BackgroundA dissociative subtype of posttraumatic stress disorder, known as “D-PTSD”, has been included in the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition. In addition to meeting criteria for PTSD, patients endorse prominent dissociative symptoms, namely depersonalization and derealization, or detachment from one's self and surroundings. At present, this population is supported by a highly heterogeneous and undeveloped literature. Targeted interventions are therefore lacking, and those indicated for PTSD are limited by poor efficacy, delayed onset of action, and low patient engagement. Here, we introduce cannabis-assisted psychotherapy (CAP) as a novel treatment for D-PTSD, drawing parallels to psychedelic therapy.Case presentationA 28-year-old female presented with complex D-PTSD. In a naturalistic setting, she underwent 10 sessions of CAP, scheduled twice monthly over 5 months, coupled with integrative cognitive behavioral therapy. An autonomic and relational approach to CAP was leveraged, specifically psychedelic somatic interactional psychotherapy. Acute effects included oceanic boundlessness, ego dissolution, and emotional breakthrough. From baseline to post-treatment, the patient showed a 98.5% reduction in pathological dissociation, as measured by the Multidimensional Inventory of Dissociation, no longer meeting criteria for D-PTSD. This was accompanied by decreased cognitive distractibility and emotional suffering, as well as increased psychosocial functioning. Anecdotally, the patient has sustained improvements for over 2 years to date.ConclusionsThere is urgency to identify treatments for D-PTSD. The present case, while inherently limited, underscores the potential of CAP as a therapeutic option, leading to robust and sustained improvement. Subjective effects were comparable to those produced by classic and non-classic psychedelics, such as psilocybin and ketamine. Further research is warranted to explore, establish, and optimize CAP in D-PTSD, and to characterize its role in the pharmacological landscape.

Anxiety disorders are the most common group of mental disorders, but they are often underrecognized and undertreated in primary care. Dysfunctional breathing is a hallmark of anxiety disorders; however, mainstays of treatments do not tackle breathing in patients suffering anxiety. This scoping review aims to identify the nature and extent of the available research literature on the efficacy of breathwork interventions for adults with clinically diagnosed anxiety disorders using the DSM-5 classification system. Using the PRISMA extension for scoping reviews, a search of PubMed, Embase, and Scopus was conducted using terms related to anxiety disorders and breathwork interventions. Only clinical studies using breathwork (without the combination of other interventions) and performed on adult patients diagnosed with an anxiety disorder using the DSM-5 classification system were included. From 1081 articles identified across three databases, sixteen were included for the review. A range of breathwork interventions yielded significant improvements in anxiety symptoms in patients clinically diagnosed with anxiety disorders. The results around the role of hyperventilation in treatment of anxiety were contradictory in few of the examined studies. This evidence-based review supports the clinical utility of breathwork interventions and discusses effective treatment options and protocols that are feasible and accessible to patients suffering anxiety. Current gaps in knowledge for future research directions have also been identified.

IntroductionA comorbid diagnosis of a depressive disorder is a negative prognostic factor for individuals with AN, and novel treatments are needed to target depressive symptoms in this population. One emerging promising treatment for depressive disorders is ketamine, although there is less research investigating the use of ketamine for alleviating depression in people with AN. Case reportThis study reports on four patients with a lifetime diagnosis of AN and a comorbid diagnosis of major depressive disorder who received either intramuscular ketamine (n = 2) or intranasal esketamine (n = 2) treatment from a private psychiatric clinic. Depressive symptomatology (PHQ-9) was measured prior to (es)ketamine administration on every dosing session and adverse effects were recorded during and after dosing. All patients reported a subjective decrease in depression, although only those administered intranasal esketamine showed a reduction in PHQ-9 depression scores over time. Number of doses ranged from 3 to 23. All patients tolerated treatment well and no serious adverse effects emerged, however nausea/vomiting was experienced by one patient on one dosing session. Weight remained stable in all cases, although notably across all patients, weight at the beginning of treatment was within a "healthy" range. DiscussionThese findings suggest that (es)ketamine may reduce depressive symptoms in people with major depressive disorder and a comorbid diagnosis of AN. Future feasibility and pilot trials are warranted in order to elicit robust data on efficacy, acceptability, safety and tolerability.

Importance:Psilocybin shows promise as a treatment for major depressive disorder (MDD).Objective:To evaluate the magnitude, timing, and durability of antidepressant effects and safety of a single dose of psilocybin in patients with MDD.Design, Setting, and Participants:In this phase 2 trial conducted between December 2019 and June 2022 at 11 research sites in the US, participants were randomized in a 1:1 ratio to receive a single dose of psilocybin vs niacin placebo administered with psychological support. Participants were adults aged 21 to 65 years with a Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition diagnosis of MDD of at least 60 days' duration and moderate or greater symptom severity. Exclusion criteria included history of psychosis or mania, active substance use disorder, and active suicidal ideation with intent. Participants taking psychotropic agents who otherwise met inclusion/exclusion criteria were eligible following medication taper. Primary and secondary outcomes and adverse events (AEs) were assessed at baseline (conducted within 7 days before dosing) and at 2, 8, 15, 29, and 43 days after dosing.Interventions:Interventions were a 25-mg dose of synthetic psilocybin or a 100-mg dose of niacin in identical-appearing capsules, each administered with psychological support.Main Outcomes and Measures:The primary outcome was change in central rater-assessed Montgomery-Asberg Depression Rating Scale (MADRS) score (range, 0-60; higher scores indicate more severe depression) from baseline to day 43. The key secondary outcome measure was change in MADRS score from baseline to day 8. Other secondary outcomes were change in Sheehan Disability Scale score from baseline to day 43 and MADRS-defined sustained response and remission. Participants, study site personnel, study sponsor, outcome assessors (raters), and statisticians were blinded to treatment assignment.Results:A total of 104 participants (mean [SD] age, 41.1 [11.3] years; 52 [50%] women) were randomized (51 to the psilocybin group and 53 to the niacin group). Psilocybin treatment was associated with significantly reduced MADRS scores compared with niacin from baseline to day 43 (mean difference,-12.3 [95% CI, -17.5 to -7.2]; P <.001) and from baseline to day 8 (mean difference, -12.0 [95% CI, -16.6 to -7.4]; P < .001). Psilocybin treatment was also associated with significantly reduced Sheehan Disability Scale scores compared with niacin (mean difference, -2.31 [95% CI, 3.50-1.11]; P < .001) from baseline to day 43. More participants receiving psilocybin had sustained response (but not remission) than those receiving niacin. There were no serious treatment-emergent AEs; however, psilocybin treatment was associated with a higher rate of overall AEs and a higher rate of severe AEs.Conclusions and Relevance:Psilocybin treatment was associated with a clinically significant sustained reduction in depressive symptoms and functional disability, without serious adverse events. These findings add to increasing evidence that psilocybin-when administered with psychological support-may hold promise as a novel intervention for MDD.Trial Registration:ClinicalTrials.gov Identifier: NCT03866174.

This study reports on 10 frontline healthcare workers, employed during the COVID-19 pandemic and experiencing symptoms of burnout and PTSD, treated with group ketamine-assisted psychotherapy (KAP) in a private outpatient clinic setting. Participants attended 6 sessions once weekly. These included 1 preparation session, 3 ketamine sessions (2 sublingual, 1 intramuscular), 2 integration sessions. Measures of PTSD (PCL-5), depression (PHQ-9), and anxiety (GAD-7) were administered at baseline and post-treatment. During ketamine sessions, the Emotional Breakthrough Inventory (EBI) and the 30-item Mystical Experience Questionnaire (MEQ-30) were recorded. Participant feedback was gathered 1-month post-treatment. We observed improvements in participants' average PCL-5 (59% reduction), PHQ-9 (58% reduction), and GAD-7 (36% reduction) scores from pre- to post-treatment. At post-treatment, 100% of participants screened negative for PTSD, 90% had minimal/mild depression or clinically significant improvement, and 60% had minimal/mild anxiety or clinically significant improvement. MEQ and EBI scores had large variations among participants at each ketamine session. Ketamine was well tolerated, and no significant adverse events were reported. Participant feedback corroborated findings of improvements observed in mental health symptoms. We found immediate improvements treating 10 frontline healthcare workers experiencing burnout, PTSD, depression, and anxiety using weekly group KAP and integration.

Aim: There is limited real-world evidence for patients with treatment-resistant depression (TRD) receiving esketamine nasal spray. Methods: This retrospective cohort study used data collected from a psychiatric clinic's EHR system. Results: A total of 171 TRD patients received esketamine July 2019-June 2021. This predominantly female, white population had several mental health comorbidities and high exposure to psychiatric medications. We observed significant reductions (p < 0.001) in average PHQ-9 and GAD-7 scores from baseline (PHQ-9: mean: 16.7; SD: 5.8; GAD-7: mean: 12.0; SD: 5.8) to last available treatment (PHQ-9: mean: 12.0; SD: 6.4; GAD-7: mean: 8.7; SD: 5.6). There were no reports of serious adverse events. Conclusion: This study found a significant disease burden for patients with TRD. Esketamine appears to be well tolerated and effective in improving depression and anxiety.

Background and Aims Ketamine and esketamine have garnered interest in both psychiatric research and clinical practice for treatment-resistant depression (TRD). In this review, we examined registered trials investigating the therapeutic use of ketamine or esketamine for TRD, with the aim of characterizing emerging trends and knowledge gaps. Methods The ClinicalTrials.gov electronic registry and results database was queried from inception to February 5, 2022, adhering to elements of the PRISMA guideline, we evaluated trial eligibility in the qualitative synthesis. Data regarding study design, drug regimens, and measures were subsequently abstracted and descriptively analyzed. Results The search returned 86 records, of which 56 trials were included in the final review. The number of trials investigating ketamine and esketamine for TRD increased since 2008, with higher peaks observed in 2015 ( n = 9) and 2021 ( n = 9). Most trials were Phase 2 (13, 23.2%) or Phase 3 (11, 19.6%), gathering preliminary data on efficacy and/or further data on safety and efficacy with variant dosing and pharmacological approaches. By and large, trials examined ketamine and esketamine as individual versus combination treatments (45% and 25%, respectively). The Montgomery-Asberg Depression Rating Scale (MADRS) was most commonly used to assess clinical outcomes (75%). Conclusions There are increasingly large-scale and late-phase trials of esketamine over ketamine for TRD, coupled with efforts to centralize evidence on these medications. Yet several trials do not assess patient characteristics that may affect treatment response, such as age, sex, and race. By understanding these design limitations, scientists and clinicians can avoid research waste and funding bodies can judiciously direct support towards high priority research.

Eating disorders (EDs) are serious, life-threatening psychiatric conditions associated with physical and psychosocial impairment, as well as high morbidity and mortality. Given the chronic refractory nature of EDs and the paucity of evidence-based treatments, there is a pressing need to identify novel approaches for this population. The noncompetitive N-methyl-D-aspartate receptor (NMDAr) antagonist, ketamine, has recently been approved for treatment-resistant depression, exerting rapid and robust antidepressant effects. It is now being investigated for several new indications, including obsessive–compulsive, post-traumatic, and substance use disorder, and shows transdiagnostic potential for EDs, particularly among clinical nonresponders. Hence, the aim of this review is to examine contemporary findings on the treatment of EDs with ketamine, whether used as a primary, adjunctive, or combination psychopharmacotherapy. Avenues for future research are also discussed. Overall, results are encouraging and point to therapeutic value; however, are limited to case series and reports on anorexia nervosa. Further empirical research is thus needed to explore ketamine efficacy across ED subgroups, establish safety profiles and optimize dosing, and develop theory-driven, targeted treatment strategies at the individual patient level.

ObjectiveThe aim of this study was to estimate the cost-effectiveness of esketamine nasal spray relative to intravenous ketamine for patients with treatment-resistant depression (TRD) in the US.MethodsWe used a Markov model with a 1-month cycle length, and we estimated quality-adjusted life years (QALYs), costs (2020 USD), and incremental cost-effectiveness ratios (ICER) of esketamine relative to ketamine over a 3-year time horizon, from both the healthcare sector and patient perspectives. We ran the model using efficacy estimates from both clinical trial and real-world effectiveness (RWE) data. One-way and probabilistic sensitivity analyses (PSAs) were performed to evaluate the robustness of findings.ResultsOver a 3-year time horizon, the use of esketamine yielded 1.98 QALYs (RWE/clinical trial efficacy), and the use of ketamine yielded 2.03 QALYs (clinical trial efficacy) or 1.99 QALYs (RWE). Esketamine was dominated by ketamine using the healthcare perspective. ICERs were above $150,000/QALY threshold with the patient perspective. Under the healthcare perspective, PSA showed there are no scenarios where esketamine was cost-effective compared to ketamine. With the patient's perspective, the probability that esketamine was cost-effective compared to ketamine was 0.0055 (clinical trial efficacy) and 0.35 (RWE).LimitationsThe data utilized for efficacy have limitations. The time horizon may fail to capture longer-term costs and benefits.ConclusionsIn this decision analytic model evaluating esketamine versus ketamine for TRD, we found esketamine unlikely to be cost-effective under a healthcare sector perspective. Under a patient perspective, esketamine had similar effectiveness and was less costly than ketamine due to insurance coverage.

BACKGROUND:Depression and anxiety outcome measures, safety/tolerability, patient satisfaction, and ease of implementation of group-based ketamine-assisted psychotherapy (G-KAP) delivered to patients in intensive residential eating disorder (ED) treatment were assessed.CASE PRESENTATION:This study reports on five participants with a diagnosis of an ED and comorbid mood and anxiety disorders who received weekly intramuscular ketamine injections in a group setting over 4 weeks. Measures of anxiety (GAD-7) and depression (PHQ-9) were administered pre-dose, 4-h post-dose, and 24-h post dose. Four of the 5 participants experienced clinically significant improvements on the PHQ-9 score (i.e., change greater than 5) while 2 of the 5 participants experienced clinically significant improvements on the GAD-7 score (i.e., change greater than 4) from pre-dose to 24-h post-dose after the last ketamine session. Dosing sessions were well tolerated, and no serious adverse events were reported. Clinical observations and participant reports corroborated improvements in depression and anxiety symptoms, good tolerability of ketamine treatment, and practical implementation of the G-KAP protocol in a residential ED treatment center.CONCLUSIONS:This study suggests the potential utility of G-KAP as an adjunct to intensive, specialized ED treatment. Overall, this novel, cross-diagnostic intervention warrants future research to further explore its appropriateness in a treatment setting.

Background Ketamine has emerged as a promising pharmacotherapy for depression and other mental illnesses, and the intramuscular (IM) administration of ketamine is now offered at many North American outpatient psychiatric clinics. However, a characterization of the outpatient population receiving IM ketamine treatment, and an evaluation of the real-world efficacy and safety of long-term IM ketamine treatment, has not been reported. This study aimed to evaluate the clinical characteristics, treatment patterns, clinical outcomes, and adverse events of patients receiving IM ketamine treatment. Methods Patient data from the electronic health records of a private outpatient psychiatric clinic network in the United States were collected and analyzed retrospectively. Adults who received ketamine treatment only by IM administration from January 2018 to June 2021 were included; a total of 452 patients were included in the cohort. Results Patients receiving IM ketamine treatment had a mean of 2.8 (SD 1.4) psychiatric diagnoses. 420 (93%) patients had a diagnosis of major depressive disorder, 243 (54%) patients had a diagnosis of generalized anxiety disorder, and 126 (28%) patients had a diagnosis of post-traumatic stress disorder. Thirty-seven percent (42/114) of patients reported a history of a previous suicide attempt, and patients had an average of 3.1 (SD 2.9) psychiatric medication prescriptions at baseline. Patients received between 1 and 48 IM ketamine treatments. Average depression and anxiety symptoms both significantly improved by 34% (p < .001) from baseline (PHQ-9: mean=16.3, SD=6.7; GAD-7: mean=12.8, SD=5.7) to patients’ last treatment (PHQ-9: mean=10.8, SD=5.9; GAD-7: mean=8.4, SD=5.5), and suicidal ideation scores also significantly improved (p < .001) . With maintenance ketamine treatments, median improvements in depression and anxiety of at least 21% and 19% were maintained for over 13 months. An adverse event occurred during 59 of 2,532 treatments (2.3%). Conclusions IM ketamine is being utilized to treat psychiatric outpatients with a moderate-to-severe mental health history and multiple mental illnesses not limited to depression. Average depression and anxiety levels significantly improve throughout IM ketamine treatment and do not regress to baseline for over one year with maintenance treatments. Prospective studies are recommended to confirm the long-term efficacy and safety of IM ketamine.

Ketamine has emerged as a promising pharmacotherapy for depression and other mental illnesses, and the intramuscular (IM) administration of ketamine is now offered at many North American outpatient psychiatric clinics. However, a characterization of the outpatient population receiving IM ketamine treatment, and an evaluation of the real-world efficacy and safety of long-term IM ketamine treatment, has not been reported.

Abstract Background Ketamine has emerged as a promising pharmacotherapy for depression and other mental illnesses, and the intramuscular (IM) administration of ketamine is now offered at many North American outpatient psychiatric clinics. However, a characterization of the outpatient population receiving IM ketamine treatment and an evaluation of the real-world depression, anxiety, and safety outcomes of long-term psychiatric IM ketamine treatment has not been reported. This study aimed to evaluate the clinical characteristics, treatment patterns, clinical outcomes, and adverse events of patients receiving IM ketamine treatment. Methods Patient data from the electronic health records of a private outpatient psychiatric clinic network in the United States were collected and analyzed retrospectively. Adults with any psychiatric diagnosis who received ketamine treatment only by IM administration from January 2018 to June 2021 were included. A total of 452 patients were included in the cohort. Results Patients receiving IM ketamine treatment had a mean of 2.8 (SD 1.4) psychiatric diagnoses. 420 (93%) patients had a diagnosis of major depressive disorder, 243 (54%) patients had a diagnosis of generalized anxiety disorder, and 126 (28%) patients had a diagnosis of post-traumatic stress disorder. Patients received a median of 4 (range 1–48) IM ketamine treatments. Median depression scores (PHQ-9) improved 38% from 16.0 (IQR 11.3–21.8) at baseline to 10.0 (IQR 6.0–15.0) at last treatment (p < .001). Median anxiety scores (GAD-7) improved 50% from 14.0 (IQR 8.0–17.0) at baseline to 7.0 (IQR 4.3–11.8) at last treatment (p < .001). With maintenance ketamine treatments, average improvements in depression (PHQ-9) and anxiety (GAD-7) scores of at least 4.7 and 4.9 points were maintained for over 7 months. An adverse event occurred during 59 of 2532 treatments (2.3%). Conclusions IM ketamine is being utilized to treat psychiatric outpatients with multiple mental illnesses not limited to depression. Average depression and anxiety levels significantly improve throughout IM ketamine treatment and do not regress to baseline during patients’ maintenance treatment phase. Prospective studies are recommended to confirm the long-term effectiveness and safety of IM ketamine.

The genetic underpinnings of most pediatric-cancer cases are unknown. Population-based studies use large sample sizes but have accounted for only a small proportion of the estimated heritability of pediatric cancers. Pedigree-based studies are infeasible for most human populations. One alternative is to collect genetic data from a single nuclear family and use inheritance patterns within the family to filter candidate variants. This approach can be applied to common and rare variants, including those that are private to a given family or to an affected individual. We evaluated this approach using genetic data from three nuclear families with 5, 4, and 7 children, respectively. Only one child in each nuclear family had been diagnosed with cancer, and neither parent had been affected. Diagnoses for the affected children were benign low-grade astrocytoma, Wilms tumor (stage 2), and Burkitt’s lymphoma, respectively. We used whole-genome sequencing to profile normal cells from each family member and a linked-read technology for genomic phasing. For initial variant filtering, we used global minor allele frequencies, deleteriousness scores, and functional-impact annotations. Next, we used genetic variation in the unaffected siblings as a guide to filter the remaining variants. As a way to evaluate our ability to detect variant(s) that may be relevant to disease status, the corresponding author blinded the primary author to affected status; the primary author then assigned a risk score to each child. Based on this evidence, the primary author predicted which child had been affected in each family. The primary author’s prediction was correct for the child who had been diagnosed with a Wilms tumor; the child with Burkitt’s lymphoma had the second-highest risk score among the seven children in that family. This study demonstrates a methodology for filtering and evaluating candidate genomic variants and genes within nuclear families that may merit further exploration.

Members of a paralogous gene family in which variation in one gene is known to cause disease are eight times more likely to also be associated with human disease. Recent studies have elucidated DHX30 and DDX3X as genes for which pathogenic variant alleles are involved in neurodevelopmental disorders. We hypothesized that variants in paralogous genes encoding members of the DExD/H-box RNA helicase superfamily might also underlie developmental delay and/or intellectual disability (DD and/or ID) disease phenotypes. Here we describe 15 unrelated individuals who have DD and/or ID, central nervous system (CNS) dysfunction, vertebral anomalies, and dysmorphic features and were found to have probably damaging variants in DExD/H-box RNA helicase genes. In addition, these individuals exhibit a variety of other tissue and organ system involvement including ocular, outer ear, hearing, cardiac, and kidney tissues. Five individuals with homozygous (one), compound-heterozygous (two), or de novo (two) missense variants in DHX37 were identified by exome sequencing. We identified ten total individuals with missense variants in three other DDX/DHX paralogs: DHX16 (four individuals), DDX54 (three individuals), and DHX34 (three individuals). Most identified variants are rare, predicted to be damaging, and occur at conserved amino acid residues. Taken together, these 15 individuals implicate the DExD/H-box helicases in both dominantly and recessively inherited neurodevelopmental phenotypes and highlight the potential for more than one disease mechanism underlying these disorders.

Human Phenotype Ontology (HPO) has risen as a useful tool for precision medicine by providing a standardized vocabulary of phenotypic abnormalities to describe presentations of human pathologies; however, there have been relatively few reports combining whole genome sequencing (WGS) and HPO, especially in the context of structural variants.

KBG syndrome is a rare autosomal dominant genetic condition characterized by neurological involvement, macrodontia and distinct facial, hand and skeletal features. Over 70 cases have been reported; however it is likely that KBG syndrome is underdiagnosed due to lack of comprehensive characterization of the heterogeneous phenotypic features. We describe the clinical manifestations in a male currently at 13 years of age, who exhibited symptoms including epilepsy, severe developmental delay, distinct facial features and hand anomalies, without positive genetic diagnosis. Subsequent exome sequencing identified a novel de novo heterozygous single base pair insertion (c.6015dupA) in ANKRD11, which was validated by Sanger sequencing. This insertion is predicted to lead to a premature stop codon and loss of function in ANKRD11, thereby implicating it as contributing to the proband’s symptoms and yielding a molecular diagnosis of KBG syndrome for the case. INTRODUCTION Whole exome sequencing (WES) is a method that sequences only regions of the genome that code for proteins and is more comprehensive than other testing methods such as microarray and CNV analyses. WES is useful for detecting disease-contributing variants in genes associated with rare genetic syndromes. Here we report our efforts in phenotypic characterization and molecular diagnosis of a previously undiagnosed pediatric patient. This case demonstrates the utility of whole exome sequencing for finding rare disease-contributing mutations that can then lead to the diagnosis of rare, previously unrecognized syndromes. In this case, we report in a single family the identification of a de novo mutation in ANKRD11, which led to the recognition of KBG syndrome in the sequenced proband. RESULTS Clinical presentation and family history The proband was born to a non-consanguineous couple, who had an unremarkable pregnancy history; however, at birth a large fontanel was reported. Parents and siblings were healthy and no significant family history was reported (Figure 1). The proband had his first epileptic episode at three years of age. After this episode, he lost all speech, began exhibiting autistic behavior, and also started to have frequent generalized tonic-clonic seizures. Over time, tonic, atonic, mild clonic, complex partial, myoclonic and gelastic seizures were reported in the proband. Other developmental skills, including throwing a ball, responding to his name, feeding himself with utensils and self-care skills were lost by 4-years of age. No significant conductive hearing loss, heart abnormalities or delayed bone age were found in the proband at that age. The proband was evaluated (by G.J.L.) at eleven years of age. He presented with several neurological and craniofacial abnormalities including epilepsy, ventriculomegaly, relative macrocephaly, prominent forehead, low hairline, thick eyebrows, wide-set eyes (Figure 2), macrodontia of upper central incisors, and full lips (Figure 3). Hand and foot abnormalities included clinodactyly of the fifth digit, bilateral single transverse palmar creases, brachydactyly (Figure 4) and flat feet. He also had a diagnosis of cerebral folate deficiency due to the presence of folate receptor autoantibodies. Genomic Analyses Blood and saliva samples from the proband as well as his parents and siblings were used as samples to be sequenced. These samples were sent to Affiliated Genetics in Salt Lake City, Utah, where genomic DNA was extracted and exons sequenced using the Life Technologies Ampliseq Exome RDY kit and the Life Technologies Proton sequencing system (see Methods). These targeted regions were sequenced using the Ion Proton sequencing system using Ion Hi-Q Chemistry with 200 bp reads. The DNA sequencing data was compared to the UCSC hg19 reference sequence using several methods of analysis (see Methods). These analyses included in-house protocols and several commercial software packages including Tute Genomics, Omicia Opal, and Cartagenia v4.1, along with the use of an OTG-SNP Caller pipeline (see Methods). The various analyses helped to provide a more comprehensive and in-depth approach to the data . As one example, for the OTG-SNP Caller pipeline, for each individual, the final VCF file contained 20,000 to 25,000 variants, of which around 300-400 variants were found to be autosomal recessive, i.e. heterozygous in both parents, and homozygous only in the proband. However, over a thousand variants were recognized as de novo, which is notably above the expected number of de novo mutations found in WES . Therefore, even with an optimized variant calling pipeline, there were still a significant number of false positives called. Autosomal variants were examined, and there was no evidence found to support any of them as possible contributing mutations. These variants are provided in supplementary files (as described in Methods). For the de novo mutations, a single base insertion of adenine (A) at position 6015 in exon 10 of ANKRD11 (c.6015dupA , p.Gly2006Argfs*26) (Figure 5) was identified as the most relevant mutation. All phenotypic analysis software, including Phenolyzer, wAnnovar, and PhenIX indicated that a heterozygous frame-shift mutation in ANKRD11, or the Ankyrin Repeat Domain 11 gene, might be a contributing factor in this individual’s disease The presence of the mutation was confirmed using Sanger sequencing (Figure 5).This mutation has a CADD score of 32, and is considered to be ‘Deleterious’ by SIFT with a confidence score of 0.858, and is therefore predicted to have a severe effect on protein structure. Although insertions have not yet been reported, other mutations in this gene have been previously identified as contributing to KBG syndrome, a rare disease that affects around 60-70 people worldwide. DISCUSSION Many syndromes affecting neurological development present with heterogeneous and non-distinct phenotypes and therefore remain undiagnosed or are misdiagnosed. The combination of whole exome sequencing combined with detailed and standardized phenotypic documentation is a powerful method to achieve accurate diagnosis. KBG syndrome (OMIM #148050) is a rare, but increasingly recognized, autosomal dominant genetic condition. It was first described in 1975, and is characterized by craniofacial features, hand abnormalities, macrodontia, and neurological involvement including developmental delay and epilepsy. The syndrome’s name was derived from the last names of the first three families found to have this syndrome. Over 70 cases have been reported, however it is likely that KBG syndrome is underdiagnosed due to the fact that dysmorphic features may be subtle and cognitive delay can vary from mild to moderate. Skjei et al. suggested that a clinical diagnosis of KBG syndrome can be made if the individual meets four out of the following eight major criteria: characteristic facial features, macrodontia of upper central incisors, short stature, delayed bone age, neurological involvement, hand abnormalities, costovertebral anomalies and the presence of a family member affected with the syndrome. Facial features include hypertelorism, short nose with broad base and bulbous nasal tip, and broad bushy eyebrows. Although the shape of the face is often described as being round, it has been noted that the shape evolves as affected children develop. Hand abnormalities typically include brachydactyly, clinodactyly of the fifth digit, small hands and nail anomalies. Skeletal anomalies frequently involve the pelvis, thorax, limbs and skull with abnormal curvature of the spine, including kyphosis and scoliosis, being reported in some cases. Minor features of KBG syndrome include cutaneous syndactyly, conductive hearing loss, palatal abnormalities, cryptorchidism, webbed/short neck, strabismus and congenital heart defects. There is some phenotypic overlap with Cornelia de Lange syndrome (CdLS) . Individuals with KBG syndrome have been found to have heterozygous mutations leading to haploinsufficiency of the ankyrin repeat domain 11 (ANKRD11) gene or a 16q24 microdeletion that encompasses ANKRD11. When mutations in the gene affect the highly conserved region of the domain for transcriptional regression, they are predicted to lead to premature stop codons which could result in haploinsufficiency and when a 16q24 microdeletion is present the haploinsufficency of ANKRD11 is confirmed to be the pathogenic mechanism of KBG syndrome. Sporadic and familial cases of KBG syndrome have been reported, with familial cases following an autosomal dominant inheritance pattern 15; . ANKRD11 (ankyrin repeat domain containing protein 11) is a chromatin regulator that controls histone acetylation and gene expression during neural development. There are two functional domains that act as transcriptional repressors and one domain that functions as a transcriptional promoter. The majority of reported mutations in KBG syndrome result in a truncated protein that affects a domain for transcriptional repression. ANKRD11 interacts with the p160 coactivator and the nuclear receptor complex and it functions to inhibit ligand-dependent transcriptional activation by recruiting histone deacytelases (HDACs). Additionally, ANKRD11 was also found to play a role in enhancing the transcriptional activity of p53. Homozygosity for a missense mutation in ANKRD11 is embryonic lethal in mice, whereas the heterozygous mice have an osteopenia-like phenotype and craniofacial abnormalities. This sporadic case of KBG syndrome demonstrates the importance of ongoing investigations of rare conditions. Each case reported in the literature will help to delineate the phenotype so that we may better identify cases in the future and determine appropriate recommendations for clinical management. Current recommendations for management of KBG syndrome include hearing tests, ophthalmologic assessments, echocardiography, an EEG, orthodontic evaluation and skeletal investigation wit

KBG syndrome is a rare autosomal dominant genetic condition characterized by neurological involvement and distinct facial, hand, and skeletal features. More than 70 cases have been reported; however, it is likely that KBG syndrome is underdiagnosed because of lack of comprehensive characterization of the heterogeneous phenotypic features. We describe the clinical manifestations in a male currently 13 years of age, who exhibited symptoms including epilepsy, severe developmental delay, distinct facial features, and hand anomalies, without a positive genetic diagnosis. Subsequent exome sequencing identified a novel de novo heterozygous single base pair duplication (c.6015dupA) in ANKRD11, which was validated by Sanger sequencing. This single-nucleotide duplication is predicted to lead to a premature stop codon and loss of function in ANKRD11, thereby implicating it as contributing to the proband's symptoms and yielding a molecular diagnosis of KBG syndrome. Before molecular diagnosis, this syndrome was not recognized in the proband, as several key features of the disorder were mild and were not recognized by clinicians, further supporting the concept of variable expressivity in many disorders. Although a diagnosis of cerebral folate deficiency has also been given, its significance for the proband's condition remains uncertain.