MF Genetikos ir molekulinės medicinos klinika (04.20)

Introduction: WAGR syndrome is a rare contiguous gene deletion disorder characterized by the classical tetrad: Wilms tumor, aniridia, genitourinary anomalies, and a spectrum of developmental delays. It is caused by a deletion on chromosome 11p13, typically involving critical genes such as WT1 and PAX6. Objectives: To present a rare clinical case of WAGR syndrome with multisystem involvement, including endocrine and metabolic abnormalities. Methods: A 4-year-old girl was referred to a pediatric endocrinologist due to rapid weight gain and metabolic disturbances. Her anthropometric measurements were: height 100.2 cm (10th–25th percentile), weight 19 kg (75th–90th percentile), and BMI 18.92 kg/m² (approaching the 97th percentile). Laboratory evaluation revealed fasting hyperglycemia (up to 5.84 mmol/L) with insulin resistance (HOMA-IR 3.63), and central hypothyroidism (low TSH and low FT4). Previous medical history: she was the first child in the family. Aniridia was diagnosed shortly after birth. At the age of 1.5 years, she was diagnosed with nephroblastoma (Wilms tumor) of the right kidney. Given the suspicion of a rare syndromic condition, the patient was referred for genetic evaluation. Results: Genetic testing identified a pathogenic heterozygous deletion >7 Mb on chromosome 11p13, encompassing multiple genes including PAX6, WT1, BDNF, FSHB, DCDC1, ELP4, RCN1, HIPK3, and CD44. Haploinsufficiency of WT1, PAX6, and BDNF is considered central to the patient’s phenotype, contributing to tumor predisposition, ocular abnormalities, and neurodevelopmental and metabolic features. Conclusions: This case highlights the critical role of genetic testing in patients with congenital anomalies and multisystem involvement. WAGR syndrome may also present with endocrine and metabolic disturbances, including obesity, insulin resistance, and hypothyroidism. Early diagnosis and coordinated multidisciplinary management are essential to optimize clinical outcomes.

Objective Hypopharyngeal squamous cell carcinoma (HSCC) is a rare cancer of the head and neck with a poor overall survival. Genetic factors that influence disease pathogenesis remain unclear. TLR4 receptors are important for innate immunity, and mutations in their regulatory genes are linked to carcinogenesis; however, their genetic variants have not been studied in HSCC development. Methods This retrospective case–control study included 79 histologically confirmed HSCC patients and 220 healthy male control group. Clinical data, including age, tumor size, lymph node involvement, extra-nodal involvement (ENE), distant metastasis, stage, histological grade, and HPV status, were obtained from medical records. Genomic DNA was isolated from peripheral venous blood samples. Genotyping of selected TLR4 rs7037225 and rs7037117 polymorphisms was performed using real-time PCR with TaqMan probes. Results TLR4 rs7037117 polymorphism is significantly associated with higher ENE risk (p = 0.030). AG genotype (vs AA genotype) carriers have a 3.526-fold increased risk of ENE (OR = 3.526, 95% CI 1.204-10.328, p = 0.022). G allele carriers are significantly associated with ENE-positive status (p = 0.011). No significant associations were observed between the TLR4 rs7037225 polymorphism and HSCC risk or pathomorphological characteristics. Conclusions TLR4 polymorphisms may contribute to HSCC progression, as rs7037117 was associated with advanced lymph node metastasis.

Background and Objectives: This study aimed to determine the diagnostic yield of familial hypercholesterolemia (FH) genotyping and to describe the clinical phenotype and pharmacogenomic (PGx) implementation readiness of adults referred for clinically suspected FH at a Lithuanian tertiary cardiology center. The study was designed as a descriptive retrospective analysis of existing clinical and genetic test results generated using heterogeneous historical sequencing platforms. Materials and Methods: This single-center descriptive cohort included 48 adults with clinically suspected FH evaluated between February 2022 and June 2023. Patients were classified according to Dutch Lipid Clinic Network criteria before genetic testing. Historical targeted next-generation sequencing (NGS) and whole-exome sequencing (WES) results were reviewed. Genetic findings were interpreted using a three-tier reporting framework: diagnostic FH-associated findings, pharmacogenomic findings relevant to future implementation, and VUS, common polymorphisms, or other non-diagnostic findings. Because harmonized PGx star-allele/diplotype calling was not available across historical platforms, PGx findings were not analysed as a mutually exclusive patient-level category. Results: Pathogenic FH-associated variants were identified in 13 of 48 patients (27.1%), including eight LDLR and five APOB variant carriers. In the remaining 35 patients (72.9%), no pathogenic or likely pathogenic FH-associated variant was identified within the analytical scope of the historical test used. This non-diagnostic category could include VUS, common polymorphisms, exploratory findings, or no reportable variant. Patients carrying pathogenic LDLR variants had higher total cholesterol and LDL-C concentrations than patients without detected pathogenic variants. PGx-related findings were considered observations relevant to reporting and future implementation; they were not evaluated as predictors of statin response, statin intolerance, or LDL-C reduction. Conclusions: In this Lithuanian real-world cohort, pathogenic LDLR or APOB variants were identified in 27.1% of patients referred for suspected FH. Retrospective analyses of heterogeneous historical genetic results can inform diagnostic reporting and implementation planning but cannot establish PGx associations with statin efficacy or intolerance without harmonized diplotype calling and prospectively collected treatment outcomes.

This study investigated the antiproliferative effects of luteolin (LUT) and its potential to modify the response of breast cancer (BC) cells to ionizing radiation (IR). LUT effects alone or with IR were evaluated in MCF-7 and MDA-MB-231 cells using clonogenic and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. Apoptosis and cell cycle distribution were assessed by flow cytometry, while reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blotting were used to determine apoptosis-related gene and protein expression. LUT reduced the clonogenic survival and metabolic activity in both BC cell lines. In MCF-7 cells, combined LUT and IR treatment produced a greater reduction in metabolic activity and clonogenic survival than either treatment alone, whereas no comparable effect was observed in MDA-MB-231 cells. LUT induced apoptosis in both cell lines. Cell cycle analysis indicated G2/M phase accumulation, reaching statistical significance in MDA-MB-231 cells. In MCF-7 cells, LUT decreased BCL2 and MCL1 gene expression, with a further reduction following combined LUT and IR treatment. Consistently, BCL-2 protein levels were reduced in MCF-7 cells. In MDA-MB-231 cells, LUT modulated the expression of selected anti-apoptotic genes. Overall, these findings demonstrate cell line-dependent effects of LUT and suggest that its ability to modify the response to IR may depend on the molecular characteristics of BC cells.

Introduction. Recurrent nephrolithiasis requires evaluation for underlying metabolic disorders. Impaired vitamin D degradation due to mutations in CYP24A1 leads to increased active vitamin D levels, resulting in hypercalciuria and stone formation. Objectives. To present a case of recurrent nephrolithiasis associated with hypercalciuria and abnormal calcium metabolism, emphasizing the role of genetic evaluation in atypical presentations. Materials and Methods. A 54-year-old male with recurrent nephrolithiasis since 2011, requiring multiple lithotripsy procedures, was evaluated. Clinical history, biochemical parameters, and 24-hour urine analysis were assessed. Results. Serum calcium was intermittently elevated (2.49-2.73 mmol/L) with non-suppressed parathyroid hormone (1.62 pmol/L). Phosphate, alkaline phosphatase, fibroblast growth factor 23, and parathyroid hormone-related peptide were within normal limits. Both 25-hydroxyvitamin D (122 nmol/L) and 1,25-dihydroxyvitamin D (55.5 ng/L) were within reference ranges. A 24-hour urine collection (2300 mL) demonstrated marked hypercalciuria (16.95 mmol/24 h), with elevated sodium (286 mmol/24 h), phosphate (45.8 mmol/24 h), and magnesium (6.91 mmol/24 h). Whole-exome sequencing identified compound heterozygous pathogenic variants in CYP24A1 (NM_000782.5: c.1186C>T;425AAG[1J). Low-dose hydrochlorothiazide reduced urinary calcium to 11.88 mmol/24 h, with serum calcium stabilizing at 2.65 mmol/L. Management included avoidance of vitamin D supplementation and excessive sun exposure. Conclusions. Recurrent nephrolithiasis may reflect inherited disorders of vitamin D metabolism, even in the absence of family history due to autosomal recessive inheritance. CYP24A1 mutation should be considered in patients with marked hypercalciuria, intermittent hypercalcemia, and non-suppressed parathyroid hormone, even when both 25-hydroxyvitamin D and 1,25-dihydroxyvitamin D levels are within normal ranges. Early diagnosis allows for targeted management and prevention of further stone formation.

Background and Objectives: Echocardiographic assessment of left ventricular diastolic dysfunction (LVDD) in hypertrophic cardiomyopathy (HCM) is challenging, as conventional parameters correlate poorly with left ventricular filling pressure. Emerging evidence suggests that left atrial (LA) strain is associated with atrial fibrillation (AF). This study aimed to compare LA reservoir strain (LASr)-based and conventional LVDD grading, evaluate associations between LA strain and clinical and echocardiographic parameters, and explore its relationship with AF. Materials and Methods: This retrospective cross-sectional single-center study included 109 adult HCM patients with comprehensive echocardiographic data. LVDD was graded using conventional echocardiographic parameters and LASr-based thresholds (≥35% (grade 0), ≥24% to <35% (grade 1), ≥19% to <24% (grade 2), and < 19% (grade 3)). Results: LASr-based grading revealed a significantly greater prevalence of advanced LVDD compared with conventional assessment (grade 3: 39% vs. 3%, p < 0.001), with poor intermethod agreement (κ = 0.07). Overall, approximately 45% of patients were reclassified into higher LVDD grades using LASr, particularly among nonobstructive HCM patients. Higher LASr-based LVDD grades were associated with worse conventional diastolic indices, reduced biventricular strain parameters, and higher natriuretic peptide levels. Reduced LA reservoir and conduit strain were independently associated with documented AF after adjustment for age, LA volume index, and left ventricular mass index. Conclusions: LASr provides additional information on diastolic dysfunction in HCM patients and is associated with documented AF. LASr-based grading assigned more patients to advanced LVDD grades than conventional assessment. The observed association with AF requires prospective validation before LA strain can be used for risk stratification.

Dyslipidemia and elevated low-density lipoprotein cholesterol (LDL-C) are major drivers of atherosclerotic cardiovascular disease, yet response to statin therapy varies between patients. We investigated clinical and genetic factors associated with LDL-C response to atorvastatin in patients with ST-elevation myocardial infarction (STEMI). A total of 93 patients underwent targeted next-generation sequencing of a pharmacogene panel, and LDL-C was measured at baseline and after 6 months. The mean ΔLDL-C was − 1.46 ± 0.97 mmol/L. After exclusion of singleton variants, 176 variants remained for analysis, including 68 rare variants (minor allele frequency < 0.05). In single-variant analyses adjusted for age, sex, smoking status, and baseline LDL-C, the CYP2C19 phenotype showed the strongest nominal association with ΔLDL-C, followed by KIF6 rs20455, ABCB1 rs3842, and APOB rs72654423; however, none remained significant after false discovery rate correction. Exploratory aggregate analyses showed associations between the transport pathway score and the weighted polygenic risk score and LDL-C response. In the burden model, CYP2C19 phenotype, transport gene variation, age, and baseline LDL-C were associated with ΔLDL-C. For the binary treatment endpoint, the full model combining clinical and genetic variables showed higher apparent in-sample discrimination than the genetic-only model (AUC 0.829 vs. 0.710; DeLong test p = 0.0044). These exploratory findings require independent validation before clinical application.

Primary bilateral macronodular adrenal hyperplasia (PBMAH) is a rare disease, accounting for <2% of cases of Cushing's syndrome (CS). Patients with PBMAH have varying levels of cortisol secretion and are often diagnosed with PBMAH due to incidental radiological findings of the adrenal glands, as in our case. During evaluation for angioedema, a CT scan in a 53-year-old woman revealed an incidental finding of nodular thickening up to ~11 mm in the left adrenal gland (most likely hyperplasia). Further abdominal MRI showed that the left adrenal gland is nodular, with a suspicious solid lesion measuring 1.2 × 0.9 cm with small fatty-density inclusions, based on MRI, a small pheochromocytoma was suspected; the right adrenal gland is structurally normal with no additional lesions. Further evaluation of this finding by an endocrinologist was carried out. An overnight dexamethasone suppression test was performed, and cortisol suppression was not achieved (56.24 nmol/L, n < 50). DXA showed normal BMD, and spinal X-ray revealed degenerative changes. Other laboratory findings included free metanephrines (metanephrine 0.08 nmol/L, n 0-0.507; normetanephrine 0.41 nmol/L, n 0-1.18), ACTH (2.1 pmol/L, n 1.63-14.15), TSH (2.03 mU/L, n 0.4-3.6), and a normal aldosterone-renin ratio (43). Whole-exome sequencing identified a heterozygous pathogenic variant in the ARMC5 gene NM_001105247.2:c.517C>T (p.Arg173Ter). This gene is associated with autosomal dominant PBMAH and has incomplete penetrance, as a second hit in the gene is required for PBMAH to develop. A repeat overnight dexamethasone suppression test again showed inadequate suppression (58 nmol/L, n < 50). A low-dose dexamethasone test was then performed, and suppression was achieved (40.6 nmol/L, n < 50); overall, the tests revealed mild autonomous cortisol hypersecretion. The patient case was discussed at a MDT on rare endocrine and adrenal diseases. Although the patient carries this variant, because adrenal hyperplasia is unilateral and she currently does not have typical clinical features of CS, she does not at present meet the criteria for PBMAH and can be considered a presymptomatic carrier of a genetic variant. In patients with pathogenic ARMC5 variants, PBMAH and CS most often present in the fifth or sixth decade of life, ie, with later onset compared with unilateral cortisol-producing adrenal adenomas. Therefore, the patient requires regular endocrinological follow-up, as there is a high likelihood of developing PBMAH and CS in the future. Monitoring facilitates early treatment initiation and helps prevent progression of CS-related complications such as cardiovascular, metabolic, and musculoskeletal disorders.

Introduction Radiotherapy is a vital therapeutic option in the treatment of breast cancer nowadays. However, a major obstacle to the full effectiveness of radiation therapy is still the radioresistance of cancer cells. Various studies have proven sulforaphane's (SFN) beneficial effects against cancer and its possible utilization as a radiosensitizer in radiotherapy. This study aimed to investigate whether SFN has a radiosensitizing effect on breast cancer cells.

Material and methods The anticancer efficiency of SFN and radiosensitizing effect in MCF-7 and MDA-MB-231 cell lines were assessed by the MTT assay. Using a flow cytometric assay, the apoptosis level and changes in the cell cycle were measured. RT-qPCR and Western blot analysis were used to determine BCL-2 and BCL-XL genes expresion and proteins level.

Results According to our results, SFN reduced the viability of cells, and combining SFN with radiation therapy (IR) caused much greater anticancer effects on cells. SFN+IR was shown to enhance the number of cells in the G2/M phase and the percentage of cells going through apoptosis. SFN reduced the expression of apoptosis-relative genes BCL-2 and BCL-XL. Consistent with this data, Western blot analysis revealed that BCL-2 and BCL-XL protein levels were decreased in tested cells. As a result of the combination treatment, the downregulation of the BCL-2 protein was observed only in MDA-MB-231 cells.

Conclusions These results indicate that SFN acts as a radiosensitizer by enhancing apoptotic cell death and reducing anti-apoptotic genes in breast cancer cells.

Background/Objectives: Dilated cardiomyopathy is a heterogeneous disorder with a substantial genetic contribution from a variety of pathogenic variants. Hereditary isolated DCM is often caused by variants in genes encoding sarcomere proteins, as well as proteins involved in desmosomes or other cardiac cell functions. Identifying genetic causes improves our understanding of DCM pathophysiology, facilitates prognostic assessment, and enables more personalized disease management. Methods: We retrospectively analyzed genetic data from adult patients with a clinical diagnosis of isolated DCM evaluated at a Lithuanian tertiary university hospital between 2019 and 2024. All patients were tested with a next-generation sequencing cardiovascular gene panel. Results: We gathered 169 patients and initially reached a 16.0% (n = 27) genetic testing diagnostic yield. We performed all genetic variant reanalyses with the most current classification guidelines, and we found an additional eight positive cases. Our final diagnostic yield was 20.7% (n = 35). TTN was the most frequently affected gene (n = 30), whereas variants in BAG3 (n = 2), DSP (n = 1), LMNA (n = 1), and FLNC (n = 1) were rare. In total, 15 variants were novel—not described in the literature or databases. We did not observe significant clinical differences between patients with pathogenic variants and those without pathogenic variants. We expected a different clinical course with variants in genes like BAG3 or LMNA, but there were only a few cases. Conclusions: Genetic testing remains an important tool for confirming complex DCM cases and allows earlier disease management for relatives at risk.