The American College of Medical Genetics and Genomics (ACMG) and other professional organisations recommend whole exome sequencing (WES) as a first-tier genetic test for paediatric patients with congenital anomalies, developmental delay and/or intellectual disability, which has contributed to rapidly increasing rates of genetic testing and diagnosis in this population. We present a case of Rett syndrome diagnosed in early childhood following an atypical presentation of the condition with no regression using WES. This diagnosis was facilitated by a multidisciplinary outpatient neurodevelopmental genetics programme. Non-genetics providers trained in consent for genetic testing allowed for ample access to a comprehensive genetics work-up. The subsequent diagnosis of Rett syndrome qualified this patient for additional resources, novel interventions and family support.

The SET domain containing the 2 (SETD2) gene encodes a histone methyltransferase responsible for H3K36me3 modification, playing key roles in transcriptional regulation, RNA splicing, and DNA repair. Pathogenic variants in SETD2 have been linked to variable phenotypes, including Luscan-Lumish syndrome (LLS, OMIM #616831), autosomal dominant intellectual developmental disorder 70 (MRD70, OMIM #620157), and Rabin-Pappas syndrome (RAPAS, OMIM #620155). Defining the severity of intellectual disability/developmental delay caused by SETD2 variants is important for accurate genetic counseling. This study aims to present a patient carrying a novel de novo nonsense variant in the SETD2 gene and to expand the clinical phenotype spectrum associated with SETD2 variants. A 17-year-old male with dysmorphic features, epilepsy, attention deficit and hyperactivity disorder (ADHD), and moderate intellectual disability underwent a detailed clinical and genetic evaluation. A novel de novo heterozygous nonsense variant in the SETD2 gene, NM_014159.7:c.7084C>T (NP_054878.5:p.Gln2362Ter), was identified by whole-exome sequencing. This variant was classified as likely pathogenic according to American College of Medical Genetics and Genomics (ACMG) guidelines. The patient exhibited clinical features overlapping with LLS. Further research is warranted to elucidate the mechanistic differences underlying various SETD2 variants, which will be essential for improving our understanding of SETD2-related disorders and for providing accurate genetic counseling and targeted management strategies.

Introduction: Charcot-Marie-Tooth disease (CMT) is one of the most common inherited Neuromuscular Disorders (NMDs), classified under peripheral neuropathies and characterised by progressive motor and sensory dysfunction. Although Wholeexome Sequencing (WES), gene panels, and conventional methods have improved detection rates, they often miss deep intronic, regulatory, and Structural Variants (SVs). Wholegenome Sequencing (WGS), with its comprehensive coverage of coding and non coding regions, enables the identification of variants that are often overlooked by other approaches. Aim: To assess the diagnostic utility of WGS in CMT cases that remain unresolved by WES, analysing both coding and non coding variants. Materials and Methods: The present cross-sectional diagnostic study was conducted between July 2023 and January 2025 at the Neuberg Center for Genomic Medicine (NCGM), Ahmedabad, Gujarat, India. WGS was performed on 31 clinically suspected CMT patients, including two who had previously tested negative by WES. Both coding and non coding variants including missense, nonsense, frameshift, in-frame, intronic, and 5’ Untranslated Region (UTR) mutations were analysed. Variants were classified according to American College of Medical Genetics and Genomics (ACMG) guidelines, incorporating Combined Annotation Dependent Depletion (CADD) scores and Minor Allele Frequency (MAF) thresholds. They were interpreted based on pathogenicity, inheritance patterns, and genotypephenotype correlations. Selected non coding variants in the Gap Junction Beta-1 (GJB1; c.-16-511G>C) and Lamin A/C (LMNA; c.-142C>A) genes were validated by Sanger sequencing. Results: Sequencing data from 31 participants were processed using a standardised bioinformatics pipeline. Variants were classified according to ACMG guidelines, and their frequencies were calculated. WES and WGS results were compared to determine the additional diagnostic yield. WGS identified clinically significant non coding variants in GJB1 (intronic) and LMNA (5’ UTR) in two cases, yielding a 6.5% increase over WES. Overall, 31 variants were detected: 11 (35.5%) classified as pathogenic, 2 (6.5%) as likely pathogenic, and 18 (58.0%) as Variants of Uncertain Significance (VUS), reflecting the genetic heterogeneity of CMT. Conclusion: The WGS enhances diagnostic accuracy in CMT by detecting clinically relevant non coding variants often missed by WES. This is the first report from India confirming a GJB1 intronic variant and a Lamin A/C (LMNA) 5´ UTR variant using WGS in CMT patients. These findings support the integration of WGS into routine diagnostic workflows and highlight the value of comprehensive variant analysis for early and precise genetic diagnosis.

ABSTRACTBackgroundPathogenic variants of IBA57 (OMIM ID: 615330) are usually associated with multiple mitochondrial dysfunction syndrome (MMDS) and hereditary spastic paraplegia type 74 (SPG74). Here, we present a novel compound heterozygous IBA57 mutation in a boy with severe global developmental delay, optic atrophy, spastic paraplegia, and focal epileptic seizures.MethodsThe clinical data of a child diagnosed with MMDS were retrospectively collected. Video electroencephalogram (VEEG), cranial magnetic resonance imaging (MRI), and family whole‐exome sequencing (WES) were performed. Suspected mutation sites were further confirmed using Sanger sequencing. The activities of mitochondrial respiratory chain complexes I–IV were determined in peripheral blood mononuclear cells. The phylogenetic conservation of the affected residues was assessed by multiple sequence alignment of IBA57 gene orthologs. Furthermore, we conducted a review of the relevant literature.ResultsWhole‐exome sequencing identified compound heterozygous variants in the IBA57 gene: c.395_400dup (p.V132_Q133dup) and c.832delC (p.R278Afs*23), inherited from his phenotypically normal father and mother, respectively. Biochemical assays demonstrated selective reduction of complexes I and II activities, with normal complexes III and IV, consistent with impaired 4Fe–4S cluster maturation. Phylogenetic alignment revealed strict conservation of residues V132–Q133 and R278 across vertebrates. These variants had not been previously reported in domestic or international databases. According to the American College of Medical Genetics and Genomics (ACMG) guidelines, the former was classified as a variant of uncertain significance (PM4 + PM2), while the latter was classified as likely pathogenic (PVS1 + PM2).ConclusionDiseases associated with IBA57 gene variants are autosomal recessive disorders with a broad clinical phenotypic spectrum. Early genetic testing and family screening are beneficial for the diagnosis, treatment, and prognosis of the disease.

22q11.2 deletion syndrome (22q11.2 DS) is the second most common cause of congenital heart disease. The American College of Medical Genetics and Genomics (ACMG) has recently recommended implementing non-invasive prenatal screening (NIPS) for 22q11.2 DS for all pregnant women. This study aims to assess the effectiveness of routine NIPS for screening 22q11.2 deletion in a cohort of 38,495 pregnancies from the general population. We conducted a retrospective analysis of 38,495 pregnant women who underwent NIPS at Longgang Maternal and Child Health Hospital in Shenzhen from December 2022 to March 2024. Chromosomal microarray analysis (CMA) was performed on fetuses and pregnant women identified as high-risk for 22q11.2 DS by NIPS, using amniotic fluid samples and the leukocyte cells, respectively. Of the 38,495 cases, 22 were identified as high risk for 22q11.2 deletion by NIPS. Of these, 17 underwent amniocentesis, and 5 refused prenatal diagnosis. Concordant results between CMA and NIPS were observed in 8 cases, giving a positive predictive value (PPV) of 47.06% (8/17). Nine fetuses did not show 22q11.2 deletion by CMA, although the mothers of three fetuses were identified as having a maternal 22q11.2 deletion. Follow-up on 5 cases without prenatal diagnosis revealed one postpartum case of congenital heart disease (ventricular septal defect and atrial septal defect), three cases that were terminated, and one case that continued with normal fetal development. Routine non-invasive prenatal screening for 22q11.2 deletion demonstrates practical clinical utility and provides valuable insights for identifying pregnant women at risk for 22q11.2DS.

PurposeThe genetic spectrum and early clinical indicators of familial exudative vitreoretinopathy (FEVR) remain incompletely defined, and few studies have investigated the genetic variants and clinical phenotypes associated with eoHM‐FEVR and anisometropia‐FEVR patients. The purpose of this study was to screen the pathogenic variations in 11 FEVR families and analyze the refractive status and pathogenic genes in patients with irregular dominantly inherited FEVR.MethodsThe patients with clinical diagnoses of eoHM‐FEVR or anisometropia‐FEVR were evaluated from October 2019 to August 2022. Comprehensive ophthalmic tests were performed on participants to confirm the phenotype. The genotype was identified using whole‐exon sequencing and further verified the results among other family members by Sanger sequencing. Normal protein structures were modeled with AlphaFold, whereas mutant variants were analyzed via PyMOL. Variant pathogenicity followed the American College of Medical Genetics and Genomics (ACMG) guidelines. The protein–protein interaction (PPI) network analysis with STRING and k‐means clustering was applied for detecting the interaction of genes in the candidate genes, and the ClusPro Server was used for protein–protein docking.ResultsA total of 11 FEVR families were included in the study, and all the probands were found to have high myopia in both eyes or one eye before the age of 7 years. The pathogenic variants were identified in the genes TSPAN12, LRP5, and FZD4 known to be associated with FEVR in six probands. Among 13 eoHM‐related genes, FZD4 and LRP2 encode proteins that can dock together as analyzed by ClusPro software.ConclusionThis study observed dominant inheritance of an irregular pattern in FEVR families, with asymmetric FEVR presenting as severe anisometropia. The eye with higher myopia often had more advanced FEVR and pronounced fundus changes. PPI network analysis revealed important modules of gene interaction, and the FZD4‐LRP2 complex protein was potentially related to high myopia development. For patients with high myopia or with obvious anisometropia in both eyes, more attention should be paid clinically to the comprehensive examination of the peripheral fundus and early genetic testing.

Idiopathic hypogonadotropic hypogonadism (IHH) and Kallmann syndrome (KS) are rare reproductive disorders with known genetic heterogeneity. Using exome sequencing, our group previously reported the prevalence of pathogenic and likely pathogenic (P/LP) variants in genes causing IHH/KS as the primary endpoint of our study. Here, we investigate the frequency of secondary findings (SF) to determine whether individuals with IHH/KS harbor an increased burden of P/LP variants in medically actionable genes (MAGs) defined by the American College of Medical Genetics and Genomics (ACMG). We analyzed exome sequencing data from 156 individuals with clinically confirmed IHH/KS. Variants were filtered for P/LP classification using ACMG guidelines across all 84 MAGs in ACMG SF v3.3. Sanger sequencing was used for orthogonal confirmation. The prevalence of MAG variants was compared to external control datasets from the U.K. Biobank (UKB, ~ 50,000 genomes) and the NIH eMERGE Network (~ 21,000 genomes), both based on the ACMG SF v2.0 59-gene list. Among 370,000 variants, 2 individuals (1.3%) carried validated P/LP variants in two distinct MAGs: SCN5A and MYBPC3. Genes 60-84, the additional 25 genes on the ACMG SF v3.3 list, yielded no additional variants. The prevalence of MAG variants in IHH/KS (1.3%) was not significantly different from UKB (2.0%) or eMERGE (2.5%) (OR vs. UKB: OR 0.64; 95% CI, 0.16-2.61; P = 0.57). The frequency of P/LP variants in MAGs among IHH/KS patients is comparable to the general population, suggesting that MAG variants are not common in IHH/KS in contrast to some other types of infertility.

Introduction Monogenic systemic autoinflammatory diseases (SAIDs) are driven by innate immune dysregulation, often due to aberrant inflammasome activation. Inflammasomopathies, those involving NLRP3, MEFV, MVK, and TNFRSF1A mutations, constitute a subgroup marked by sterile inflammation and overlapping symptoms like fever, serositis, rash, and arthritis. However, diagnostic complexity from phenotypic variability and incomplete penetrance makes genetic and immunological confirmation essential. This study aimed to characterize the clinical, biochemical, immunological, and genetic profiles of pediatric patients with confirmed or suspected inflammasomopathies at a tertiary care center. Methods Data were obtained from a monogenic SAID registry. Genetic analysis was performed using panel-based exome sequencing, and inflammatory biomarkers (C-reactive protein [CRP], erythrocyte sedimentation rate [ESR], MRP8/14) were evaluated. Results Over a 10-year period, 36 patients were assessed, of whom 13 (36%) had definitive diagnoses. Median age at symptom onset was 18 months, while diagnosis occurred at a mean age of 99 months, highlighting diagnostic delay. Predominant symptoms included fever (25%), ocular inflammation (12.5%), rash, arthritis, and aphthous ulcers (9% each). Despite identical NLRP3 mutations, phenotypic variation suggested roles for modifier genes or environmental factors. All patients showed laboratory signs of systemic inflammation—elevated ESR, CRP, MRP8/14, leukocytosis, and thrombocytosis—with absence of autoantibodies, consistent with autoinflammatory etiology. Overall variants were classified according to the ACMG (American College of Medical Genetics and Genomics) in inflammasome-related genes. Tailored to treatment strategies included IL-1 and TNF inhibitors. Conclusions This cohort reinforces the pivotal role of the inflammasome in monogenic autoinflammatory diseases. Clinical heterogeneity, even among patients with identical mutations, underscores the need for individualized evaluation. Early integration of clinical, immunological, and genetic data guiding targeted cytokine therapies that can mitigate innate immune dysregulation improving outcomes in children.

IntroductionProgressive myoclonic epilepsies (PME) are rare genetic disorders typically presenting with myoclonus, seizures, and cognitive decline. While several genes are associated with PME, the NUS1 gene has recently emerged as a potential cause. We report the case of a 41-year-old woman who presented with tics, myoclonus, and language difficulties followed by gait instability, tremor, absence seizures, and psychotic symptoms including persistent hallucinations and delusional misidentification.MethodsNeurology and psychiatry specialists reviewed clinical data. Brain MRI, scalp video-EEG monitoring, and [18F]-FDG-PET/MRI were performed following standardized protocols. Whole exome sequencing (WES) guided by human phenotype ontology (HPO) terms was performed, and variants were interpreted according to American College of Medical Genetics and Genomics (ACMG) guidelines. Additionally, we conducted a review of previously reported cases of NUS1 pathogenic/likely pathogenic variants associated with PME to better characterize the clinical and paraclinical features and to explore potential management strategies.ResultsA novel heterozygous frameshift likely pathogenic variant in the NUS1 gene, c.248del, p. (His83Profs*22), was identified in the patient. This finding led to the introduction of a targeted therapeutic strategy, including the initiation of metformin and a thorough revision of the patient’s existing psychiatric treatment. The patient showed an improvement in her psychiatric manifestations. However, neurological examination revealed either stable or slightly worsened signs, and she did not achieve seizure freedom.DiscussionThis is the first review of NUS1 from a PME perspective and the first report describing the exploratory use of metformin as a potential therapeutic intervention. In our case, metformin was introduced simultaneously with a change in antipsychotic treatment, so its specific clinical impact cannot be determined. Additional studies are needed to improve understanding of the benefits of using metformin and other therapeutic strategies in NUS1-related disorders. Further studies are essential to clarify the full phenotypic spectrum associated with NUS1 variants and to improve our understanding of how specific variant types and locations contribute to clinical presentation.

Synonymous single nucleotide variants (sSNVs) are increasingly recognized as contributors to disease, yet existing variant annotation databases offer limited functional insights for sSNVs. We present SynMall, a comprehensive resource designed to decipher the functional impact of synonymous variation. SynMall catalogs 25 million potential human sSNVs and integrates evolutionary and population information of sSNVs from 45 non-human species. For each human sSNV, SynMall provides multilevel annotations that combine American College of Medical Genetics and Genomics (ACMG) aligned variant interpretation information, such as allele frequencies and functional effects, with over 100 descriptors at the DNA, RNA, and protein levels. These include both handcrafted features and embeddings from large language models to support advanced representation learning. To prioritize pathogenic sSNVs, we develop SynScore, a machine learning framework that integrates ACMG guidelines and diverse biological characteristics. Benchmark comparisons show that SynScore achieves state-of-the-art performance, validating its effectiveness for genome-wide pathogenicity inference. Furthermore, SynMall enables mechanistic exploration by investigating in silico assessments and curated literature evidence to evaluate sSNV effects on miRNA-mRNA interactions, mRNA splicing, mRNA stability, and codon usage. By consolidating these features into a unified platform, we anticipate that SynMall will serve as a valuable resource for elucidating the functional role of synonymous mutations.

Background: Congenital disorders of glycosylation (CDG) encompass a broad spectrum of rare inborn errors of metabolism (IEMs), resulting in defective glycosylation of various proteins and/or lipids. PMM2-CDG is the most prevalent subtype of CDG, characterized by gene mutations leading to deficiency of the phosphomannomutase 2 (PMM2) enzyme. This research identified two Iranian patients from a consanguineous family with multiple organ dysfunctions, diagnosed with PMM2-CDG due to a pathogenic variant in the PMM2 gene. The study underscores the importance of identifying specific variants in different ethnic groups for effective genetic counseling for this disorder. Methods: The study utilized exome sequencing (ES) to identify pathogenic variants. Verification of the potential variant in affected siblings, along with segregation analysis involving the parents, affected individuals, and healthy siblings of the family, was performed using Sanger sequencing. Interpretation of the variant was informed by multiple in silico analysis tools, as well as by adhering to the guidelines set forth by the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP). Results: In the genomic investigations of the two brothers with intellectual disability (ID) and progressive movement disability, the NM_000303.3:c.647A > T (NP_000294.1:p.Asn216Ile) homozygous variant in the PMM2 gene was identified. The variant was confirmed in the affected and other family members by segregation analysis. Multiple in silico tools supported the pathogenic classification of the identified variant. Conclusions: The findings provide broader insight into variants within the PMM2 gene and offer a thorough characterization of the phenotype related to this specific variant. The data have important applications for genetic diagnosis and counseling in relevant clinical contexts, as well as for identifying common gene variants associated with various ethnic groups. Additionally, this information could serve as a valuable resource in guiding therapeutic interventions.

Migraine and epilepsy are distinct neurological disorders that co-occur as comorbid conditions as well. Despite their clinical differences, these disorders exhibit some overlapping symptoms and share underlying pathophysiological mechanisms driven by a common genetic contribution. The current study aimed to explore the genetic predisposition associated with epilepsy, migraine, and their comorbidity in both familial and sporadic cases. Whole exome sequencing carried out in 191 individuals, comprising familial and sporadic cases diagnosed with migraine (n = 63), epilepsy (n = 62), and comorbid (n = 39) involving unaffected first-degree relatives (n = 16) and healthy controls (n = 11). Variant interpretation was performed in accordance with the American College of Medical Genetics and Genomics (ACMG) guidelines. Segregation analysis was carried out by Sanger sequencing. Clinically relevant pathogenic and likely pathogenic variants were observed in the genes associated with ion channel functioning and neurotransmitter regulation in migraine as well as in epilepsy. Apart from these, variations in other genes regulating glucose transport, synaptic organization and signaling were also identified. In the epilepsy group, variants were detected in sodium channel genes (SCN1A, SCN1B, SCN2A), G protein-coupled receptor (ADGRV1), GLUT-1, and GABA transporters (SLC2A1, SLC6A1), synaptic transporter (STXBP1), and others (ICK, EFHC1, SETD1B, and DEPDC5). In the migraine group, genes including ion channel encoding gene (SCN9A, ATP1A2), GABA receptor-encoding gene (GABRA5) were noted. In individuals with migraine and epilepsy comorbidity alterations were observed in ion channel encoding gene (SCN1A, KCNMA1, and KIF1A) and other gene (COL4A1) highlighting that ion channel genes are common genetic markers shared by all three disorders. The identified variants predominantly involve genes encoding sodium, potassium, and GABA receptors that result in ion channel dysfunction and neurotransmitter imbalance. These findings highlight shared molecular pathways contributing to the pathogenesis of epilepsy, migraine, and their comorbidity. The convergence of genetic factors suggests potential avenues for the development of unified therapeutic strategies.

Background/aimHereditary neuropathies exhibit significant genetic heterogeneities, often making molecular diagnosis challenging. Clinical Exome Sequencing (CES) allows the simultaneous evaluation of a wide range of candidate genes, and can be considered an efficient approach to identifying underlying genetic causes. The present study assesses the diagnostic utility of CES in patients with clinically suspected hereditary neuropathy.Materials and methodsIncluded in the study were 21 patients with clinically suspected hereditary neuropathy who underwent CES. DNA samples were isolated from peripheral blood and subjected to CES on an Illumina NextSeq 2000 system, while a bioinformatics analysis and variant interpretation were performed using Sophia DDM® software. All identified variants were classified according to the most recent American College of Medical Genetics and Genomics (ACMG) guidelines.ResultsPathogenic or likely pathogenic variants were identified in seven patients (33.3%), of whom six (28.5%) had a confirmed molecular diagnosis consistent with the clinical phenotype. Variants of uncertain significance (VUS) were detected in nine patients (42.8%), while no clinically relevant variants were found in five (23.8%).ConclusionCES contributed significantly to the establishment of molecular diagnoses in nearly one-third of the studied cohort. However, the high prevalence of VUS underscores the limitations of current interpretation frameworks, revealing a need for functional validation and familial segregation analyses. Despite these challenges, CES remains a valuable and appropriate diagnostic tool, particularly in resource-limited healthcare settings.

This study investigates FLNC mutations in Chinese cardiomyopathy patients. Background: Inherited cardiomyopathies, including dilated cardiomyopathy (DCM), hypertrophic cardiomyopathy (HCM), restrictive cardiomyopathy (RCM), and arrhythmogenic right ventricular cardiomyopathy (ARVC) are major heart failure causes. FLNC, critical for muscle structure, is implicated in myofibrillar myopathy and isolated DCM (3–4% cases) with ventricular arrhythmias. Missense variants are linked to HCM and protein aggregation. A cohort of 25 patients with pathogenic/likely pathogenic FLNC mutations (2022–2025, Beijing Anzhen Hospital) underwent whole-exome sequencing (WES) using IDT kit 1.0/Hiseq 4000. Variants were classified via the American College of Medical Genetics and Genomics (ACMG) guidelines. Clinical data (echocardiography, CMR, labs) and follow-up data (prognosis, meds, and family history) were collected. The statistics used SPSS (p < 0.05). The mean age was 38 ± 14.6 years (13 males). There were 25 FLNC mutations: 12 single nucleotide polymorphisms (SNPs), 5 deletions, 2 duplications, and 3 deletion-insertions, classified as 6 pathogenic, 16 likely pathogenic, and 3 variants of uncertain significance (VUS). Diagnoses: 24% dilated cardiomyopathy (DCM), 8% hypertrophic cardiomyopathy (HCM), and 4% left ventricular non-compaction. Nonsense mutation carriers exhibited significantly higher tricuspid regurgitation prevalence compared to frameshift mutation carriers (6/9 vs. 2/10; p = 0.04). Echocardiography revealed reduced left ventricular ejection fraction (LVEF) (41.5 ± 14.1%), with statistically significant differences in fractional shortening (p = 0.024) and aortic root diameter (p = 0.028). Pedigree analysis confirmed that a frameshift mutation (LP) co-segregated with familial DCM and was associated with severe phenotypes, including sudden cardiac death. Furthermore, nonsense FLNC mutations correlated with increased tricuspid regurgitation severity, smaller aortic root dimensions, and reduced pulmonary artery flow velocity.

ABSTRACTAimThis study aims to expand the clinical and genetic spectrum of Krabbe disease (KD) in Chinese adult patients and to improve diagnosis and understanding of its phenotypic diversity.MethodsPatients clinically suspected of leukodystrophy were recruited between 2015 and 2025. Clinical features were collected, and whole‐exome sequencing (WES) was performed to identify potential variants. The pathogenicity of detected variants was classified according to the American College of Medical Genetics and Genomics (ACMG) standards and guidelines. Functional assays assessing protein expression, processing, secretion, subcellular localization, and enzymatic activity were conducted to further validate variant pathogenicity.ResultsFourteen unrelated patients were genetically diagnosed with KD, and their genetic and clinical features were summarized. Eleven variants in GALC were identified, including a novel missense variant c.1019C>T (p.P340L) which is not reported in the Human Gene Mutation Database (HGMD). Unlike most adult patients who typically present with spastic paraplegia, the patient carrying this variant exhibited initial symptoms of peripheral neuropathy. Functional experiments demonstrated that the variant led to impaired protein processing and localization, as well as reduced GALC enzymatic activity. Other variants including p.D56H, p.L377X, p.L441X, and p.L634S also affected GALC functions to varying degrees.ConclusionThis study enhances the genotypic and phenotypic characterization of KD in China, aiding in differential diagnosis and genetic counseling. Functional data reinforce the pathogenicity of identified variants.

BackgroundPrimary ciliary dyskinesia (PCD) is a rare genetic disorder caused by structural or functional abnormalities of motile cilia, characterized by considerable clinical and genetic heterogeneity. Although exome sequencing (ES) can improve the diagnostic rate of PCD, more than 30% of patients with clinically suspected PCD remain undiagnosed by initial ES. The American College of Medical Genetics and Genomics (ACMG) recommends periodic reanalysis of ES data to increase the diagnostic yield.MethodsWe investigated a 35-year-old female patient with bronchiectasis and a strong clinical suspicion of PCD from Peking Union Medical College Hospital. The patient’s ES data, which had initially yielded negative results in August 2024, was reanalyzed in early 2025 employing a “genotype-phenotype-inheritance pattern” strategy. A minigene assay was conducted to validate the pathogenicity of the identified CFAP54 splicing variant. Variant pathogenicity was classified according to the ACMG/AMP guidelines.ResultsThe patient had a history of rhinitis and neonatal pneumonia. Pulmonary function tests revealed moderate obstructive ventilatory dysfunction. ES reanalysis identified a homozygous variant, CFAP54 (NM_001306084.2):c.6965 + 5G >A, which was initially classified as a variant of uncertain significance. Minigene assays confirmed that this variant induced exon 50 skipping, resulting in a frameshift and a premature termination codon (loss-of-function). This variant was subsequently reclassified as “Likely Pathogenic”.ConclusionThis study is the first to describe CFAP54:c.6965 + 5G >A and confirm its pathogenicity. This finding brings the total number of CFAP54-associated PCD patients to eight and the number of distinct CFAP54 mutations to twelve, thereby enriching the phenotypic and genotypic spectrum of this gene. Furthermore, this effective strategy of “ES reanalysis + minigene verification” resolved the diagnostic dilemma in initially ES-negative PCD cases, providing a replicable molecular diagnostic framework for similar scenarios.

How do the genetic complexities of neurofibromatosis type 1 (NF1) impact reproductive counseling, preimplantation genetic testing (PGT) design, and PGT treatment? We established association between both incidence and tissue mosaicism with multiple exon deletions and specific single-nucleotide variants (SNVs) in neurofibromin 1 (NF1), a clinical actionable finding that we structured as a flowchart outlining challenges in and an approach for reproductive counseling, PGT design, and PGT treatment for NF1. NF1 has a prevalence of 1 in 2500-3000 and is one of the most frequently requested autosomal dominant indications for PGT. NF1 is a large gene with a high mutation rate, resulting in a 50% de novo occurrence, many different reported variants scattered across the gene and relatively frequent mosaicism. We conducted a retrospective, observational cohort study on PGT molecular design for NF1 in three large PGT centers (n = 281 couples), starting from the first assay for NF1 developed in 2004 until 2022. A PGT assay was developed for 281 couples with 218 different variants in NF1. Newly described variants (n = 76) were scored using the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) classification system and submitted prior to publication in the Leiden Open Variation Database (LOVD). The employed techniques were PCR-based PGT using short tandem repeat markers (n = 230), SNP-array-based PGT (n = 39), and next-generation sequencing (NGS)-based PGT (n = 12). Minisequencing (SNAPshot) or double amplification refractory mutation system (D-ARMS) was used to incorporate SNVs. Small deletions and insertions were incorporated using fragment length analysis. All PGT assays were designed and validated according to local protocols and ESHRE guidelines. Mosaicism was present in 8% of the sporadic cases (n = 13/168), of which about half were unknown prior to PGT (n = 6/13). Mosaicism was significantly higher in patients with multiple exon deletions (n = 4/6) as compared to patients with SNVs (n = 9/162) (P < 0.001, Fisher's exact test). Additionally, two recurrent SNVs were significantly associated with mosaicism (P <0.0167, Fisher's exact test). Importantly, three unrelated families with different NF1 variants in close relatives were identified. Due to its retrospective design, not all details on the genetic test results and clinical phenotype could be retrieved for some cases (n = 6). The extent to which our findings are applicable to centers worldwide depends on their local procedures and legislation. Our findings substantially impact reproductive counseling for couples with NF1, enabling informed reproductive decision-making. For couples affected with NF1 proceeding with PGT, our findings alert colleagues worldwide on NF1-specific pitfalls in PGT molecular design and treatment. There was no funding involved in the research for this publication. M.Z.E. is an inventor on two patent applications: ZL910050-PCT/EP2011/060211-WO/2011/157846 'Methods for haplotyping single cells' and ZL913096-PCT/EP2014/068315-WO/2015/028576 'Haplotyping and copy-number typing using polymorphic variant allelic frequencies'. The other authors have no competing interests to disclose. N/A.

Referral to genetics specialists plays a pivotal role in the diagnostic journey of pediatric patients with conditions such as autism spectrum disorder (ASD), developmental delay, intellectual disability, and hearing loss. Although there are referral guidelines in place from the American College of Medical Genetics and Genomics (ACMG), not all eligible children receive referrals. Among individuals with ASD, developmental delay, or intellectual disability, only about 20-50% of eligible patients are referred for genetic testing. Additionally, further barriers hinder those who are referred from accessing care. Our study delves into these often-overlooked barriers impeding pediatric patients' access to genetics services. We invited the parents of young children with one of the aforementioned conditions, identified through early intervention and related programs, including family-professional organizations, in South Carolina and Florida, to complete an online survey about referral to genetic services and barriers to attending a genetics consultation. Insurance coverage for genetic testing was a barrier both for referral to genetics and attendance at a genetics consultation. The child's race was a predictor of attendance at a genetics consultation after referral. Our study highlights the critical need to address barriers, such as insurance coverage and racial disparities, that prevent pediatric patients from accessing genetics services. We investigated barriers to genetic services for pediatric patients who were not referred or who were referred but never attended, study populations that are under-represented in the literature. This study sheds light on key factors influencing referral to and attendance at genetics consultations for pediatric patients, including race and health insurance coverage for genetic testing. Our findings emphasize the need for genetics education and improved accessibility to genetics services.

Introduction: RASopathies are among the most prevalent genetic syndromes caused by variants in the Ras/MAPK signaling pathway, affecting various systems such as the heart, craniofacial features, skin, musculoskeletal system, hearing, and vision. They can also increase the risk of secondary malignancies. Despite clinical overlaps, distinguishing features are crucial for diagnosis, as different variants lead to distinct clinical implications. This study reviews the molecular and clinical characteristics of RASopathies, focusing on neurofibromatosis type 1 (NF1) and non-NF1 RASopathies. Methods: The study analyzed 76 patients referred to our outpatient clinic over a 6-year period, all of whom were clinically diagnosed with RASopathy and confirmed in most cases by molecular testing. Patient files, clinical photographs, and laboratory results were reviewed and analyzed. A targeted multigene next-generation sequencing panel test was performed, followed by Sanger sequencing for both confirmation and segregation analysis. Multiplex ligation-dependent probe amplification was conducted in a patient with normal sequence results but strong clinical suspicion, to identify potential deletions. Results: We identified 44 pathogenic, 25 likely pathogenic variants, and 6 variants of uncertain significance based on American College of Medical Genetics and Genomics (ACMG) criteria. Among these, 14 novel variants were found – 13 in the NF1 gene and one in SOS1. NF1 variants were present in 51 cases. Additional variants, likely to represent clinically significant findings, were identified in PTPN11 (n = 11), RAF1 (n = 4), SOS1 (n = 3), RIT1 (n = 3), KRAS (n = 1), NRAS (n = 1), SOS2 (n = 1), and BRAF (n = 1). Diagnoses included 49 patients with NF1, 21 with Noonan syndrome, 2 with neurofibromatosis-Noonan syndrome, 2 with Noonan syndrome with multiple lentigines, and 1 with cardiofaciocutaneous syndrome. Here, 12% of NF1 variants were located in exon 21, 36% of PTPN11 variants in exon 3, and 75% of RAF1 variants in exon 7. Conclusion: RASopathies have a broad molecular and clinical spectrum, complicating diagnosis and management. Accurate clinical correlation and molecular analysis are essential, as different RASopathy syndromes can result from variants in the same genes, while the same syndrome may arise from different genetic alterations. This study identifies novel variants and emphasizes the need for precise diagnostic approaches in these complex disorders.

GBA1 variants cause Gaucher's disease (GD) in biallelic forms and increase Parkinson's disease (PD) risk in heterozygous carriers. Carriers of mild or severe variants (causing GD type 1 or types 2-3) can enroll in clinical trials, whereas those with GBA1 variants classified as unknown are typically excluded. We assessed the contribution of unknown variants to PD risk and their relevance for trial stratification. We meta-analyzed 34 case-control studies (24,060 PD cases, 14,465 controls). Odds ratios (ORs) were estimated using random-effects models and stratified by the American College of Medical Genetics and Genomics (ACMG) criteria. Unknown variants also classified as variants of uncertain significance (VUSs) per ACMG criteria were associated with PD (OR = 1.59, 95% confidence interval [CI]: 1.25-2.02; I2 = 0%). VUSs + likely pathogenic + pathogenic also showed an association (OR = 1.63, 95% CI: 1.28-2.06; I2 = 0%). Unknown GBA1 variants may be considered provisionally in clinical trials if also classified as VUS, likely pathogenic, or pathogenic per ACMG criteria. © 2025 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.