Genetics
Abstract
BACKGROUND. Primary adrenal insufficiency (PAI) is commonly caused by autoimmunity in adults, known as autoimmune Addison’s disease (AAD). Autoantibodies against 21-hydroxylase are a hallmark of AAD but are absent in some patients diagnosed with AAD, posing a challenge to their clinical management. METHODS. We tested a polygenic risk score for AAD (PRS14AAD) to prioritize seronegative patients for etiologic reevaluation. The PRS14AAD was calculated in patients diagnosed with AAD but lacking 21-hydroxylase autoantibodies (n = 124). Patients with low (n = 35) or high (n = 10) genetic susceptibility to AAD underwent clinical re-evaluation and whole-genome sequencing. RESULTS. Of the 34 patients with low PRS14AAD, 14 (41%) were re-diagnosed with non-autoimmune adrenal insufficiency, whereas no alternative etiologies were identified in patients with high PRS14AAD. The low PRS group included six patients (17%) with confirmed monogenic PAI and eight patients (23%) with adrenal insufficiency due to either tuberculosis or non-primary causes. Notably, four of the six confirmed monogenic cases developed PAI in adulthood, consistent with late-onset disease caused by hypomorphic variants. Additionally, seven patients (20%) carried genetic variants suspected of causing monogenic disease. CONCLUSION. These findings support considering genetic evaluation in patients with seronegative PAI, regardless of the age at diagnosis.
Authors
Maribel Aranda-Guillén, Ileana Ruxandra Botusan, Venuja Fernando, Michela Barbaro, Maria Blomqvist, Ellen Christine Røyrvik, Anette Susanne Bøe Wolff, Eystein Sverre Husebye, Stephan Kemp, Anna Wedell, Olov Ekwall, Magnus Isaksson, Jeanette Wahlberg, Ola Lindgren, Jakob Skov, Ragnhildur Bergthorsdottir, Per Dahlqvist, Sophie Bensing, Olle Kämpe, Daniel Eriksson
Abstract
SEC16A encodes a scaffold protein that organizes the endoplasmic reticulum (ER) exit sites, specialized ER subdomains that mediate export of newly synthesized proteins and regulate early secretory pathway steps. The role of SEC16A in human nervous system diseases has not been established yet. Here, we report six individuals with bi-allelic SEC16A variants who presented with early-onset neurological symptoms, including cognitive impairment, motor delays, and seizures. Using Drosophila, we show the Sec16 ortholog is essential for normal development and that its loss causes early lethality. Neuron-specific knockdown of Sec16 causes neurological phenotypes including locomotor defects, seizure-like bang sensitivity, and impaired synaptic transmission, which can be associated with the clinical features observed in affected individuals. Neurons differentiated from patient-derived induced pluripotent stem cells (iPSCs) carrying SEC16A variants display reduced SEC16A protein, impaired neurite outgrowth and increased excitability, which are rescued in isogenic control neurons when the variants are corrected, consistent with a loss-of-function mechanism. Increased ER stress was observed in both patient iPSC-derived neurons and fly models. In summary, our findings indicate a conserved requirement for SEC16A in neurodevelopment and neuronal function and implicate ER stress as a key cellular phenotype underlying the SEC16A-associated neurodevelopmental disorder.
Authors
Mengqi Ma, Xiaowei Lei, Juan Li, Yiming Zheng, Yu Yuan, Yohei Nitta, Xiao Mao, Zhenming Guo, Shenzhao Lu, Mingxi Deng, Xueyang Pan, Oguz Kanca, Shiyi Liu, Hua Wang, Bo Xiao, Rachel Schot, Serwet Demirdas, Yolande van Bever, Marjon van Slegtenhorst, Tahsin Stefan Barakat, Shan Bian, Hugo J. Bellen, Lili Long
Abstract
Triple negaJve breast cancer (TNBC) is characterised by poor prognosis and limited treatment opJons, parJally due to therapy-resistant stem/progenitor cells driving metastasis. Elevated expression of the transcripJon factor MYBL2 is associated with aggressive disease, yet its role in BC stem/progenitor cells remains elusive. Here, we employed inducible MYBL2 downregulaJon in TNBC cell lines and paJent-derived models to invesJgate its funcJon and idenJfy potenJal vulnerabiliJes associated with its high expression. Our funcJonal and mechanisJc analyses reveal that high MYBL2 levels are criJcal for stem cell self-renewal, DNA repair, and replicaJon stress response and lowering MYBL2 impairs stemness and self-renewal both in vitro and in vivo. Accordingly, high-MYBL2 stem/progenitor cells exhibit increased sensiJvity to Polθ inhibitors which is lost upon MYBL2 downregulaJon due to transcripJonal suppression. Combining Polθ inhibitors with ATR inhibitors further enhances this sensiJvity, in vitro, ex vivo and in vivo. These findings suggest that dual Polθ/ATR inhibiJon represents a promising therapeuJc strategy to target TNBC stem/progenitor cells with high MYBL2 expression, highlighJng MYBL2 as a potenJal biomarker to straJfy paJents for Polθ inhibitor-based therapies in TNBC.
Authors
Rachel Bayley, Xiaoping Qu, Adrija Chakraborty, Menna Shamma, Ciara Ward, Kinga Koscielniak, Anna Munsey, Suman Ahmed, Amber Stiby, Bohdana-Myroslava Briantseva, Kirendeep Jawanda, Christopher J. Lord, Abeer M. Shaaban, Marco Saponaro, Graeme C. M. Smith, Robert B. Clarke, Clare Davies, Paloma Garcia
Abstract
Hypoxia-inducible factor 1 (HIF-1) orchestrates the transcriptional regulation of thousands of genes involved in breast cancer (BC) progression. Here, we identified protein phosphatase 2A (PP2A) methylesterase 1 (PPME1) as a critical HIF-1 target gene that drives oncogenic signaling under hypoxic conditions. In BC cells, HIF-1–dependent PPME1 expression caused inhibition of the PP2A catalytic subunit (PP2Ac), thereby diminishing PP2A activity, which led to AKT activation, phosphorylation of β-catenin, and its nuclear translocation. Nuclear β-catenin cooperates with HIF-1 to promote BC stem cell specification by activating transcription of the NANOG and KLF4 genes, which encode pluripotency factors, and to drive immune evasion by activating transcription of VEGFA, which recruits and polarizes immunosuppressive tumor-associated macrophages and ISG20, which represses STAT1/IRF1-dependent expression of CXCL10, thereby impairing CD8+ T cell recruitment. In vivo, PPME1 knockdown altered the tumor immune microenvironment, enhanced antitumor immunity, and synergized with anti–CTLA-4 immunotherapy to enable complete tumor eradication. These findings establish PPME1 as a critical regulator linking hypoxia signaling, stemness, and immune evasion and highlight its potential as a BC therapeutic target in combination with immune checkpoint blockade.
Authors
Yajing Lyu, Varen Talwar, Yongkang Yang, Si-Sim Kang, Shuyi Li, Shaima Salman, Daiana Drehmer, Yufeng Wang, Chelsey Chen, Vijay Ramu, Sujin Kim, Dylan Park, Tina Yi-Ting Huang, Emmanuel Datan, Dominic Dordai, Jonathan P. Schneck, Gregg L. Semenza
Abstract
Vascular endothelial cells (ECs) encode a homeostatic fluid shear stress (FSS) set point that is essential for vascular stability. Deviations above or below this threshold trigger adaptive remodeling to restore physiological shear levels. Disruption of this control mechanism leads to enlarged arteriovenous malformations (AVMs) in hereditary hemorrhagic telangiectasia, a vascular disorder caused by heterozygous loss-of-function (LOF) mutation in ALK1, ENG, or SMAD4. Mechanistically, Smad4-deficient ECs are reset to a lower FSS set point value, resulting in AVMs that show characteristics of high-FSS remodeling with elevated KLF4 and high activation of the downstream Akt. Here, we investigated the KLF4/Akt upstream mechanisms by which SMAD4 sets the physiological FSS set point. We identified the receptor tyrosine kinase c-KIT as a component and regulator of the junctional mechanosensory receptor complex, which is highly upregulated in murine and human AVMs. SMAD4 restrains flow signaling by limiting c-KIT–dependent ERK5 activation and KLF4 induction. Thus, SMAD4 LOF leads to sustained c-KIT engagement in the sensory junctional apparatus, driving enhanced and prolonged activation of the ERK5/KLF4/Akt signaling axis. These results show that Smad4-LOF mutations induce malformations by disabling a key homeostatic mechanism and identify c-KIT as a potentially previously unrecognized therapeutic target.
Authors
Johannes Gahn, Fan Wu, Qing Zhang, Yuxi Di, Tanmaya Behera, Yonggang Ren, Zohrah Hashemi, Kuheli Banerjee, Julio Cordero, Claudia Gherman, Kornelia Andorfer, Caroline T. Seebauer, Fatemeh Mirzapour-Shafiyi, Gergana Dobreva, Martin A. Schwartz, Roxana Ola
Abstract
Familial Episodic Pain Syndrome (FEPS) is a rare inherited disorder characterized by recurrent episodes of severe upper-body pain triggered by stimuli such as cold, stress, or fasting. A gain-of-function mutation (N855S) in the Transient Receptor Potential Ankyrin 1 (TRPA1) ion channel has been identified in affected individuals, yet the cellular mechanisms driving FEPS pain remain poorly defined. Although TRPA1 has been primarily studied in sensory neurons, emerging evidence suggests that non-neuronal cells may contribute to pain signaling. To define the cell-specific role of mutant TRPA1 (TRPA1mut), we generated mouse models selectively expressing TRPA1mut in Schwann cells or sensory neurons, using CRISPR-based and Cre-loxP strategies. Patch-clamp recordings confirmed enhanced agonist-evoked currents in TRPA1mut compared with wild-type channels. Behavioral analyses revealed that neuronal TRPA1mut mediates acute nociceptive responses, whereas Schwann cell TRPA1mut drives mechanical allodynia induced by subthreshold TRPA1 agonists and physiological triggers relevant to FEPS, including fasting, cold exposure, and restraint stress. These responses were associated with increased reactive oxygen species and accumulation of 4-hydroxynonenal in sciatic nerves and were attenuated by antioxidant treatment. These findings uncover a previously unrecognized Schwann cell-dependent mechanism of TRPA1-driven pain and identify non-neuronal TRPA1 signaling as a potential therapeutic target in FEPS and related pain channelopathies.
Authors
Matilde Marini, Martina Chieca, Elisabetta Coppi, Marco Montini, Lorenzo Bonacchi, Lorenzo Landini, Irene Scuffi, Kelvin Y. Kwan, Alice Papini, Gaetano De Siena, Elisa Bellantoni, Lucrezia Timotei, Elena Colzi, Valentina Albanese, Gabriele Ferroni, Evelynn D. N. Melo, Marie-Christine Birling, Romain Lorentz, Romina Nassini, Francesco De Logu
Abstract
Background. Myelodysplastic syndromes (MDS) are characterized by aberrant DNA methylation, and mutations in epigenetic modifiers are frequently found in these patients. Although DNA methyltransferase inhibitors (DNMTi) are used to treat MDS, response variability remains a challenge in the clinic, with limited predictive markers. Methods. We integrated genomic, epigenomic, and transcriptomic analyses of 98 MDS patients. Patients were classified into epigenetic subtypes via hierarchical clustering. Random forest classifiers were developed and validated using internal stratified testing and an independent external cohort to predict AZA response. Results. MDS is characterized by widespread DNA hypomethylation affecting distal regulatory elements. We identified seven epigenetic clusters correlated with distinct molecular drivers. Notably, Cluster VI exhibited low mutational burden but a high AZA response rate of 71% (P ≤ 0.01). While transcriptional profiles alone failed to distinguish responders, a DNAme-based classifier achieved an area under the curve (AUC) of 0.82. An integrative model combining DNAme, gene expression, mutations, and clinical parameters achieved an AUC of 0.93 in internal validation and 0.88 in the external cohort. Conclusion. Epigenetic signatures at distal genomic elements provide superior predictive power for AZA response compared to promoter-centric or transcriptional analyses. These findings establish a robust framework for personalized treatment strategies in MDS.
Authors
Qin Yang, Miguel Torres-Martin, Masataka Taguchi, Alice Brogi, Irene Casalin, Eleonora Ceneri, Stephanie Halene, Amy E. DeZern, Elizabeth A. Griffiths, Matilde Y. Follo, Carlo Finelli, Jerald P. Radich, Michael J. Rauh, Rafael Bejar, Mikkael A. Sekeres, Valeria Santini, Maria E. Figueroa
Abstract
Despite enormous advances in clinical genomics, idiopathic scoliosis remains an enigmatic condition with poorly understood genetic and pathophysiological underpinnings impeding molecular diagnosis and the development of targeted treatments. We performed linkage analysis, exome, genome and short- and long-read RNA sequencing in a multigenerational family affected by autosomal dominant early-onset scoliosis (EOS) with a unique pattern of spondylodysplastic elements and progressive endplate erosion and identified the LRR binding FLII interacting protein 1 gene (LRRFIP1) as the disease-causing gene. The underlying cause is a rare noncoding variant altering transcription factor binding of NR3C1 (glucocorticoid receptor) leading to changes in LRRFIP1-isoform expression. Transcriptomic changes in fibroblasts of affected individuals indicated a combination of disturbed Wnt-signaling during somitogenesis, planar cell polarity signaling and postnatal inflammatory dysregulation with clinical and molecular overlaps with Ankylosing Spondylitis and Scheuermann kyphosis. We conducted a rare variant enrichment analysis using genome data from 484,903 UK Biobank participants and found an enrichment of rare risk-increasing LRRFIP1-variants in individuals with scoliosis. Our analysis of an Lrrfip1tm1.1(KOMP)Wtsi KO mouse model showed increased prevalence of idiopathic kyphoscoliosis in Lrrfip1-deficient mice. Our work provides insights into the pathophysiology of rare and common spinal disorders and hints to potential future therapeutic approaches with selective NR3C1-inhibitors.
Authors
Tanja Frey, Elena M. Cabello, Gabriele Siegel, Carla Bello, Rike Schiller, Martina A. Trippel, Neguin Ranjbar, Paranchai Boonsawat, Stephanie E. van Gijn, Ivan Ivanovski, Michael Papik, Markus Zweier, Kan Min, Katharina Steindl, Anita Rauch
Abstract
Atrial fibrillation (AF) is the most common sustained human cardiac arrhythmia and linked to a drastic increase in stroke and heart failure risk. While sequence variations in the PITX2 non-coding region are the strongest genetic signature of AF risk, the direct role of PITX2 in AF remains a topic of debate. Here, we generated a mouse model (Pitx2Pro41Ser) of a human PITX2 coding variant linked to increased AF risk in the Finnish population. The Pitx2Pro41Ser mice exhibit near-complete penetrance of pacing-induced AF, and transcriptional profiling indicates that Pitx2Pro41Ser is a loss-of-function mutation. In vivo cleavage under targets and tagmentation (CUT&Tag) reveals that PITX2 acts as a transcriptional repressor in developing left atrial cardiomyocytes independent of DNA methylation. Ectopic Pitx2 expression in postnatal right atrial cardiomyocytes via adeno-associated virus (AAV) delivery or genetic overexpression represses right atrial genes and induces a left atrial transcriptome, revealing unexpected plasticity of postnatal atrial cardiomyocytes. Strikingly, delivery of Pitx2 AAV into Pitx2Pro41Ser mice rescues AF inducibility uncovering a direct link between PITX2 activity and AF susceptibility.
Authors
Jeffrey D. Steimle, Yue Yuan, Shaohai Fang, Vaibhav Deshmukh, Christine Rodriguez, Fansen Meng, Taotao Tan, Md. Abul Hassan Samee, Yun Huang, Na Li, James F. Martin
Abstract
STXBP1 variants are a frequent cause of early-onset developmental and epileptic encephalopathies and related neurodevelopmental disorders, but the clinical interpretation of these variants remains a major challenge. Most reported STXBP1 missense variants are classified as variants of uncertain significance (VUS), complicating diagnosis, counseling, and patient eligibility for precision therapies. Here, we developed EpiPred, a gene-specific machine learning classifier that predicts the pathogenicity of STXBP1 missense variants and tests these predictions using empirical evidence from well-established cellular assays. Trained on a curated set of pathogenic and benign variants, EpiPred outperformed global prediction tools in accuracy, sensitivity, and specificity. We validated the model’s predictions using variant effect assays that measure protein abundance, solubility, stability, and interaction with the SNARE complex partner syntaxin 1. These biochemical readouts aligned closely with model outputs and enabled reclassification of several possibly misdiagnosed variants, which warrant further validation and clinical reevaluation. We deployed EpiPred in an interactive web application that allows clinicians, researchers, and patients to explore predictions for all possible STXBP1 missense variants. By identifying likely pathogenic STXBP1 variants, including those that may respond to emerging therapies such as protein stabilizers. By coupling gene-calibrated machine learning with orthogonal variant-effect assays and public deployment, EpiPred provides a transferable framework for VUS resolution, trial enrichment, and precision diagnosis across clinically actionable Mendelian disease genes.
Authors
Jeffrey D. Calhoun, Chengbing Wang, Carina G. Biar, Jonathan R. Gunti, John S. Lee, Aaron M. Geller, Jung H. Hong, Santiago Schnell, Louis T. Dang, Yu Wang, Jack M. Parent, Lori L. Isom, Michael D. Uhler, Heather C. Mefford, M. Elizabeth Ross, Vanessa Aguiar-Pulido, Gemma L. Carvill
Copyright © 2026 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)