Deletion Carriers Research Articles

A burden of rare copy number variants in obsessive-compulsive disorder.

Current genetic research on obsessive-compulsive disorder (OCD) supports contributions to risk specifically from common single nucleotide variants (SNVs), along with rare coding SNVs and small insertion-deletions (indels). The contribution to OCD risk from rare copy number variants (CNVs), however, has not been formally assessed at a similar scale. Here we describe an analysis of rare CNVs called from genotype array data in 2248 deeply phenotyped OCD cases and 3608 unaffected controls from Sweden and Norway. Cases carry an elevated burden of CNVs ≥30 kb in size (OR = 1.12, P = 1.77 × 10-3). The excess rate of these CNVs in cases versus controls was around 0.07 (95% CI 0.02-0.11, P = 2.58 × 10-3). This signal was largely driven by CNVs overlapping protein-coding regions (OR = 1.19, P = 3.08 × 10-4), particularly deletions impacting loss-of-function intolerant genes (pLI >0.995, OR = 4.12, P = 2.54 × 10-5). We did not identify any specific locus where CNV burden was associated with OCD case status at genome-wide significance, but we noted non-random recurrence of CNV deletions in cases (permutation P = 2.60 × 10-3). In cases where sufficient clinical data were available (n = 1612) we found that carriers of neurodevelopmental duplications were more likely to have comorbid autism (P < 0.001), and that carriers of deletions overlapping neurodevelopmental genes had lower treatment response (P = 0.02). The results demonstrate a contribution of rare CNVs to OCD risk, and suggest that studies of rare coding variation in OCD would have increased power to identify risk genes if this class of variation were incorporated into formal tests.

A comprehensive overview of neuropsychiatric symptoms in adolescents with 22q11.2 deletion syndrome.

The 22q11.2 deletion syndrome (22q11DS) is associated with a variety of neuropsychiatric outcomes that vary across deletion carriers. We adopted a dimensional approach to provide a comprehensive overview of neuropsychiatric symptom expression in adolescents with 22q11DS and further our understanding of the observed phenotypical heterogeneity. Participants were 208 adolescents with 22q11DS between 10 and 19years old. Semi-structured clinical interviews and IQ tests were used to quantify symptom expression on multiple symptom dimensions, some reflecting DSM-IV diagnostic domains. We investigated symptom expression in those with and without a formal DSM-IV classification and examined between and within symptom dimensions. We used correlation analyses to explore associations between different symptom dimensions. We demonstrated inter-individual differences in symptom expression, both between and within neuropsychiatric symptom dimensions. On most symptom dimensions, more than 50% of adolescents expressed at least one clinically relevant symptom. In addition, a significant proportion of youth without a formal DSM-IV diagnosis reported clinically relevant symptoms (e.g. >85% of those without an ADHD diagnosis reported ADHD symptoms). The exploratory correlation analysis indicated mostly positive correlations between symptom dimensions. The finding that most adolescents with 22q11DS express neuropsychiatric symptoms, even in the absence of a DSM-IV classification, has substantial ramifications for guiding adequate support. Findings may spur further research into the dimensional structure of neuropsychiatric symptoms in 22q11DS and aid in uncovering mechanisms that contribute to symptom expression. Ultimately, this provides leads to improve clinical care for 22q11DS and to understand phenotypical variation in other high-risk genetic variants.

TAOK2 Drives Opposing Cilia Length Deficits in 16p11.2 Deletion and Duplication Carriers.

Copy number variation (CNV) in the 16p11.2 (BP4-BP5) genomic locus is strongly associated with autism. Carriers of 16p11.2 deletion and duplication exhibit several common behavioral and social impairments, yet, show opposing brain structural changes and body mass index. To determine cellular mechanisms that might contribute to these opposing phenotypes, we performed quantitative tandem mass tag (TMT) proteomics on human dorsal forebrain neural progenitor cells (NPCs) differentiated from induced pluripotent stem cells (iPSC) derived from 16p11.2 CNV carriers. Differentially phosphorylated proteins between unaffected individuals and 16p11.2 CNV carriers were significantly enriched for centrosomal and cilia proteins. Deletion patient-derived NPCs show increased primary cilium length compared to unaffected individuals, while stunted cilium growth was observed in 16p11.2 duplication NPCs. Through cellular shRNA and overexpression screens in human iPSC derived NPCs, we determined the contribution of genes within the 16p11.2 locus to cilium length. TAOK2, a serine threonine protein kinase, and PPP4C, a protein phosphatase, were found to regulate primary cilia length in a gene dosage-dependent manner. We found TAOK2 was localized at centrosomes and the base of the primary cilium, and NPCs differentiated from TAOK2 knockout iPSCs had longer cilia. In absence of TAOK2, there was increased pericentrin at the basal body, and aberrant accumulation of IFT88 at the ciliary distal tip. Further, pharmacological inhibition of TAO kinase activity led to increased ciliary length, indicating that TAOK2 negatively controls primary cilium length through its catalytic activity. These results implicate aberrant cilia length in the pathophysiology of 16p11.2 CNV, and establish the role of TAOK2 kinase as a regulator of primary cilium length.

Large deletions and small insertions and deletions in the factor VIII gene predict unfavorable immune tolerance induction outcome in people with severe hemophilia A and high-responding inhibitors

IntroductionHemophilia A is an inherited bleeding disorder caused by pathogenic variants in the factor VIII gene (F8), which leads to factor VIII (FVIII) deficiency. Immune tolerance induction (ITI) is a therapeutic approach to eradicate alloantibodies (inhibitors) against exogenous FVIII in people with inherited hemophilia A. Few studies have evaluated the role of F8 variants on ITI outcome. Material and methodsWe included people with severe hemophilia A (FVIII ˂ 1 international units/dL) and high-responding inhibitors (≥ 5 Bethesda units/mL lifelong) who underwent a first course of ITI. Socio-demographic, clinical and laboratory data were collected. ITI outcomes were defined as total, partial successes, and failure. Detection of intron 1 and 22 inversions was performed by polymerase-chain reaction, followed by F8 sequencing. ResultsWe included 168 people with inherited hemophilia A and high-responding inhibitors, median age 6 years at ITI start. Intron 22 inversion was the most prevalent variant (53.6 %), followed by nonsense (16.1 %), small insertion/deletion (11.3 %), and large deletion (10.7 %). In comparison with intron 22 inversion, the odds of ITI failure were 15.5 times higher (odds ratio [OR] 15.50; 95 % confidence interval [95 % CI] 4.59–71.30) and 4.25 times higher (95 % CI, 1.53–12.3) among carriers of F8 large deletions and small insertions and deletions, respectively. ConclusionF8 large deletions and small insertions/deletions predicted ITI failure after a first course of ITI in patients with severe hemophilia A and high-responding inhibitors. This is the first study to show F8 large deletions and small insertions/deletions as predictors of ITI failure.

Differential inflammatory profiles in carriers of reciprocal 22q11.2 copy number variants

BackgroundGenetic copy number variants (CNVs; i.e., a deletion or duplication) at the 22q11.2 locus confer increased risk of neuropsychiatric disorders and immune dysfunction. Inflammatory profiles of 22q11.2 CNV carriers can shed light on gene-immune relationships that may be related to neuropsychiatric symptoms. However, little is known about inflammation and its relationship to clinical phenotypes in 22q11.2 CNV carriers. Here, we investigate differences in peripheral inflammatory markers in 22q11.2 CNV carriers and explore their relationship with psychosis risk symptoms and sleep disturbance. MethodsBlood samples and clinical assessments were collected from 22q11.2 deletion (22qDel) carriers (n=45), 22q11.2 duplication (22qDup) carriers (n=29), and typically developing (TD) control participants (n=92). Blood plasma levels of pro-inflammatory cytokines, including interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ), and anti-inflammatory cytokine interleukin-10 (IL-10) were measured using a MesoScale Discovery multiplex immunoassay. Plasma levels of C-reactive protein (CRP) were measured using Enzyme-linked Immunosorbent Assay (ELISA). Linear mixed effects models controlling for age, sex, and body mass index were used to: a) examine group differences in inflammatory markers between 22qDel, 22qDup, and TD controls, b) test differences in inflammatory markers between 22qDel carriers with psychosis risk symptoms (22qDelPS+) and those without (22qDelPS-), and c) conduct an exploratory analysis testing the effect of sleep disturbance on inflammation in 22qDel and 22qDup carriers. A false discovery rate correction was used to correct for multiple comparisons. Results22qDup carriers exhibited significantly elevated levels of IL-8 relative to TD controls (q<0.001) and marginally elevated IL-8 levels relative to 22qDel carriers (q=0.08). There were no other significant differences in inflammatory markers between the three groups (q>0.13). 22qDelPS+ exhibited increased levels of IL-8 relative to both 22qDelPS- (q=0.02) and TD controls (p=0.002). There were no relationships between sleep and inflammatory markers that survived FDR correction (q>0.14). ConclusionOur results suggest that CNVs at the 22q11.2 locus may have differential effects on inflammatory processes related to IL-8, a key mediator of inflammation produced by macrophages and microglia. Further, these IL-8-mediated inflammatory processes may be related to psychosis risk symptoms in 22qDel carriers. Additional research is required to understand the mechanisms contributing to these differential levels of IL-8 between 22q11.2 CNV carriers and IL-8’s association with psychosis risk.

Loss of transient receptor potential channel 5 causes obesity and postpartum depression

Hypothalamic neural circuits regulate instinctive behaviors such as food seeking, the fight/flight response, socialization, and maternal care. Here, we identified microdeletions on chromosome Xq23 disrupting the brain-expressed transient receptor potential (TRP) channel 5 (TRPC5). This family of channels detects sensory stimuli and converts them into electrical signals interpretable by the brain. Male TRPC5 deletion carriers exhibited food seeking, obesity, anxiety, and autism, which were recapitulated in knockin male mice harboring a human loss-of-function TRPC5 mutation. Women carrying TRPC5 deletions had severe postpartum depression. As mothers, female knockin mice exhibited anhedonia and depression-like behavior with impaired care of offspring. Deletion of Trpc5 from oxytocin neurons in the hypothalamic paraventricular nucleus caused obesity in both sexes and postpartum depressive behavior in females, while Trpc5 overexpression in oxytocin neurons in knock-in mice reversed these phenotypes. We demonstrate that TRPC5 plays a pivotal role in mediating innate human behaviors fundamental to survival, including food seeking and maternal care.

Discovering the gene-brain-behavior link in autism via generative machine learning.

Autism is traditionally diagnosed behaviorally but has a strong genetic basis. A genetics-first approach could transform understanding and treatment of autism. However, isolating the gene-brain-behavior relationship from confounding sources of variability is a challenge. We demonstrate a novel technique, 3D transport-based morphometry (TBM), to extract the structural brain changes linked to genetic copy number variation (CNV) at the 16p11.2 region. We identified two distinct endophenotypes. In data from the Simons Variation in Individuals Project, detection of these endophenotypes enabled 89 to 95% test accuracy in predicting 16p11.2 CNV from brain images alone. Then, TBM enabled direct visualization of the endophenotypes driving accurate prediction, revealing dose-dependent brain changes among deletion and duplication carriers. These endophenotypes are sensitive to articulation disorders and explain a portion of the intelligence quotient variability. Genetic stratification combined with TBM could reveal new brain endophenotypes in many neurodevelopmental disorders, accelerating precision medicine, and understanding of human neurodiversity.

Longitudinal Analysis of Brain Function-Structure Dependencies in 22q11.2 Deletion Syndrome and Psychotic Symptoms

BackgroundCompared with conventional unimodal analysis, understanding how brain function and structure relate to one another opens a new biologically relevant assessment of neural mechanisms. However, how function-structure dependencies (FSDs) evolve throughout typical and abnormal neurodevelopment remains elusive. The 22q11.2 deletion syndrome (22q11.2DS) offers an important opportunity to study the development of FSDs and their specific association with the pathophysiology of psychosis. MethodsPreviously, we used graph signal processing to combine brain activity and structural connectivity measures in adults, quantifying FSD. Here, we combined FSD with longitudinal multivariate partial least squares correlation to evaluate FSD alterations across groups and among patients with and without mild to moderate positive psychotic symptoms. We assessed 391 longitudinally repeated resting-state functional and diffusion-weighted magnetic resonance images from 194 healthy control participants and 197 deletion carriers (ages 7–34 years, data collected over a span of 12 years). ResultsCompared with control participants, patients with 22q11.2DS showed a persistent developmental offset from childhood, with regions of hyper- and hypocoupling across the brain. Additionally, a second deviating developmental pattern showed an exacerbation during adolescence, presenting hypocoupling in the frontal and cingulate cortices and hypercoupling in temporal regions for patients with 22q11.2DS. Interestingly, the observed aggravation during adolescence was strongly driven by the group with positive psychotic symptoms. ConclusionsThese results confirm a central role of altered FSD maturation in the emergence of psychotic symptoms in 22q11.2DS during adolescence. The FSD deviations precede the onset of psychotic episodes and thus offer a potential early indication for behavioral interventions in individuals at risk.

Memory, mood and associated neuroanatomy in individuals with steroid sulphatase deficiency (X-linked ichthyosis).

Steroid sulphatase (STS) cleaves sulphate groups from steroid hormones, and steroid (sulphate) levels correlate with mood and age-related cognitive decline. In animals, STS inhibition or deletion of the associated gene, enhances memory/neuroprotection and alters hippocampal neurochemistry. Little is known about the consequences of constitutive STS deficiency on memory-related processes in humans. We investigated self-reported memory performance (Multifactorial Memory Questionnaire), word-picture recall and recent mood (Kessler Psychological Distress Scale, K10) in adult males with STS deficiency diagnosed with the dermatological condition X-linked ichthyosis (XLI; n = 41) and in adult female carriers of XLI-associated genetic variants (n = 79); we compared results to those obtained from matched control subjects [diagnosed with ichthyosis vulgaris (IV, n = 98) or recruited from the general population (n = 250)]. Using the UK Biobank, we compared mood/memory-related neuroanatomy in carriers of genetic deletions encompassing STS (n = 28) and non-carriers (n = 34,522). We found poorer word-picture recall and lower perceived memory abilities in males with XLI and female carriers compared with control groups. XLI-associated variant carriers and individuals with IV reported more adverse mood symptoms, reduced memory contentment and greater use of memory aids, compared with general population controls. Mood and memory findings appeared largely independent. Neuroanatomical analysis only indicated a nominally-significantly larger molecular layer in the right hippocampal body of deletion carriers relative to non-carriers. In humans, constitutive STS deficiency appears associated with mood-independent impairments in memory but not with large effects on underlying brain structure; the mediating psychobiological mechanisms might be explored further in individuals with XLI and in new mammalian models lacking STS developmentally.

Motor difficulties in 16p11.2 copy number variation.

The rare genetic variants 16p11.2 duplication and 16p11.2 deletion have opposing effects on brain structure and function, yet are associated with broadly similar clinical phenotypes that include autism, intellectual impairment, psychiatric illness, and motor difficulties. In recent years, studies have identified subtle distinctions between the phenotypic effects of 16p11.2 duplication and 16p11.2 deletion with respect to patterns of autism, intellectual impairment, and psychiatric illness. However, although divergent phenotypic findings in some motor domains have been reported, no study has yet made a comprehensive comparison of motor difficulties between 16p11.2 deletion and 16p11.2 duplication carriers to elucidate points of convergence and divergence. We sought to make such a comparison in a group of 133 16p11.2 deletion carriers, 122 duplication carriers, and 388 familial controls, hypothesizing that motor impairment would overall be greater in deletion than duplication carriers. In a series of regression models, we found that 16p11.2 deletion status tended to predict greater impairment along indices of gross motor function, but less impairment along indices of fine motor function. These findings point to a potential pattern of performance difficulties that could be investigated in future studies. Elucidating motor differences between 16p11.2 duplication and 16p11.2 deletion carriers may help in understanding the complex effect of 16p11.2 copy number variation and other rare genetic causes of autism.

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Proband-independent haplotyping based on NGS-based long-read sequencing for detecting pathogenic variant carrier status in preimplantation genetic testing for monogenic diseases.

Preimplantation genetic testing for monogenic diseases (PGT-M) can be used to select embryos that do not develop disease phenotypes or carry disease-causing genes for implantation into the mother's uterus, to block disease transmission to the offspring, and to increase the birth rate of healthy newborns. However, the traditional PGT-M technique has some limitations, such as its time consumption, experimental procedural complexity, and the need for a complete family or reference embryo to construct the haplotype. In this study, proband-independent haplotyping based on NGS-based long-read sequencing (Phbol-seq) was used to effectively construct haplotypes. By targeting the mutation sites of single gene disease point mutations and small fragment deletion carriers, embryos carrying parental disease-causing mutations were successfully identified by linkage analysis. The efficiency of embryo resolution was then verified by classical Sanger sequencing, and it was confirmed that the construction of haplotype and SNP linkage analysis by Phbol-seq could accurately and effectively detect whether embryos carried parental pathogenic mutations. After the embryos confirmed to be nonpathogenic by Phbol-seq-based PGT-M and confirmed to have normal copy number variation by Phbol-seq-based PGT-A were transplanted into the uterus, gene detection in amniotic fluid of the implanted embryos was performed, and the results confirmed that Phbol-seq technology could accurately distinguish normal genotype embryos from genetically modified carrier embryos. Our results suggest that Phbol-seq is an effective strategy for accurately locating mutation sites and accurately distinguishing between embryos that inherit disease-causing genes and normal embryos that do not. This is critical for Phbol-seq-based PGT-M and could help more single-gene disease carriers with incomplete families, de novo mutations or suspected germline mosaicism to have healthy babies with normal phenotypes. It also helps to reduce the transmission of monogenic genetic diseases in the population.

Using transcranial alternating current stimulation to enhance working memory skills in youths with 22q11.2 deletion syndrome: A randomized double-blind sham-controlled study

Abnormal cognitive development, particularly working memory (WM) deficits, is among the first apparent manifestations of psychosis. Yet, cognitive impairment only shows limited response to current pharmacological treatment. Alternative interventions to target cognition are highly needed in individuals at high risk for psychosis, like carriers of 22q11.2 deletion syndrome (22q11.2DS). Here we applied theta-tuned transcranial alternating current stimulation (tACS) between frontal and temporal regions during a visual WM task in 34 deletion carriers. We conducted a double-blind sham-controlled study over three consecutive days. The stimulation parameters were derived from individual structural MRI scan and HD-EEG data acquired at baseline (Day 1) to model current intensity and individual preferential theta peak. Participants were randomized to either sham or tACS (Days 2 and 3) and then completed a visual WM task and a control task. Our findings reveal that tACS was safe and well-tolerated among participants. We found a significantly increased accuracy in the visual WM but not the control task following tACS. Moreover, this enhancement in WM accuracy was greater after tACS than during tACS, indicating stronger offline effects than online effects. Our study therefore supports the application of repeated sessions of brain stimulation in 22q11.2DS.

Hyperinsulinemia in Sotos Syndrome with a de novo NSD1 Deletion.

Sotos syndrome belongs to the group of diseases characterised by features such as facial dysmorphism, intellectual disability, hypotonia and overgrowth. Usually, Sotos syndrome is caused by heterozygous mutations in the NSD1 gene at chromosome 5q35 or by large genomic deletions of the same region. Genotype-phenotype correlations have mainly been reported as an association of significant or major abnormalities and presence of 5q35 deletions rather than intragenic deletions or point mutations in NSD1. The congenital hyperinsulinaemic hypoglycaemia (CHI) has been described as an uncommon feature in the presentation of Sotos syndrome. Most of the patients with Sotos syndrome and transient CHI were carriers of 5q35 deletions while persistent CHI has been recently reported in individuals with point mutations or small NSD1 deletions. We report the clinical features and medical treatment in a new-born child with Sotos syndrome and CHI that was present for almost two years. Genetic cause of Sotos syndrome in this case was a novel, large genomic deletion encompassing 24 OMIM genes including the entire NSD1 gene and 6 other Morbid genes. Our report shows challenges in diagnostics and management of this rare genetic condition. We propose, that in neonatal diagnostics, the phenotypic spectrum of Sotos syndrome should include CHI as a characteristic feature and molecular genetic testing should be done by whole genome analysis.

Establishment of a Pilot Newborn Screening Program for Spinal Muscular Atrophy in Saint Petersburg.

Spinal muscular atrophy 5q (SMA) is one of the most common neuromuscular inherited diseases and is the most common genetic cause of infant mortality. SMA is associated with homozygous deletion of exon 7 in the SMN1 gene. Recently developed drugs can improve the motor functions of infants with SMA when they are treated in the pre-symptomatic stage. With aim of providing an early diagnosis, newborn screening (NBS) for SMA using a real-time PCR assay with dried blood spots (DBS) was performed from January 2022 through November 2022 in Saint Petersburg, which is a representative Russian megapolis. Here, 36,140 newborns were screened by the GenomeX real-time PCR-based screening test, and three genotypes were identified: homozygous deletion carriers (4 newborns), heterozygous carriers (772 newborns), and wild-type individuals (35,364 newborns). The disease status of all four newborns that screened positive for the homozygous SMN1 deletion was confirmed by alternate methods. Two of the newborns had two copies of SMN2, and two of the newborns had three copies. We determined the incidence of spinal muscular atrophy in Saint Petersburg to be 1 in 9035 and the SMA carrier frequency to be 1 in 47. In conclusion, providing timely information regarding SMN1, confirmation of disease status, and SMN2 copy number as part of the SMA newborn-screening algorithm can significantly improve clinical follow-up, testing of family members, and treatment of patients with SMA.

Large Deletion in the Factor VIII Gene Is a Predictor of Immune Tolerance Induction Failure in People with Severe and Moderately-Severe Hemophilia a and High-Responding Inhibitors

Introduction Immune tolerance induction (ITI) is a therapeutic approach to eradicate inhibitors against factor VIII (FVIII) in people with inherited hemophilia A (PwHA). The success of ITI is highly variable, ranging from 60% to 80% across studies. Beyond inhibitor titers, other predictors of ITI outcome are unknown. Deleterious variants in FVIII gene ( F8) are well established risk factors for alloantibodies development in PwHA, although few studies have investigated the role of F8 variants on ITI outcome.Therefore we aimed to comprehensively analyze the association of F8 pathogenic variants on ITI outcome in severe PwHA with high-responding inhibitors. Methods We included severe (FVIII &amp;lt; 1 international units [IU]/dL) and moderately-severe (FVIII 1-2 IU/dL) unrelated PwHA and high-responding inhibitors who completed ITI from a large, admixed population of two well-characterized cohorts - the HEMFIL and the Brazilian Immune Tolerance (BrazIT) studies. We collected socio-demographic, clinical and laboratory data. ITI outcomes were defined according to previous definitions as failure, partial and total successes. Inversions of intron 1 and 22 (Inv22) were detected by polymerase chain reaction (PCR), and high-throughput sequencing approaches were used to unveil the additional F8 variants. The association between F8 pathogenic variants and ITI outcome was adjusted for inhibitor levels. To investigate whether F8 variants associated with inhibitor development were also related to ITI outcome, variants were categorized as “high-risk”, “intermediate-risk”, and “low-risk” categories. These categories for inhibitor development were then compared with the outcomes of ITI (total and partial successes and failure) using data from this study. Results We included158 PwHA, median age 6.6 years at ITI start, 90.5% were severe (Table 1). Inv22 was the most prevalent variant (55.1%) (Table 1). In comparison with Inv22-1, the risk of ITI failure was about 9 times higher (adjusted odds ratio [adjOR] 9.29; 95% confidence interval [95% CI] 1.95-53.70) among carriers of large deletions (Table 2). Conversely, Inv22-2 was associated with favorable ITI outcome in a univariate analysis (OR 0.15; 0.01-0.84), and after adjustment (adjOR 0.32; 95% CI 0.02-1.96) (Table 2). F8 deleterious variants sorted as high-risk and intermediate-risk according to a previously published classification on inhibitor development were associated with failure and successful outcomes, respectively. Conclusion Our study showed that F8 large deletions are independent predictors of ITI failure, and Inv22-2 is likely to be a predictor of successful ITI. We found a correspondence between variants classified as high-risk and intermediate-risk to inhibitor development with ITI failure and success, respectively. We suggest that F8 genotyping should be considered before indication of ITI, as ITI outcome can vary according to individual variant burden.

Whole genome sequencing resolves 10 years diagnostic odyssey in familiar myxoma

Carney complex (CNC) is an ultrarare disorder causing cutaneous and cardiac myxomas, primary pigmented nodular adrenocortical disease, hypophyseal adenoma, and gonadal tumours. Genetic alterations are often missed under routine genetic testing. Pathogenic variants in PRKAR1A are identified in most cases, while large exonic or chromosomal deletions have only been reported in a few cases. Our aim was to identify the causal genetic alteration in our kindred with a clinical diagnosis of CNC and prove its pathogenic role by functional investigation. Targeted testing of PRKAR1A gene, whole exome and whole genome sequencing (WGS) were performed in the proband, one clinically affected and one unaffected relative. WGS identified a novel, large, 10,662 bp (10.6 kbp; LRG_514t1:c.-10403_-7 + 265del; hg19, chr17:g.66498293_66508954del) deletion in the promoter of PRKAR1A in heterozygous form in the affected family members. The exact breakpoints and the increased enzyme activity in deletion carriers compared to wild type carrier were proved. Segregation analysis and functional evaluation of PKA activity confirmed the pathogenic role of this alteration. A novel deletion upstream of the PRKAR1A gene was proved to be the cause of CNC. Our study underlines the need for WGS in molecular genetic testing of patients with monogenic disorders where conventional genetic analysis fails.

Beyond the Global Brain Differences: Intraindividual Variability Differences in 1q21.1 Distal and 15q11.2 BP1-BP2 Deletion Carriers

BackgroundCarriers of the 1q21.1 distal and 15q11.2 BP1-BP2 copy number variants exhibit regional and global brain differences compared with noncarriers. However, interpreting regional differences is challenging if a global difference drives the regional brain differences. Intraindividual variability measures can be used to test for regional differences beyond global differences in brain structure. MethodsMagnetic resonance imaging data were used to obtain regional brain values for 1q21.1 distal deletion (n = 30) and duplication (n = 27) and 15q11.2 BP1-BP2 deletion (n = 170) and duplication (n = 243) carriers and matched noncarriers (n = 2350). Regional intra-deviation scores, i.e., the standardized difference between an individual’s regional difference and global difference, were used to test for regional differences that diverge from the global difference. ResultsFor the 1q21.1 distal deletion carriers, cortical surface area for regions in the medial visual cortex, posterior cingulate, and temporal pole differed less and regions in the prefrontal and superior temporal cortex differed more than the global difference in cortical surface area. For the 15q11.2 BP1-BP2 deletion carriers, cortical thickness in regions in the medial visual cortex, auditory cortex, and temporal pole differed less and the prefrontal and somatosensory cortex differed more than the global difference in cortical thickness. ConclusionsWe find evidence for regional effects beyond differences in global brain measures in 1q21.1 distal and 15q11.2 BP1-BP2 copy number variants. The results provide new insight into brain profiling of the 1q21.1 distal and 15q11.2 BP1-BP2 copy number variants, with the potential to increase understanding of the mechanisms involved in altered neurodevelopment.

Long-read sequencing identifies a common transposition haplotype predisposing for CLCNKB deletions

BackgroundLong-read sequencing is increasingly used to uncover structural variants in the human genome, both functionally neutral and deleterious. Structural variants occur more frequently in regions with a high homology or repetitive segments, and one rearrangement may predispose to additional events. Bartter syndrome type 3 (BS 3) is a monogenic tubulopathy caused by deleterious variants in the chloride channel gene CLCNKB, a high proportion of these being large gene deletions. Multiplex ligation-dependent probe amplification, the current diagnostic gold standard for this type of mutation, will indicate a simple homozygous gene deletion in biallelic deletion carriers. However, since the phenotypic spectrum of BS 3 is broad even among biallelic deletion carriers, we undertook a more detailed analysis of precise breakpoint regions and genomic structure.MethodsStructural variants in 32 BS 3 patients from 29 families and one BS4b patient with CLCNKB deletions were investigated using long-read and synthetic long-read sequencing, as well as targeted long-read sequencing approaches.ResultsWe report a ~3 kb duplication of 3′-UTR CLCNKB material transposed to the corresponding locus of the neighbouring CLCNKA gene, also found on ~50 % of alleles in healthy control individuals. This previously unknown common haplotype is significantly enriched in our cohort of patients with CLCNKB deletions (45 of 51 alleles with haplotype information, 2.2 kb and 3.0 kb transposition taken together, p=9.16×10−9). Breakpoint coordinates for the CLCNKB deletion were identifiable in 28 patients, with three being compound heterozygous. In total, eight different alleles were found, one of them a complex rearrangement with three breakpoint regions. Two patients had different CLCNKA/CLCNKB hybrid genes encoding a predicted CLCNKA/CLCNKB hybrid protein with likely residual function.ConclusionsThe presence of multiple different deletion alleles in our cohort suggests that large CLCNKB gene deletions originated from many independently recurring genomic events clustered in a few hot spots. The uncovered associated sequence transposition haplotype apparently predisposes to these additional events.The spectrum of CLCNKB deletion alleles is broader than expected and likely still incomplete, but represents an obvious candidate for future genotype/phenotype association studies.We suggest a sensitive and cost-efficient approach, consisting of indirect sequence capture and long-read sequencing, to analyse disease-relevant structural variant hotspots in general.

Chromosomal deletions on 16p11.2 encompassing SH2B1 are associated with accelerated metabolic disease

New approaches are needed to treat people whose obesity and type 2 diabetes (T2D) are driven by specific mechanisms. We investigate a deletion on chromosome 16p11.2 (breakpoint 2-3 [BP2-3]) encompassing SH2B1, a mediator of leptin and insulin signaling. Phenome-wide association scans in the UK (N= 502,399) and Estonian (N= 208,360) biobanks show that deletion carriers have increased body mass index (BMI; p= 1.3×10-10) and increased rates of T2D. Compared with BMI-matched controls, deletion carriers have an earlier onset of T2D, with poorer glycemic control despite higher medication usage. Cystatin C, a biomarker of kidney function, is significantly elevated in deletion carriers, suggesting increased risk of renal impairment. In a Mendelian randomization study, decreased SH2B1 expression increases T2D risk (p= 8.1×10-6). We conclude that people with 16p11.2 BP2-3 deletions have early, complex obesity and T2D and may benefit from therapies that enhance leptin and insulin signaling.