Journal of Medical Genetics

Heterozygous FOXL2 (non-)coding sequence and structural variants (SVs) lead to blepharophimosis, ptosis and epicanthus inversus syndrome (BPES), a rare, autosomal dominant developmental disorder characterized by a completely penetrant eyelid malformation and incompletely penetrant primary ovarian insufficiency (POI). We collected variants from our in-house database, generated via clinical genetic testing and downstream research testing in the Center for Medical Genetics Ghent, Belgium (2001-2024), and via literature and other resources in the same period. All retrieved variants were categorized using ACMG/AMP classifications to increase the knowledge of pathogenicity. We collected 413 unique genetic defects of the FOXL2 region, including 76 novel variants, in 864 index patients. Of these, 87% of patients were identified with a coding FOXL2 sequence variant. The polyalanine tract is a known mutational hotspot of FOXL2, illustrated here by the high percentage of pathogenic polyalanine expansions (24%). Furthermore, the molecular spectrum in typical BPES index patients is characterized by 8% coding deletions and 3% deletions located up- and downstream of FOXL2. The remaining 2% carry translocations along with chromosomal rearrangements of 3q23. This uniform and structured reclassification, incorporating the largest dataset of variants implicated in FOXL2-associated disease so far, will improve both the diagnosis as well as genetic counselling for individuals with BPES.

IRF2BPL-related disorder is a neurodevelopmental disorder caused by heterozygous variants in the IRF2BPL (Interferon Regulatory Factor 2 Binding Protein-Like) gene. The few reports available in the literature suggest that common symptoms include developmental delay, intellectual disability and developmental regression. There are no reports of genotype-phenotype correlations. We developed a retrospective and prospective patient-reported survey to assess diagnostic information, presenting symptoms and longitudinal follow-up of neurological symptoms for up to two years. Clinical information was available for all 32 participants and was highly variable in regards to age at symptom onset, severity of neurologic manifestations, and progressivity. For 27 of the 32 participants, diagnostic genetic test results were available. Genetic mutation analysis revealed 22 individuals with truncating variants and five participants with unique missense variants in IRF2BPL. The study data support the hypothesis that IRF2BPL missense variants are associated with a less severe disease presentation and progression than participants with truncating variants. The purpose of this study is to further define IRF2BPL-related disorder and provide more clinical and molecular insight into this ultra-rare disease. Highlights- Patient-reported clinical history at diagnosis and up to two years of follow up - The clinical spectrum is increasingly heterogeneous - We report 32 patients, 27 with noted IRF2BPL variants, 14 being novel to literature. - Data supports the notion that IRF2BPL missense variants may be associated with less severe disease than truncations (nonsense/frameshifts).

Lipoedema is a chronic adipose tissue disorder mainly affecting women with excess subcutaneous fat deposition on the lower limbs, associated with pain and tenderness. There is often a family history of lipoedema, suggesting a genetic origin, but the contribution of genetics is not well studied. We conducted a genome-wide association study (GWAS) for this disorder in a clinically ascertained cohort from Spain and performed a meta-analysis with the UK lipoedema cohort GWAS. We then used the results of this study as a replication of the inferred UK Biobank "lipoedema phenotype" study. Whilst our meta-analysis alone did not identify any genome-wide significant associations, our clinical cohorts provide support for three loci identified through the UKBB study: the chr2q24.3 GRB14-COBLL1 locus (rs6753142, PMETA=1.64x10-6), chr6p21.1 VEGFA locus (rs4711750, PMETA=8.99x10-7) and the chr5q11.2 ANKRD55-MAP3K1 locus (rs3936510, PMETA=1.67x10-5). We identify numerous rare SNPs with strong association signals in our meta-analysis (P<1x10-6) with support in both UK and Spanish datasets, three of which also show nominal support in the UKBB (P<0.05). These findings provide a starting point towards understanding the genetic basis of clinical lipoedema and demonstrate the utility of the interplay of large-scale biobanks genetic data and clinically ascertained cohorts to elucidate the genetic architecture of lipoedema.

CTNNB1 syndrome is a rare neurodevelopmental disorder caused by pathogenic variants in the CTNNB1 gene. Although its core clinical manifestations have been increasingly recognised, longitudinal data on cognitive, behavioural and motor trajectories remain limited, and the craniofacial phenotype has not previously been quantitatively characterised. This study provides longitudinal evidence on the cognitive, clinical and psychological profile of individuals with CTNNB1 syndrome, together with a detailed three-dimensional morphometric analysis of facial morphology. Cognitive, clinical, psychological and neuropsychological data were collected at two time points (T0 and T1), separated by a one-year interval, using a comprehensive and standardised assessment protocol. Longitudinal analyses indicated stability across most domains, with no evidence of systematic regression. A significant improvement in gross motor functioning was observed, particularly among younger participants. Linear mixed-effects models showed that age moderated developmental change, with younger individuals exhibiting greater gains over time in gross motor skills and adaptive behaviour compared to older participants. Three-dimensional facial morphometric analyses revealed a distinctive and statistically significant craniofacial pattern associated with CTNNB1 syndrome, independent of age and facial size. This phenotype was characterised by midfacial narrowing, reduced midface projection and mandibular retrusion. Importantly, facial shape variation was significantly associated with externalising behavioural problems and clinically relevant behavioural difficulties, suggesting a link between craniofacial morphology and behavioural severity. This study represents the first integrated longitudinal characterisation of CTNNB1 syndrome combining neurodevelopmental follow-up with quantitative craniofacial phenotyping. The findings indicate slow but progressive improvement in specific clinical domains during childhood and adolescence, alongside relative stability in global adaptive functioning, and highlight three-dimensional facial morphology as a sensitive structural biomarker for phenotypic stratification and clinical monitoring in CTNNB1 syndrome. Lay summaryThis study is the first to describe how children with CTNNB1 syndrome, a rare genetic condition that leads to global developmental delays, develop over time. We also performed advanced facial analysis to look for common facial features among patients.

BackgroundGenetic skeletal disorders (GSDs) comprise a heterogeneous group of rare, predominantly monogenic conditions that are increasingly diagnosed through high-throughput sequencing. While gene discovery has progressed rapidly, interpretation of pathogenic and uncertain variants remains a major bottleneck, in part because their functional consequences are determined at the protein structure level. However, a systematic assessment of structural knowledge across GSD-associated genes is currently lacking. Here, we present a comprehensive protein structure-centric analysis of 674 protein-coding genes implicated in GSDs. MethodsWe integrated experimental structures, AlphaFold2 (AF2) models, multimeric states, protein-protein interactions, and ClinVar variant annotations. ResultsWe quantify experimental structural availability and sequence coverage, revealing that 37% of GSD proteins lack any experimental structure and that, among proteins with structures, sequence coverage is often incomplete. We show that AF2 models provide high-confidence structural information for a substantial subset of proteins lacking experimental data, but that model reliability strongly correlates with existing structural coverage. Analysis of multimeric assemblies and co-occurring partners demonstrates that many GSD proteins function as obligate multimers, highlighting the importance of interface-level interpretation of variants. Finally, mapping clinically annotated missense variants onto representative protein structures illustrates how structural context can inform the interpretation of pathogenic and uncertain variants, particularly at interaction interfaces. ConclusionsTogether, this work provides a structure-aware reference framework for GSD genes, highlighting systematic gaps in current protein knowledge and demonstrating how integration of structural data can guide genomic variant interpretation. Our observations support a broader principle of structural equivalence, whereby distinct variants converge on shared structural perturbations that explain clustering patterns and enable mechanistic interpretation of nearby variants of uncertain significance.

BackgroundHeterozygous c.283+1G>A and c.283G>A variants in the THRB gene, encoding for thyroid hormone receptor (TR){beta}1 and {beta}2, lead to autosomal dominant macular dystrophy (ADMD). We report the detailed clinical characterization of two first-degree relatives with ADMD, heterozygous for THRB c.283+1G>A, and an unrelated ADMD patient with a novel variant, c.283G>C. The genomic and molecular consequences of both variants were studied. MethodsgDNA and mRNA were obtained from leukocytes. Clinical characterization included biochemistry, bone density and body composition, ECG, echocardiography, ultrasound, audiometry and color-vision. In vitro assays investigated TR function and DNA binding. ResultsThe patients manifested no resistance to thyroid hormone beta (RTH{beta}) and had normal FT4 and TSH. Detailed studies in two patients showed no goiter, tachycardia, hypercholesterinemia or hepatic steatosis. Hearing was not impaired. Both had impaired color vision and reduced bone density. RT-PCR from all three patients revealed skipping of exon 4 exclusive to TR{beta}1, producing a deletion of 87 amino acids in the N-terminal domain (TR{beta}1{Delta}NTD). In vitro, DNA-binding affinity of TR{beta}1{Delta}NTD to DR4-TRE with or without RXR was comparable to TR{beta}1WT. Surprisingly, TR{beta}1{Delta}NTD was transcriptionally twice more active than TR{beta}1WT with a similar EC50 for T3, demonstrating gain-of-function of TR{beta}1{Delta}NTD. THRA expression in leukocytes was increased by 3-fold compared to unrelated controls and different from RTH{beta} patients. ConclusionThese THRB splice site variants produce TR{beta}1 exon 4 skipping, resulting in a gain-of-function mutant, TR{beta}1{Delta}NTD. This explains the dominant ADMD phenotype devoid of RTH{beta} and suggests a TR{beta}1 gain-of-function syndrome.

Cancer Variant Interpretation Group UK was established in 2017 in response to the publication of the 2015 ACMG/AMP v3 guidance for the interpretation of sequence variants. Its initial purpose was to ensure consistency in the UK clinical-laboratory community implementation of ACMG/AMP v3 guidance for cancer susceptibility genes (CSGs). Still convening for monthly national meetings, the remit of CanVIG-UK now encompasses additional activities delivered under the following objectives: O_LICreation of a national multidisciplinary professional network and regular forum. C_LIO_LIDelivery of training and education. C_LIO_LIEstablishment of a consensus approach to the fundamentals of variant interpretation in cancer susceptibility genes. C_LIO_LIDevelopment and ratification of gene-specific frameworks for variant interpretation for cancer susceptibility genes. C_LIO_LIDevelopment and maintenance of an online platform to facilitate information sharing and variant interpretation within the UK clinical-laboratory community. C_LIO_LIFacilitation of UK contribution to international variant interpretation endeavours. C_LI A survey of CanVIG-UK members evaluating the impact of these activities conducted in November 2025 had 163 responses, including 113 clinical scientists/trainees and 27 Clinical Genetics consultants/trainees. The utility of the CanVIG-UK consensus recommendations for variant interpretation in cancer susceptibility genes was highly rated, with 89/145=61.4% of survey respondents reporting using the guidance at least weekly ([&ge;]4 times/month) and 124/128=96.9% rating it as extremely/very useful. The usage frequency and utility of the gene-specific guidance reported by survey respondents were similar to those reported for the main consensus specification. Both qualitative and quantitative survey responses clearly demonstrate the value of the CanVIG-UK activities to the clinical-diagnostic community. Key messagesO_LIWhat is already known on this topic: Cancer Variant Interpretation Group UK (CanVIG-UK) is a national subspeciality multidisciplinary network first established in 2017. It brings together members of the UK clinical-laboratory community to improve accuracy and consistency in the interpretation of variants in cancer susceptibility genes (CSG) C_LIO_LIWhat this study adds: this article presents the results of a survey of CanVIG-UK members, demonstrating the impact of CanVIG-UK activities on their services, as well as a review of progress in the six updated objectives of CanVIG-UK C_LIO_LIHow this study might affect research, practice or policy: this article presents current priorities and practices and potential future directions for variant interpretation in CSGs across the UK and Republic of Ireland C_LI

PIK3CA-related overgrowth spectrum (PROS) comprises a heterogeneous group of disorders caused by postzygotic activating variants in PIK3CA, leading to mosaic activation of the PI3K pathway. PROS natural history is highly variable across patients and depends on the timing and distribution of the somatic mutation. To better characterize this natural history, we analyzed a cohort of patients with PROS. This multicenter study was conducted at Hopital Femme Mere Enfant (Lyon, France) and Hopital Necker-Enfants Malades (Paris, France). We included pediatric and adult patients with PROS who had a documented PIK3CA variant and at least two MRI examinations performed at least 2 months apart. Patients undergoing interventional surgical or radiological procedures, or receiving targeted therapies were excluded. In all patients, target lesions were identified on baseline MRI scans, and assessed on follow-up scans according to the RECIST criteria. Among 67 PROS patients screened from 2008 to 2021, 30 met the inclusion criteria, including 43.3% female patients. The median age at first MRI was 19 years (interquartile range, 5 to 34) and the median interval between the two scans was 75.7 months (range, 2.1 to 160.3 months). Recurrent localizations included the face (n = 6; 20%), ear, nose and throat region (n = 3; 10%), upper limbs (n = 5; 16.7%), thorax (n = 3; 10%), abdomen (n = 4; 13.3%), pelvis (n = 5; 16.7%), and lower limbs (n = 10; 33.3%), with some patients presenting multisite involvement. During the observation period, 86.6% (n = 26) of patients exhibited an increase in target lesion volume, with a median progression of 37.8% (range, 2.6 to 233.0%) and a mean progression of 52% (standard error of the mean, 7.2%), reflecting a right-skewed distribution driven by a subset of rapidly enlarging lesions. In conclusion, this study provides the first radiological description of the natural history of PROS, demonstrating that tissue malformations most often enlarge over time, with sustained progression persisting into adulthood.

Interpreting HNF1B variants is challenging in clinical practice. We aimed to integrate functional, clinical, and family data to improve variant classification, describe clinical features of carriers and report registry-level prevalence of HNF1B alterations. Clinical, genetic, and family data were analyzed from the Norwegian MODY Registry (NMR) and the Norwegian Childhood Diabetes Registry (NCDR). Clinical features of sequence variant and 17q12 deletion (17q12del) carriers were summarized, and variants were classified using ACMG-AMP-ClinGen criteria. Registry-level prevalence was reported with 95% confidence intervals. HNF1B sequence variants were functionally assessed, showing that the lower transactivation (TA) was associated with higher clinical severity. Eleven variants demonstrated impaired functional activity, with TA inversely correlated with clinical burden ({varrho} = -0.701, p = 0.002). We identified 28 individuals with 17q12del (21 in NMR, seven in NCDR) and 15 individuals carrying 14 unique (LP/P) sequence variants, all detected in the NMR. Overall, 36/486 probands (7.4%) with genetically confirmed monogenic diabetes in the NMR carried an LP/P HNF1B sequence variant or 17q12del. In the NCDR, [~] 0.2% carried 17q12del (7/3,583; 3/7 GADA/IA-2A-positive). Functional data enabled reclassification of three variants. Since many pediatric 17q12del carriers in the NMR were referred for testing due to structural renal anomalies without diabetes, HNF1B screening should be considered in children with renal/extra-renal features, irrespective of diabetes or autoantibody status. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=86 SRC="FIGDIR/small/26348894v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@41feforg.highwire.dtl.DTLVardef@ccc48borg.highwire.dtl.DTLVardef@17ba2e9org.highwire.dtl.DTLVardef@4919b_HPS_FORMAT_FIGEXP M_FIG C_FIG

ImportanceGenome-wide association studies have identified hundreds of common single nucleotide polymorphisms (SNPs) and small insertions/deletions (indels) associated with primary open-angle glaucoma (POAG) risk, though these variants have modest effect sizes and individually may have minor contributions to disease development. As whole-genome sequencing data is becoming more readily available, structural variants and other complex genomic features can be interrogated for contribution to disease risk. ObjectiveTest the association of structural variants in known glaucoma loci with disease risk. DesignCross-sectional study. SettingA multicenter cohort of individuals from the United States who contributed genomic and electronic health record data to the All of Us Research Program. ParticipantsPOAG case/control cohorts were generated in the All of Us Researcher Workbench using age (>40 for cases, >65 for controls) and ICD 9/10 diagnosis codes. Main Outcomes and MeasuresLogistic regression analyses adjusted for age, sex, and the top 10 principal components of ancestry were used to test association of structural variants within 500 kilobases of 309 known open-angle glaucoma risk loci. The significance threshold after Bonferroni correction was set at p<1.6x10-4. Results516 POAG cases and 18,716 controls of European ancestry from the All of Us v8 data release were included in the analysis. Mean age was 77.0 years among cases and 74.7 years among controls. Females comprised 45.7% of cases and 56.5% of controls. An 8,732 base pair deletion upstream of PITX2 (chr4:110680827-110689558) was associated with 7.3-fold higher odds of POAG (95% confidence interval: 2.9-18.5, p= 2.4x10-5, variant carrier frequency= 1.6% in cases and 0.25% in controls). Functional annotation identified multiple enhancers overlapping the deletion, suggesting that this structural variant likely impacts gene regulation and expression. Conclusion and RelevanceWhole genome sequencing data captures rare structural variants with large effect sizes that are missed by conventional SNP and indel genotyping approaches, enabling improved POAG risk stratification. These data also expand the phenotypic spectrum of structural variation in the PITX2 locus from childhood glaucoma to adult-onset disease, where age at diagnosis and clinical severity may be influenced by the extent of disrupted regulatory elements.

PURPOSE To evaluate cancer risk, age-specific penetrance, and mortality associated with heterozygous pathogenic or likely pathogenic (P/LP) germline PALB2 variants identified through genomic ascertainment and to assess modification by family history of cancer. PATIENTS AND METHODS We conducted a case-control study in two large population-based adult cohorts: the UK Biobank (n=469,580) and Geisinger MyCode (n=167,050). Individuals with heterozygous PALB2 P/LP variants were identified via exome sequencing and compared with non-carriers. Cancer diagnoses and vital status were obtained from linked registry and electronic health record data. We used multivariable logistic regression to estimate odds ratios (ORs) for cancer outcomes and Cox proportional hazards models to estimate hazard ratios (HRs) for all-cause mortality. Age-specific cumulative incidence (penetrance) was estimated using Kaplan-Meier methods. Models were adjusted for birth year, sex (when applicable), smoking status, and body mass index; stratified analyses assessed modification by family history of cancer. RESULTS PALB2 P/LP variant prevalence was 1:571 in UK Biobank and 1:940 in MyCode, with the higher prevalence in the UK cohort driven by the PALB2 p.Trp1038Ter founder variant. Compared with non-carriers, heterozygotes had significantly increased odds of any cancer, female breast cancer, pancreatic cancer, and cancers of ill-defined or secondary sites in both cohorts (P < 0.01). Adjusted hazard ratios for any cancer and female breast cancer ranged from 1.7 to 3.6. All-cause mortality was increased among PALB2-heterozygotes (HR 1.61-1.67), and survival after cancer diagnosis was reduced. Family history further modified cancer risk. CONCLUSION Genomic ascertainment of PALB2-heterozygotes identifies elevated risk for multiple cancers and increased mortality, although risks were lower than estimates from familial ascertainment. These findings inform risk management for individuals identified through genomic screening.

Introduction: Nonsyndromic cleft lip with or without cleft palate (NSCL/P) is a common congenital malformation with complex etiology involving both genetic and environmental factors. Epigenetic mechanisms may mediate environmental contributions, but separating genetic from environmental effects remains challenging. Methods: We present an epigenome-wide association study with 32 monozygotic and 22 dizygotic twin pairs discordant for NSCL/P on blood and saliva samples. Differential methylation analysis was conducted using linear models to identify CpG sites showing significant methylation differences between affected and unaffected twins followed by functional annotation and pathway enrichment analysis. Results: The top-ranked finding is a differentially methylated region comprising two CpG sites at the CYP26A1 locus, cg12110262 (P = 3.21x10-7) and cg15055355 (P = 1.39x10-3). CYP26A1 is essential for retinoic acid catabolism and craniofacial patterning. The chromatin regulator ANKRD11, which causes KBG syndrome featuring cleft palate was the second best hit. Differentially methylated CpG sites showed significant enrichment in craniofacial enhancers and overlap with multiple GWAS-validated cleft genes including VAX1, PVRL1, SMAD3, and PRDM16. Conclusions: Our findings implicate retinoic acid signaling and chromatin regulation in NSCL/P etiology and demonstrate the value of discordant twin designs for distinguishing environmental from genetic epigenetic contributions to complex malformations.

Wolfram syndrome is a rare genetic disorder characterized by antibody-negative early-onset atypical diabetes mellitus, optic nerve atrophy, sensorineural hearing loss, diabetes insipidus (arginine vasopressin deficiency), and progressive neurodegeneration, with significant variability in disease severity. We assessed the accuracy of a genotype-based severity scoring system to predict the onset of cardinal symptoms in Wolfram syndrome. This system is based on the type of WFS1 variants (in-frame or out-of-frame) and their location relative to transmembrane domains. Severity scores were assigned to 324 patients with documented onset ages for diabetes mellitus, optic atrophy, hearing loss, and diabetes insipidus. Our analysis revealed a clear correlation between severity scores and earlier onset of diabetes mellitus and optic atrophy. Patients with in-frame variants outside transmembrane domains exhibited milder symptoms, especially WFS1 c.1672C>T (p.Arg558Cys) variant, whereas those with out-of-frame variants showed the earliest onset. Severity scores 3 and 4 did not follow the expected progression, suggesting that transmembrane domain involvement in both alleles may result in greater severity. These findings suggest that this scoring system provides valuable insights into the progression of Wolfram syndrome and may guide clinical care. Further refinement may improve its utility for predicting the onset of non-diabetic symptoms.

Protocadherin-12 (PCDH12), a cell-adhesion protein belonging to the non-clustered protocadherin family, plays a crucial role in the establishment and regulation of neuronal connections and communication. Bi-allelic loss-of-function (LoF) variants in the PCDH12 gene have been associated with several neurodevelopmental disorders (NDDs) such as diencephalic-mesencephalic junction dysplasia syndrome, cerebral palsy, and cerebellar ataxia, often accompanied by ocular abnormalities. However, genotypes exhibit variable expressivity. Affected individuals sharing the same PCDH12 variant presenting differing phenotypic severities have posed major challenges towards identification of the underlying pathogenic mechanisms. Here, we report three affected individuals from two families, each harbouring non-truncating pathogenic missense variants in PCDH12. The patients are compound heterozygous, with each individual carrying one extracellular [c.1742T>G (p.Val581Gly) and c.1861_2del/insCA (p.Ile621His)] and one intracellular variant [c.3370C>T (p.Arg1124Cys) and c.3445G>A (p.Asp1149Asn] on each allele. The children present with a range of phenotypes similar to those associated with LoF variants. One child exhibited microcephaly and seizures, while the two siblings displayed developmental delays and severe behavioral disorders. All three children experienced some degree of visual impairment. The missense variants provided new insights into the neurodevelopmental consequences of compromised PCDH12 function by distinguishing the specific consequences associated with dysfunction in the extracellular versus intracellular domains of PCDH12. All identified missense variants are predicted to be deleterious and destabilizing. The expression of PCDH12 in HEK293T and HeLa cells demonstrated that PCDH12 is expressed effectively, regardless of the presence of missense variants. However, the extracellular variants p.Val581Gly and p.Ile621His compromised the stability of PCDH12s homophilic adhesion. Additionally, we found evidence of an interaction between PCDH12 and the extracellular domain of the epilepsy-associated PCDH19 protein. PCDH12 extracellular missense variants also affect PCDH19 stability. Our study provides evidence that PCDH12 mediates both homophilic and heterophilic interactions. Our findings also highlight the importance of stable PCDH12-mediated adhesion, emphasizing the need to further study the functional consequences of PCDH12 missense variants on brain and visual system development.

AimBesides registries, healthcare databases can provide useful information for assessing major congenital malformations (MCMs) frequency and investigating their risk factors, particularly medications exposures. This study aimed to assess the validity of MCMs identification based on French national, comprehensive healthcare databases. MethodsUsing information on hospital discharge diagnoses, medical procedures (e.g. surgery) and death causes from the EPI-MERES register nested in the French National Health Data System, 72 specific MCMs grouped in 11 organ groups were assessed among all births occurred after 22 weeks of amenorrhea in France between 2010 and 2023. MCMs prevalence rates were estimated and compared to those from EUROCAT, and associations with prenatal exposure to valproate were assessed. ResultsAmong 10.5 million births, 213,153 live born infants with at least one MCM, i.e. 203.0 cases per 10,000 births, were identified. MCMs prevalence rates among live births were close to those reported in EUROCAT overall (difference: -1.76 per 10,000 births [-1%]), for each organ group (differences ranging from -9.10 [-13%] to +3.44 [+16%] per 10,000 births), and for the 72 specific MCMs (median prevalence difference: 1%). Prenatal exposure to valproate was significantly associated with increased risks of any MCM (adjusted odds ratio (aOR) 2.82, 95% CI [2.33-3.41]) and of 15 specific MCMs including spina bifida (aOR 17.88 [7.88-40.53]). ConclusionThis study supports the validity of MCMs identification based on data of the EPI-MERES register. The EPI-MERES register provides a highly powerful, reactive and operational tool complementing MCMs registries for improving real-life knowledge on drug teratogenicity. Plain language summaryMajor congenital malformations are serious structural abnormalities present at birth that can have lasting consequences on childrens health. Better understanding their risk factors, particularly medication exposures during pregnancy, is crucial. Population-based registries are today the primary source of information on malformations, but healthcare databases could offer a faster and broader alternative. This study tested whether the EPI-MERES register, built upon the French National Health Data System (SNDS), could accurately identify 72 specific malformations across 10.5 million births between 2010 and 2023. Prevalence estimates closely matched those from the European EUROCAT registry, confirming good data accuracy. As expected, valproate (a known teratogen) was associated with an increased risk of various malformations, including spina bifida, EPI-MERES thus constitutes a promising tool for studying medication risks during pregnancy.

STUDY QUESTION[Do structural genomic variants, that can be identified by using optical genome mapping, contribute to male infertility?] SUMMARY ANSWER[By using optical genome mapping we can identify several types of structural variants, both known and new, that may contribute to male infertility.] WHAT IS KNOWN ALREADY[Traditional approaches such as karyotyping, CFTR and chromosome Y microdeletion testing are successful in explaining clinical findings in [~]30% of MI patients, leaving the rest without a genetic diagnosis. Recent research suggests at least 265 genes may play a role in male fertility. While the assessment of the roles of copy number variants and single nucleotide variants in monogenic forms of disease in these genes is underway, much less is known about structural variants.] STUDY DESIGN, SIZE, DURATION[We performed a longitudinal case/control study on a total of 220 individuals; 88 patients with male infertility, negative for cytogenetic abnormalities using karyotyping, and molecular testing for chrY microdeletions, and CFTR gene variants, and 132 healthy male individuals that underwent optical genomic mapping for other reasons. Exclusion criteria for the control cohort were low-sperm quality and/or inclusion in IVF procedures. The study was approved by the National Medical Ethics Committee of the Republic of Slovenia (reference number: 0120-213/2022/6). Optical genome mapping was performed from an aliquot of whole blood collected for routine testing purposes at the Clinical Institute of Genomic Medicine (CIGM), UMC Ljubljana from January 2023 to November 2024.] PARTICIPANTS/MATERIALS, SETTING, METHODS[We examined structural variants in 220 participants by using optical genome mapping, which was performed with DLE-1 SP-G2 chemistry and the Saphyr instrument. The de novo assembly and Variant Annotation Pipeline were executed on Bionano Solve3.7_20221013_25 while reporting and direct visualization of structural variants was done on Bionano Access 1.7.2. All obtained variants were filtered using the Bionano Access software and in-house generated gene/regions of interest panel bed files. The first filter was applied to include variants below a population frequency of 10%, and overlapping the regions of interest. Subsequently, all variants occurring with frequency 0% in the internal manufacturer variant dataset were manually evaluated for possible involvement of the overlapping genes or regions in biological processes involved in MI. The male infertility cohort also underwent research whole exome analyses as previously reported. All results of optical genomic mapping were confirmed by an appropriate alternative method where available.] MAIN RESULTS AND THE ROLE OF CHANCE[We show that the overall number of structural variants in MI patients does not differ from that of healthy individuals. By looking in detail at genes and regions associated with MI, we identified 21 rare variants absent from controls in 25.0 % of MI patients, of which five were likely causative, and two would be missed by using traditional approaches. These variants include inversions, duplications, amplifications, deletions (e.g. SPAG1), and insertions/expansions (e.g. DMPK), that were validated using additional methods. While the remaining SV cannot be currently classified as pathogenic according to existing criteria, they open a new avenue in genetic research of MI. LARGE SCALE DATA[Variants reported in this study were deposited into ClinVar under accession numbers SUB15650956 (https://www.ncbi.nlm.nih.gov/clinvar/)] LIMITATIONS, REASONS FOR CAUTION[Technical limitations of optical genome mapping include the lack of DLE-1 labelling of centromeric and telomeric regions, the inability to detect Robertsonian translocations, the unclear exact location of smaller structural variants located between the DLE-1 labels, and unclear boundaries in case of their location in segmentally duplicated regions (this limitation is shared with other methods). The ACGM criteria of rarity are also hard to apply, as the fertility status of the individuals in healthy population databases such as GnomAD and DGV is unknown. Similarly, gene-associated phenotype and the proposed inheritance model both need to be considered as parts of the ACMG criteria, but for many candidate genes associated with MI, no model of inheritance has yet been proposed.] WIDER IMPLICATIONS OF THE FINDINGS[Currently, with the established diagnostic approaches we are able to resolve [~]30% of male infertility cases, with [~]70% of patients remaining undiagnosed. The significance of our work is in showing that rare structural variants can be identified in MI, by using optical genome mapping, opening new avenues of research of the genetics of this important contributor to human fertility.] STUDY FUNDING/COMPETING INTEREST(S)[All authors declare having no conflict of interest in regard to this research. This work was funded by the Slovenian Research and Innovation Agency (ARIS) Programme grant P3-0326: Gynecology and Reproduction: Genomics for personalized medicine] Lay summaryMale infertility affects about 5% of adult males and has complex causes, including genetic ones, such as mutations in the CFTR gene, small deletions on chromosome Y, and balanced translocations, but currently we can only find a genetic cause in [~]30% of patients. This means [~]70% of cases remain undiagnosed but potentially, they too may have a yet unknown genetic cause. Indeed, so far research has shown at least 265 genes have been proposed to play a role in male fertility. In these genes, there has so far been limited research of single nucleotide variants and of copy number variants, but many structural variants are not visible using commonly used methods in clinical genetic testing. Therefore, apart from chromosome Y microdeletions and chromosomal numerical and structural anomalies, such as balanced translocations, the role of smaller structural variants in male infertility is unknown, but based from what we know from other diseases, they also may play a role in male infertility. Optical genome mapping is a novel method for the detection of structural variants, such as balanced and unbalanced translocations, insertions, duplications, deletions, and complex structural rearrangements in a wide range of sizes. By using optical genome mapping to test a cohort of 88 infertile men and 132 healthy controls, we aimed to provide the first insights into the range of SV that may be associated with MI. We found, by using optical genome mapping, the overall number of structural variants in MI patients not to be significantly different to the control group. However, by looking at genes and regions associated with MI, we can find rare structural variants that are absent from controls in 25.0% of MI patients. These variants include inversions, duplications, amplifications, deletions (e.g. deletion in SPAG1), and insertions/expansions (e.g. in DMPK), that were validated using additional methods. Five of these variants (5.6%) were likely causative, and two would be missed by traditional approaches. While the remaining SV cannot be currently classified as pathogenic according to existing criteria, they open a new avenue in genetic research of MI.

Abstract Background Understanding the phenotypic spectrum of disease-associated genes is essential for accurate diagnosis and targeted therapy. FRMPD4 (FERM and PDZ Domain Containing 4) has previously been associated with intellectual disability and epilepsy. However, its potential role in non-syndromic hearing loss has not been explored. Methods We performed genetic analysis in two unrelated families presenting with non-syndromic sensorineural hearing loss, identifying maternally inherited missense variants in FRMPD4. Clinical phenotyping included audiological assessment and evaluation for neurodevelopmental involvement. Cross-species expression analyses were conducted in Drosophila, zebrafish, and mouse. Functional characterization included quantitative evaluation of sound-evoked responses in Drosophila nicht gut hoerend (ngh) mutants, assessment of neuronal development and acoustic startle responses in zebrafish loss of function models, and morphological cochlear analyses with auditory brainstem response measurements in knockout mice. Results Three affected males from two unrelated families presented with prelingual, bilaterally symmetrical sensorineural hearing loss, with confirmed congenital onset in one individual and no evidence of neurodevelopmental abnormalities. Cross-species analyses demonstrated evolutionarily conserved expression of FRMPD4 in auditory structures. In Drosophila, quantitative analysis of sound-evoked responses in ngh mutants revealed impaired auditory function. Zebrafish loss of function models exhibited reduced neuronal populations in the otic vesicle and posterior lateral line, abnormal neuromast development, and diminished acoustic startle responses. In mice, Frmpd4 knockout resulted in high-frequency hearing loss and cochlear abnormalities consistent with the human phenotype. Conclusions Our findings expand the phenotypic spectrum of FRMPD4 to include non-syndromic sensorineural hearing loss and establish its evolutionarily conserved role in auditory function. These results have direct implications for genetic diagnosis and variant interpretation in patients with hearing loss.

Copy number variants (CNV) contribute significantly to the pathogenic variation associated with developmental disorders. CNV detection is often not included in standard exome sequencing (ES) analysis. Complementary methods such as chromosomal microarray are typically offered in diagnostic laboratories to diagnose pathogenic CNV. In this study, we aimed to develop an optimal approach for incorporating CNV detection within our ES analysis process for the Deciphering Developmental Disorders in Africa (DDD-Africa) cohort. We analyzed ES data from 505 probands with a developmental disorder, applying a CNV detection approach that assessed data generated using the tools CANOES and XHMM. When available, parental ES data was used to assess inheritance patterns. We confirmed a diagnosis in 42/505 (8,3%) patients with 44 pathogenic CNV identified in the probands. There were 31 deletions and 13 duplications. Among the 27 probands with parental data, all identified CNV were de novo. The addition of CNV analysis to our ES analysis pipeline resulted in an 8.3% increase in diagnostic yield in the DDD-Africa cohort without additional laboratory cost. This approach offers a feasible approach which is likely to reduce analytical cost and is suitable for low- and middle-income countries where funding and resources for genomic medicine initiatives are limited.

Primary open-angle glaucoma (POAG) disproportionately affects individuals of African ancestry, yet rare coding variation in this population remains understudied. To address this gap, we performed a multi-cohort exome-wide meta-analysis across POAAGG, PMBB, All of Us, and UK Biobank, including 4,815 POAG cases and 22,922 controls of genetically inferred African ancestry. Although no gene reached exome-wide significance, we identified several suggestive gene-level associations driven by rare variants (minor allele frequency [&le;]0.1% or singletons),including signals in SRF, BLTP3A, METTL2A, and KRT10. Among these, SRF demonstrated the strongest association and was driven by rare missense variants with moderate effect sizes. Given its role in cytoskeletal organization and actin dynamics; processes central to trabecular meshwork function and intraocular pressure regulation SRF represents a biologically plausible candidate gene. Notably, these genes have not been previously highlighted in predominantly European ancestry POAG association studies, suggesting potential ancestry-specific rare variant contributions. Overall, our findings highlight the critical importance of investigating rare coding variation in POAG, in disproportionately affected populations to deepen understanding of POAG etiology and genetic risk.

RationaleNeurodevelopmental disorders (NDDs) are characterised by significant challenges in communication, social interaction, and adaptive function, often impacting quality of life. Previous studies support genetic influences on the communication abilities of individuals with NDD, but were either limited to single genetic conditions or to small cohorts with a limited selection of communication measures. MethodsWe analysed caregiver-reported communication abilities in 79,518 individuals with NDD from the Simons Searchlight and SPARK registries: 4,439 with a CNV-based or monogenic NDD and 75,079 with autism spectrum disorder (ASD) without a known genetic cause (idiopathic ASD) as controls. For analysis, we a priori selected 10 communication-related measures based on their availability in the study cohorts, coverage of distinct communication aspects, and their frequent use in neurodevelopmental phenotyping, yielding 177,328 data points across all study cohorts. The individuals in the Searchlight registry were divided into a Discovery cohort (the 15 most prevalent genetic NDD conditions) and a Confirmation cohort (all other genetic NDD conditions). A second Confirmation cohort was generated using all individuals with genetic ASD forms from the SPARK registry. We then tested each of the three case cohorts and each genetic condition represented in the Discovery cohort against the ASD control cohort. Developmental trajectories were assessed through testing of participants grouped by age at evaluation. ResultsMeasure-level analyses demonstrated significant associations between genetic status and communication abilities, differences in communication abilities between classes of genetic variants (monogenic vs. CNV-based NDDs), and variability between specific genetic NDD conditions. CNV-based NDDs showed milder communication impairment, outperforming idiopathic ASD controls in 9/10 communication measures, whereas monogenic NDD conditions had more pervasive impairments, especially in verbal communication. Although impaired in verbal communication, five monogenic NDD conditions showed at least suggestive strengths in nonverbal and social communication relative to idiopathic ASD controls (CSNK2A1, CTNNB1, SETBP1, MED13L, and PPP2R5D), specifically in using gestures. Developmental trajectory analyses revealed STXBP1 as the gene group at highest risk of developmental stagnation in communication abilities. ConclusionsThese findings underscore the potential of precision speech-language pathology (SLP) approaches tailored to the specific verbal and nonverbal communication strengths and weaknesses of genetic groups. We also provide evidence for measurable improvements and declines in communication abilities with age at the group level, highlighting the need for developmentally informed care. By integrating genetic insights into clinical practice, precision SLP approaches may enhance communication outcomes and developmental progress and improve quality of life for individuals with genetic NDDs.