi syndrome participants.

Prader-Willi syndrome (PWS), a genetic disorder of obesity, intellectual disability and sleep abnormalities, is caused by loss of non-coding RNAs on paternal chromosome 15q11-q13. The imprinted minimal PWS locus encompasses a long non-coding RNA (lncRNA) transcript processed into multiple SNORD116 small nucleolar RNAs and the spliced exons of the host gene, 116HG. However, both the molecular function and the disease relevance of the spliced lncRNA 116HG are unknown. Here, we show that 116HG forms a subnuclear RNA cloud that co-purifies with the transcriptional activator RBBP5 and active metabolic genes, remains tethered to the site of its transcription and increases in size in post-natal neurons and during sleep. Snord116del mice lacking 116HG exhibited increased energy expenditure corresponding to the dysregulation of diurnally expressed Mtor and circadian genes Clock, Cry1 and Per2. These combined genomic and metabolic analyses demonstrate that 116HG regulates the diurnal energy expenditure of the brain. These novel molecular insights into the energy imbalance in PWS should lead to improved therapies and understanding of lncRNA roles in complex neurodevelopmental and metabolic disorders.

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are oppositely imprinted autism-spectrum disorders with known genetic bases, but complex epigenetic mechanisms underlie their pathogenesis. The PWS/AS locus on 15q11-q13 is regulated by an imprinting control region that is maternally methylated and silenced. The PWS imprinting control region is the promoter for a one megabase paternal transcript encoding the ubiquitous protein-coding Snrpn gene and multiple neuron-specific noncoding RNAs, including the PWS-related Snord116 repetitive locus of small nucleolar RNAs and host genes, and the antisense transcript to AS-causing ubiquitin ligase encoding Ube3a (Ube3a-ATS). Neuron-specific transcriptional progression through Ube3a-ATS correlates with paternal Ube3a silencing and chromatin decondensation. Interestingly, topoisomerase inhibitors, including topotecan, were recently identified in an unbiased drug screen for compounds that could reverse the silent paternal allele of Ube3a in neurons, but the mechanism of topotecan action on the PWS/AS locus is unknown. Here, we demonstrate that topotecan treatment stabilizes the formation of RNA:DNA hybrids (R loops) at G-skewed repeat elements within paternal Snord116, corresponding to increased chromatin decondensation and inhibition of Ube3a-ATS expression. Neural precursor cells from paternal Snord116 deletion mice exhibit increased Ube3a-ATS levels in differentiated neurons and show a reduced effect of topotecan compared with wild-type neurons. These results demonstrate that the AS candidate drug topotecan acts predominantly through stabilizing R loops and chromatin decondensation at the paternally expressed PWS Snord116 locus. Our study holds promise for targeted therapies to the Snord116 locus for both AS and PWS.

PurposePrader-Willi syndrome (PWS) and Angelman syndrome (AS) are complex neurodevelopmental disorders caused by loss of expression of imprinted genes from the 15q11-q13 region depending on the parent of origin. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) kits from MRC-Holland (Amsterdam, The Netherlands) were used to detect PWS and AS deletion subtypes. We report our experience with two versions of the MS-MLPA-PWS/AS kit (original A1 and newer B1) in determining methylation status and deletion subtypes in individuals with PWS.MethodsMS-MLPA analysis was performed on DNA isolated from a large cohort of PWS subjects with the MS-MLPA-PWS/AS-A1 and -B1 probe sets.ResultsBoth MS-MLPA kits will identify deletions in the 15q11-q13 region but the original MS-MLPA-A1 kit has a higher density of probes at the telomeric end of the 15q11-q13 region, which is more useful for identifying individuals with atypical deletions. The newer B1 kit contains more probes in the imprinting center (IC) and adjoining small noncoding RNAs useful in identifying small microdeletions.ConclusionThe A1 kit identified the typical deletions and smaller atypical deletions, whereas the B1 kit was more informative for identifying microdeletions including the IC and SNORD116 regions. Both kits should be made available for accurate characterization of PWS/AS deletion subtypes as well as evaluating for IC and SNORD116 microdeletions.

We evaluated an inexpensive, efficient, and noninvasive technique for measuring tissue damage produced by self-injurious behavior (SIB). The technique involved computerized measurement of wound surface area (WSA) based on digital photographs. In Study 1, we compared photographic measurement to a more commonly used procedure, transparency measurement, in estimating WSA of 20 wound models. Results showed that both methods were reliable and that there was a high degree of correspondence between the 2 sets of measures. In Study 2, we compared photographic WSA measures to direct-observation measures in documenting changes over time in the SIB exhibited by a woman with Prader-Willi syndrome. Results showed that increases and decreases in observed SIB during baseline and treatment conditions corresponded with changes in WSA measures, indicating that the computer-assisted photographic technique may be useful as a corroborative measure or as a primary measure when direct observation of SIB is not feasible.

Prader-Willi syndrome (PWS) is a complex neurobehavioral condition which has been classically described as having two nutritional stages: poor feeding, frequently with failure to thrive (FTT) in infancy (Stage 1), followed by hyperphagia leading to obesity in later childhood (Stage 2). We have longitudinally followed the feeding behaviors of individuals with PWS and found a much more gradual and complex progression of the nutritional phases than the traditional two stages described in the literature. Therefore, this study characterizes the growth, metabolic, and laboratory changes associated with the various nutritional phases of PWS in a large cohort of subjects. We have identified a total of seven different nutritional phases, with five main phases and sub-phases in phases 1 and 2. Phase 0 occurs in utero, with decreased fetal movements and growth restriction compared to unaffected siblings. In phase 1 the infant is hypotonic and not obese, with sub-phase 1a characterized by difficulty feeding with or without FTT (ages birth-15 months; median age at completion: 9 months). This phase is followed by sub-phase 1b when the infant grows steadily along a growth curve and weight is increasing at a normal rate (median age of onset: 9 months; age quartiles 5-15 months). Phase 2 is associated with weight gain-in sub-phase 2a the weight increases without a significant change in appetite or caloric intake (median age of onset 2.08 years; age quartiles 20-31 months;), while in sub-phase 2b the weight gain is associated with a concomitant increased interest in food (median age of onset: 4.5 years; quartiles 3-5.25 years). Phase 3 is characterized by hyperphagia, typically accompanied by food-seeking and lack of satiety (median age of onset: 8 years; quartiles 5-13 years). Some adults progress to phase 4 which is when an individual who was previously in phase 3 no longer has an insatiable appetite and is able to feel full. Therefore, the progression of the nutritional phases in PWS is much more complex than previously recognized. Awareness of the various phases will aid researchers in unraveling the pathophysiology of each phase and provide a foundation for developing rational therapies. Counseling parents of newly diagnosed infants with PWS as to what to expect with regard to these nutritional phases may help prevent or slow the early-onset of obesity in this syndrome.

ObjectiveTo generate and report standardized growth curves for weight, length, head circumference, weight/length, and BMI for non-growth hormone-treated white infants (boys and girls) with Prader-Willi syndrome (PWS) between 0 and 36 months of age. The goal was to monitor growth and compare data with other infants with PWS.MethodsAnthropometric measures (N = 758) were obtained according to standard methods and analyzed from 186 non-growth hormone-treated white infants (108 boys and 78 girls) with PWS between 0 and 36 months of age. Standardized growth curves were developed and the 3rd, 10th, 25th, 50th, 75th, 90th, and 97th percentiles were calculated by using the LMS (refers to λ, μ, and σ) smoothing procedure method for weight, length, head circumference, weight/length, and BMI along with the normative 50th percentile using Centers for Disease Control and Prevention national growth data from 2003. The data were plotted for comparison purposes.ResultsFive separate standardized growth curves (weight, length, head circumference, weight/length, and BMI) representing 7 percentile ranges were developed from 186 non-growth hormone-treated white male and female infants with PWS aged 0 to 36 months, and the normative 50th percentile was plotted on each standardized infant growth curve.ConclusionsWe encourage the use of these growth standards when examining infants with PWS and evaluating growth for comparison purposes, monitoring for growth patterns, nutritional assessment, and recording responses to growth hormone therapy, commonly used in infants and children with PWS.

Imprinting, non-coding RNA and chromatin organization are modes of epigenetic regulation that modulate gene expression and are necessary for mammalian neurodevelopment. The only two known mammalian clusters of genes encoding small nucleolar RNAs (snoRNAs), SNRPN through UBE3A(15q11-q13/7qC) and GTL2(14q32.2/12qF1), are neuronally expressed, localized to imprinted loci and involved in at least five neurodevelopmental disorders. Deficiency of the paternal 15q11-q13 snoRNA HBII-85 locus is necessary to cause the neurodevelopmental disorder Prader-Willi syndrome (PWS). Here we show epigenetically regulated chromatin decondensation at snoRNA clusters in human and mouse brain. An 8-fold allele-specific decondensation of snoRNA chromatin was developmentally regulated specifically in maturing neurons, correlating with HBII-85 nucleolar accumulation and increased nucleolar size. Reciprocal mouse models revealed a genetic and epigenetic requirement of the 35 kb imprinting center (IC) at the Snrpn-Ube3a locus for transcriptionally regulated chromatin decondensation. PWS human brain and IC deletion mouse Purkinje neurons showed significantly decreased nucleolar size, demonstrating the essential role of the 15q11-q13 HBII-85 locus in neuronal nucleolar maturation. These results are relevant to understanding the molecular pathogenesis of multiple human neurodevelopmental disorders, including PWS and some causes of autism.

BackgroundPrader-Willi syndrome (PWS) is a complex genetic disorder with errors in genomic imprinting, generally due to a paternal deletion of chromosome 15q11-q13 region. Maternal disomy 15 (both 15s from the mother) is the second most common form of PWS resulting from a trisomic zygote followed by trisomy rescue in early pregnancy and loss of the paternal chromosome 15. However, trisomy 15 or mosaicism for trisomy 15 may be present in the placenta possibly leading to placental abnormalities affecting gestational age and delivery.Methods and subjectsWe examined growth and gestational data from 167 PWS infants (93 males and 74 females; 105 infants with 15q11-q13 deletion and 62 infants with maternal disomy 15) to determine if there are differences in gestation between the two genetic subtypes.ResultsNo significant differences in growth data (birth weight, length, head circumference) or average gestational ages were found between the two genetic subgroups. However, post-term deliveries (> 42 weeks gestation) were more common in the maternal disomy group (i.e., 12 of 62 infants) compared with the deletion group (i.e., 7 of 105 infants) (chi-square test = 6.22; p < 0.02). The distribution of gestational ages in the 15q11-q13 deletion group was more bell-shaped or normal while the distribution in the maternal disomy group suggested a bimodal pattern.ConclusionsMaternal disomy 15 in PWS may contribute to disturbances in gestational age and delivery by impacting on placental structure or function secondary to the abnormal chromosomal number in the placental cells or in mechanisms leading to the maternal disomy status in PWS infants.

BackgroundMaternally-derived duplications that include the imprinted region on the proximal long arm of chromosome 15 underlie a complex neurobehavioral disorder characterized by cognitive impairment, seizures and a substantial risk for autism spectrum disorders1. The duplications most often take the form of a supernumerary pseudodicentric derivative chromosome 15 [der(15)] that has been called inverted duplication 15 or isodicentric 15 [idic(15)], although interstitial rearrangements also occur. Similar to the deletions found in most cases of Angelman and Prader Willi syndrome, the duplications appear to be mediated by unequal homologous recombination involving low copy repeats (LCR) that are found clustered in the region. Five recurrent breakpoints have been described in most cases of segmental aneuploidy of chromosome 15q11-q13 and previous studies have shown that most idic(15) chromosomes arise through BP3:BP3 or BP4:BP5 recombination events.ResultsHere we describe four duplication chromosomes that show evidence of atypical recombination events that involve regions outside the common breakpoints. Additionally, in one patient with a mosaic complex der(15), we examined homologous pairing of chromosome 15q11-q13 alleles by FISH in a region of frontal cortex, which identified mosaicism in this tissue and also demonstrated pairing of the signals from the der(15) and the normal homologues.ConclusionInvolvement of atypical BP in the generation of idic(15) chromosomes can lead to considerable structural heterogeneity.