Neurofibromin 1 (NF1) is a tumor suppressor gene frequently altered across diverse cancer types, yet its biological significance in ovarian cancer remains incompletely characterized. Here, we integrated cohort‐scale somatic mutation profiling with functional validation to characterize the mutational and cellular consequences of NF1 loss in ovarian cancer. Somatic mutation data from the TCGA ovarian cancer cohort were analyzed to define NF1‐associated mutation types, tumor mutational burden, mutational signatures, and co‐occurring alterations in DNA damage repair (DDR) pathways, together with pathway‐ and gene set–level functional enrichment analyses. NF1 alterations were predominantly truncating and consistent with loss‐of‐function events. NF1‐mutant tumors did not exhibit increased global tumor mutational burden or uniform APOBEC hypermutation but showed distinct single‐nucleotide substitution patterns and frequent comutations in core DDR‐related genes. Functional enrichment analyses further highlighted coordinated involvement of pathways related to DNA replication, RNA processing, and proteostasis. Clinically, NF1 mutation was not independently associated with overall survival. Stable NF1 knockdown ovarian cancer models showed that NF1 depletion did not affect basal proliferation but increased sensitivity to hydroxyurea‐induced replication stress, accompanied by increased γH2AX accumulation. Together, these findings indicate that NF1 loss defines a DNA damage–associated mutational and cellular state in ovarian cancer. Rather than acting as a direct prognostic determinant, NF1 mutation appears to increase vulnerability to replication stress and DNA damage, providing functional insight into its role in ovarian tumor biology.

Constitutional epimutations of the MLH1 gene are an alternative cause of Lynch syndrome, in which inactivation of an allele of a mismatch repair (MMR) gene results from MLH1 promoter methylation, rather than a pathogenic genetic variant. These epimutations are often mosaic, and methylation levels ranging from ~50% monoallelic methylation to low‐level methylation (1%–5%) are observed in the blood of MLH1 epimutation carriers. Using a specific and highly sensitive droplet digital methyl‐specific PCR (ddMSP) assay, six patients with very low methylation levels (< 1%) were identified in a series of 142 patients with a MLH1‐methylated tumor diagnosed before age 61, who had been referred to the clinical lab between 2020 and 2024. These patients were initially missed by standard pyrosequencing assay, emphasizing the need for highly sensitive assays for constitutional epimutation screening. To confirm that methylated DNA molecules detected by ddMSP did not correspond to circulating tumor DNA rather than germline DNA, multiple validation analyses were performed, including validation of the constitutional origin of methylation on other sources of germline DNA and tumoral analysis. Taking into account the other patients identified as epimutation carriers by pyrosequencing during the same 5‐year period, 13.1% of patients with a MLH1‐methylated tumor before age 61 were diagnosed as Lynch syndrome patients, which changed their clinical follow‐up. These findings highlight the relevance of recommendations for systematic MLH1 epimutation screening using highly sensitive assays in patients with MLH1‐methylated tumors diagnosed before age 61. Such screening will increase the number of patients diagnosed with Lynch syndrome caused by a MLH1 constitutional epimutation, improving patient care and outcomes, as well as genetic counseling.

ObjectiveTo investigate the protective effects of Buyang Huanwu decoction (BYHW) on myocardial injury induced by chronic intermittent hypoxia (CIH) in rats and to analyze its potential regulatory mechanism through the NF‐κB/LOX signaling pathway.MethodsThen, 36 adult male Sprague–Dawley (SD) rats (weighing 200–250 g) were randomly divided into six groups (n = 6): normal control (NC), CIH model, BYHW, BYHW + lipopolysaccharide (LPS), BYHW + pyrrolidinedithiocarbamate (PDTC), and LPS. Except for the NC group, all groups underwent 5 weeks of intermittent hypoxia (8 h/day) alongside their respective drug treatments. Postintervention, systolic blood pressure and heart rate were recorded. Cardiac function was evaluated by echocardiography to measure left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), and left ventricular internal dimension at end‐diastole (LVDd) and left ventricular internal dimension at end‐systole (LVDs). Myocardial sections were analyzed by HE and Sirius Red staining for quantitative assessment of inflammatory cell infiltration and collagen volume fraction (CVF). The protein and mRNA expression levels of NF‐κB, LOX, Collagen I, and Collagen III in cardiac tissue were analyzed by Western blot and qPCR.ResultsCompared with the NC group, rats exposed to CIH or LPS showed elevated blood pressure and an increased heart rate, along with impaired cardiac function, as evidenced by reduced LVEF (p < 0.05) and LVFS (p < 0.05), along with increased LVDd (p < 0.05) and LVDs (p < 0.05). BYHW treatment significantly ameliorated the CIH‐induced cardiac dysfunction and ventricular dilation (p < 0.05), and these improvements were further enhanced by cotreatment with BYHW and the NF‐κB inhibitor PDTC (p < 0.05). Conversely, cotreatment with BYHW and the NF‐κB agonist LPS attenuated the cardioprotective effects of BYHW (p < 0.05). Histologically, BYHW significantly reduced CIH‐induced myocardial structural disruption, inflammatory cell infiltration, and fibrosis (CVF) (p < 0.05), effects that were potentiated by cotreatment with BYHW and PDTC, and counteracted by cotreatment with BYHW and LPS. At the molecular level, Western blot and qPCR analyses revealed that BYHW significantly reduced both the protein and mRNA levels of NF‐κB, LOX, Collagen I, and Collagen III (p < 0.05) compared with the NC group. This suppression was further enhanced by cotreatment with BYHW and PDTC and attenuated by cotreatment with BYHW and LPS.ConclusionBYHW improves cardiac function and reduces myocardial inflammation and fibrosis in SD rats with CIH. This effect may be related to the inhibition of the NF‐κB/LOX signaling pathway. Cotreatment with BYHW and PDTC enhances this therapeutic effect, while cotreatment with BYHW and LPS attenuates this effect. This study provides preliminary experimental evidence for the cardioprotective effects of BYHW.

ObjectiveThe aim of this study is to evaluate the efficacy of accurate circular consensus long‐read sequencing in the detection of rare thalassemia.MethodsConventional molecular analysis on globin genes has limitations because of the broad spectrum of genetic variants, complex genetics, and genotype–phenotype correlation. Accurate circular consensus long‐read sequencing is a novel tool that detects complex variants in the thalassemia gene based on third‐generation sequencing. In this study, we screen out suspected rare thalassemia carriers by hemoglobin analysis and conventional molecular analysis, and evaluate the efficacy of accurate circular consensus long‐read sequencing in the detection of rare thalassemia.ResultsBased on the traditional screening of thalassemia gene, an additional 16 (17.67%) cases of clinically significant variants of rare thalassemia were identified by accurate circular consensus long‐read sequencing in this study, including 12‐point variants and 4 deletion variants: HBB: (SEA)‐HPFH, HBB: c.268_281delAGTGAGCTGCACTG, HBB: (Chinese) Gγ + (Aγδβ)0, and HBA2:c.91‐93delGAG.ConclusionAccurate circular consensus long‐read sequencing has a promising prospect in detecting rare thalassemia gene variants and may improve the detection rate of carriers.

The genetic architecture underlying traits associated with Type 1 Gaucher disease (GD1) remains insufficiently explored. We integrated genomic structural equation modeling and multiple post-genome-wide association study (GWAS) methodologies to prioritize candidate SNPs associated with GD1-related variation, identifying 15 loci with strong statistical support. Subsequently, diverse transcriptome-wide association approaches were employed to pinpoint susceptibility gene signals strongly correlated with GD1. For selected candidate genes, we explored the potential structural consequences of missense variants using integrated structure prediction, molecular dynamics simulations, and AI-based thermodynamic stability analyses. These analyses suggested that the mutations may alter protein structure and dynamics, with possible consequences for protein stability and biological function. Next, we screened a large set of publicly available traits to identify GD1-related factors and biomarkers with potential relevance. Finally, a summary data-based polygenic risk score (PRS) was utilized to examine risk associations between 22 autosomes and GD1. Collectively, by modeling a GD1-related phenotype without direct prior measurement, this study provides an initial overview of the shared genetic architecture associated with GD1.

Neurodevelopmental disorders (NDDs) encompass a wide range of conditions often linked to genetic causes, with mutations in the X-linked ARX gene representing a recurrent contributor. ARX encodes a transcription factor critical for GABAergic neuron development and functioning, regulating the expression of key neurodevelopmental target genes. Variants in ARX result in a wide clinical spectrum, ranging from asymptomatic female carriers to severe developmental syndromes in both sexes. This study investigates the functional impact of 16 ARX variants, including known pathogenic, likely pathogenic, and novel de novo variants, some associated with atypical presentations such as sudden infant death. Using transient expression in N2a neuroblastoma cells, we employed a comprehensive functional approach-including luciferase reporter assays, Western blotting, immunofluorescence, and RT-qPCR-to assess protein expression levels, subcellular localization, their interaction with known corepressors (TLE1 and CtBP1), and their transcriptional activity on selected ARX-known targets. Our results demonstrate that all tested variants disrupt normal ARX transcriptional function, with several also altering protein localization or expression. Remarkably, a subset of variants exhibited dominant-negative effects, offering a compelling explanation for unexpectedly severe phenotypes in female patients, likely exacerbated by skewed X-inactivation and other modifying factors. By integrating experimental data with a literature-based review of published ARX variants, we provide refined genotype-phenotype correlations, highlighting the importance of mutation type, positional context within key ARX functional domains, and patient sex. Altogether, this study advances our understanding of the molecular mechanisms driving ARX-related NDDs, emphasizing the importance of functional testing for accurate variant interpretation and paving the way toward informed genetic counseling and potential therapeutic development.

Neuroblastoma is a pediatric malignancy characterized by significant clinical heterogeneity. Although MYCN amplification is a well-established marker of high-risk disease, its interplay with the tumor immune microenvironment-particularly tumor-associated macrophages (TAMs)-remains poorly understood. In this study, we developed an integrated gene signature incorporating genes associated with both MYCN amplification status and TAM infiltration, leading to the identification of 16 differentially expressed genes implicated in both biological processes. Six of these genes (CMBL, LY6E, KLRB1, CTSH, CD3D, and PTGDS) were utilized to construct a risk-scoring model that effectively stratified neuroblastoma patients into high- and low-risk groups with significantly distinct clinical outcomes (p < 0.001). Notably, LY6E emerged as the most prognostically significant gene within the signature. More importantly, we revealed that LY6E modulates M2-type macrophage polarization in neuroblastoma for the first time, suggesting a novel mechanism through which it may contribute to shaping an immunosuppressive tumor microenvironment.

Radiotherapy is a fundamental component of rectal cancer treatment, yet patient responses remain highly heterogeneous due to the lack of reliable biomarkers supported by genomic variation evidence. Integrating multi-cohort transcriptomic data with machine-learning approaches enables systematic identification of genes with both predictive and therapeutic relevance. This study aimed to develop a robust model for predicting radiotherapy response and to functionally characterize CES1 as a key regulator of radiosensitivity and a potential therapeutic target. Three independent GEO cohorts were standardized and integrated, followed by a comprehensive machine-learning pipeline incorporating LASSO, Elastic Net, Random Forest, XGBoost, and information gain. A consensus-ranked five-gene model was constructed using nested cross-validation. CES1, identified as the top-ranked contributor to model performance, was selected for biological validation. To connect transcriptomic findings with genetic variation, copy number alteration (GISTIC2) and somatic mutation analyses were performed in TCGA-READ. Functional assays-including quantitative PCR, CCK-8 viability assays, colony formation, wound-healing migration assays, Annexin V/PI flow cytometry, and rescue by CES1 overexpression-were performed in HT-29 and SW480 cells to evaluate its mechanistic role in radiotherapy response. The machine-learning model demonstrated high discriminative accuracy across datasets and consistently highlighted CES1 as a dominant contributor to radiosensitivity prediction. CES1 expression increased after clinical chemoradiotherapy and showed dose-dependent induction following irradiation in vitro. CES1 knockdown significantly reduced radiation-induced apoptosis, enhanced clonogenic survival, and promoted migratory capacity, collectively indicating a radioresistant and more aggressive phenotype. Restoration of CES1 expression in CES1-silenced cells reversed radioresistance and re-established irradiation sensitivity. Genomic analysis in TCGA-READ further demonstrated that CES1 expression was positively associated with copy number status, whereas coding-sequence mutations in CES1 were infrequent, suggesting dysregulation primarily through copy-number and transcriptional mechanisms. This integrative computational and experimental study identifies CES1 as a predictive biomarker and copy number-linked regulator of radiosensitivity in rectal cancer. Modulation of CES1 directly alters cellular responses to irradiation, supporting its role as a mechanistically interpretable biomarker for response stratification. These findings align with the emerging concept that integrating genetic variation profiling with functional validation can accelerate variant-to-biomarker translation in precision oncology.

Genetic variants are the leading cause of congenital cataract (CC). To date, numerous genes have been implicated in the development of CC. The objective of the present study was to report two previously unrecognized gene variants associated with CC in two unrelated Chinese families, identified through whole exome sequencing (WES). Two unrelated Chinese families affected by CC were recruited. Cytogenetic and molecular genetic analyses were performed using karyotyping, chromosomal microarray analysis (CMA), and WES. In addition, RNA sequencing was conducted to assess differentially expressed genes in affected individuals compared with healthy controls. Karyotype and CMA elicited none of chromosome abnormalities in both of the families. However, WES revealed a novel missense variant NM_006891.4:c.154 T > C(p.S52P) in the CRYGD gene in the proband of Family 1, which was inherited from her mother with CC. In Family 2, a novel frameshift variant NM_000276.4:c.1046dup(p.M349Ifs∗36) in the OCRL gene was identified in the fetus via WES, which was inherited from the mother who had CC. RNA sequencing further demonstrated significantly reduced OCRL mRNA expression in the fetus compared with age-matched controls. The present study reports, for the first time, two novel variants in CRYGD and OCRL that were identified in the Chinese families with CC. These findings may expand the mutational spectrum of CC and highlight the utility of WES for the genetic diagnosis of patients with CC.

Most of pituitary adenomas are biologically benign, but some grow local‐invasively and can invade important adjacent tissues, resulting in clinical symptoms such as hormone secretion disorders and visual field defects. MicroRNA‐221/222 (miR‐221/222) is tandemly encoded on the X chromosome in humans, mice and rats, and is highly conserved in vertebrates with the same seed sequence. To date, miR‐221/222 has been reported as either a tumor suppressor or a tumor promoter in different tumors, however, its role in pituitary tumors has not been elucidated. Our study aimed to investigate the effect and mechanism of miRNA‐221/222 in pituitary tumor cells. Results of real‐time quantitative PCR showed that the expression level of miRNA‐221/222 in plasma exosomes from patients with pituitary tumor was significantly higher than that from healthy people. Results of cell function experiments indicated that miRNA‐221/222 significantly promoted cell proliferation and migration, inhibited apoptosis and significantly inhibited the expression of Cleaved‐Caspase3 and E‐cadherin, while promoted the expression level of N‐cadherin. With transcriptome sequencing and comprehensive bioinformatics analysis, PHACTR4 was identified as the potential target gene of miRNA‐221/222 in regulating biological functions of pituitary adenoma cells. Dual luciferase reporter assay confirmed that PHACTR4 was the direct target gene of miRNA‐221/222 and overexpression of PHACTR4 gene reversed the regulatory effects of miRNA‐221/222. In vivo experiment of subcutaneous tumor formation in nude mice verified that miRNA‐221/222 promoted tumor growth by targeting PHACTR4. In conclusion, miRNA‐221/222 played the role of proto‐oncogene in the occurrence and development of pituitary tumors by targeting PHACTR4, which provided a new target for the diagnosis and molecular treatment of pituitary adenomas.