Shift of N-MYC Oncogene Expression in AML Patients Carrying the FLT3-ITD Mutation.

Mutations in the FLT3 gene not only lead to abnormalities in its structure and function, but also affect the expression of other genes involved in leukemogenesis. This study evaluated the expression of genes that are more characteristic of neuroblastoma but less studied in leukemia. N-MYC oncogene expression was found to be more than 3-fold higher in primary AML patients carrying the FLT3-ITD mutation compared to carriers of other mutations as well as patients with normal karyotype (p = 0.03946). In contrast to the expression of several genes (C-MYC, SPT16, AURKA, AURKB) directly correlated to the allelic load of FLT3-ITD, the expression of the N-MYC oncogene is extremely weakly related or independent of it (p = 0.0405). Monitoring of N-MYC expression in some patients with high FLT3-ITD allelic load receiving therapy showed that a decrease in FLT3-ITD allelic load is not always accompanied by a decrease in N-MYC expression. On the contrary, N-MYC expression may remain elevated during the first three months after therapy, which is additional evidence of the emergence of resistance to therapy and progression of AML.

Characterization of the novel heterozygous SCN5A genetic variant Y739D associated with Brugada syndrome.

Genetic variants in SCN5A gene were identified in patients with various arrhythmogenic conditions including Brugada syndrome. Despite significant progress of last decades in studying the molecular mechanism of arrhythmia-associated SCN5A mutations, the understanding of relationship between genetics, electrophysiological consequences and clinical phenotype is lacking. We have found a novel genetic variant Y739D in the SCN5A-encoded sodium channel Nav1.5 of a male patient with Brugada syndrome (BrS). The objective of the study was to characterize the biophysical properties of Nav1.5-Y739D and provide possible explanation of the phenotype observed in the patient. The WT and Y739D channels were heterologously expressed in the HEK-293T cells and the whole-cell sodium currents were recorded. Substitution Y739D reduced the sodium current density by 47 ± 2% at -20 mV, positively shifted voltage-dependent activation, accelerated both fast and slow inactivation, and decelerated recovery from the slow inactivation. The Y739D loss-of-function phenotype likely causes the BrS manifestation. In the hNav1.5 homology models, which are based on the cryo-EM structure of rat Nav1.5 channel, Y739 in the extracellular loop IIS1-S2 forms H-bonds with K1381 and E1435 and pi-cation contacts with K1397 (all in loop IIIS5-P1). In contrast, Y739D accepts H-bonds from K1397 and Y1434. Substantially different contacts of Y739 and Y739D with loop IIIS5-P1 would differently transmit allosteric signals from VSD-II to the fast-inactivation gate at the N-end of helix IIIS5 and slow-inactivation gate at the C-end of helix IIIP1. This may underlie the atomic mechanism of the Y739D channel dysfunction.

Different Expressions of Pericardial Fluid MicroRNAs in Patients With Arrhythmogenic Right Ventricular Cardiomyopathy and Ischemic Heart Disease Undergoing Ventricular Tachycardia Ablation.

Introduction: Pericardial fluid is enriched with biologically active molecules of cardiovascular origin including microRNAs. Investigation of the disease-specific extracellular microRNAs could shed light on the molecular processes underlying disease development. Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited heart disease characterized by life-threatening arrhythmias and progressive heart failure development. The current data about the association between microRNAs and ARVC development are limited. Methods and Results: We performed small RNA sequence analysis of microRNAs of pericardial fluid samples obtained during transcutaneous epicardial access for ventricular tachycardia (VT) ablation of six patients with definite ARVC and three post-infarction VT patients. Disease-associated microRNAs of pericardial fluid were identified. Five microRNAs (hsa-miR-1-3p, hsa-miR-21-5p, hsa-miR-122-5p, hsa-miR-206, and hsa-miR-3679-5p) were found to be differentially expressed between patients with ARVC and patients with post-infarction VT. Enrichment analysis of differentially expressed microRNAs revealed their close linkage to cardiac diseases. Conclusion: Our data extend the knowledge of pericardial fluid microRNA composition and highlight five pericardial fluid microRNAs potentially linked to ARVC pathogenesis. Further studies are required to confirm the use of pericardial fluid RNA sequencing in differential diagnosis of ARVC.

A 300-kb microduplication of 7q36.3 in a patient with triphalangeal thumb-polysyndactyly syndrome combined with congenital heart disease and optic disc coloboma: a case report.

BACKGROUND: Triphalangeal thumb-polysyndactyly syndrome (TPT-PS) is a rare well-defined autosomal dominant disorder characterized by long thumbs with three phalanges combined with pre- and postaxial polydactyly/syndactyly of limbs. By now, the syndrome has been reported in several large families from different ethnic backgrounds, with a high degree of inter- and intrafamilial variability. The genome locus responsible for TPT-PS has been mapped to the 7q36.3 region harboring a long-range sonic hedgehog (SHH) regulatory sequence (ZRS). Both single-nucleotide variants and complete duplications of ZRS were shown to cause TPT-PS and similar limb phenotypes. TPT-PS usually forms as isolated limb pathology not associated with additional malformations, in particular, with cardiovascular abnormalities. CASE PRESENTATION: Here we report on a rare Russian neonatal case of TPT-PS combined with severe congenital heart disease, namely double outlet right ventricle, and microphthalmia with optic disc coloboma. Pedigree analysis revealed TPT-PS of various expressivity in 10 family members throughout five generations, while the cardiac defect and the eye pathology were detected only in the proband. To extend the knowledge on genotype-phenotype spectrum of TPT-PS, the careful clinical and genomic analysis of the family was performed. High-resolution array-based comparative genomic hybridization (array-CGH) revealed a ~ 300 kb microduplication of 7q36.3 locus (arr[GRCh37] 7q36.3(156385810_156684811) × 3) that co-segregated with TPT-PS in the proband and her mother. The duplication encompassed three genes including LMBR1, the intron 5 of which is known to harbor ZRS. Based on whole-exome sequencing data, no additional pathogenic mutations or variants of uncertain clinical significance were found in morbid cardiac genes or genes associated with a microphthalmia/anophthalmia/coloboma spectrum of ocular malformations. CONCLUSIONS: The results support the previous data, indicating that complete ZRS duplication underlies TPT-PS, and suggest a broader phenotypic impact of the 7q36.3 microduplication. Potential involvement of the 7q36.3 microduplication in the patient's cardiac and eye malformations is discussed. However, the contribution of some additional genetic/epigenetic factors to the complex patient`s phenotype cannot be excluded entirely. Further comprehensive functional studies are needed to prove the possible involvement of the 7q36.3 locus in congenital heart disease and eye pathology.

Genetic Spectrum of Left Ventricular Non-Compaction in Paediatric Patients.

INTRODUCTION: Left ventricular non-compaction (LVNC) represents a genetically heterogeneous cardiomyopathy which occurs in both children and adults. Its genetic spectrum overlaps with other types of cardiomyopathy. However, LVNC phenotypes in different age groups can have distinct genetic aetiologies. The aim of the study was to decipher the genetic spectrum of LVNC presented in childhood. Patient Group and Methods: Twenty patients under the age of 18 years diagnosed with LVNC were enrolled in the study. Target sequencing and whole-exome sequencing were performed using a panel of 108 cardiomyopathy-associated genes. Pathogenic, likely pathogenic, and variants of unknown significance found in genes highly expressed in cardiomyocytes were considered as variants of interest for further analysis. RESULTS: The median age at presentation was 8.0 (0.1-17) years, with 6 patients presenting before 1 year of age. Twelve (60%) patients demonstrated reduced ejection fraction. Right ventricular (RV) dilation was registered in 6 (30%), often in combination with reduced RV contractility (25%). Almost half (45%) of the patients demonstrated biventricular involvement already at disease presentation. For pathogenic and likely pathogenic variants, the positive genotyping rate was 45%, and these variants were found mainly in non-contractile structural sarcomeric genes (ACTN2, MYPN, and TTN) or in metabolic and signal transduction genes (BRAF and TAZ). Likely pathogenic TAZ variants were detected in all 5 patients suspected of having Barth syndrome. No pathogenic or likely pathogenic variants were found in genes encoding for sarcomeric contractile proteins, but variants of unknown significance were detected in 3 out of 20 patients (MYH6, MYH7, and MYLK2). In 4 patients, variants of unknown significance in ion-channel genes were detected. CONCLUSION: We detected a low burden of contractile sarcomeric variants in LVNC patients presenting below the age of 18 years, with the major number of variants residing in non-contractile structural sarcomeric genes. The identification of the variants in ion-channel and related genes not previously associated with LVNC in paediatric patients requires further examination of their functional role.

Generation of two iPSC lines (FAMRCi007-A and FAMRCi007-B) from patient with Emery-Dreifuss muscular dystrophy and heart rhythm abnormalities carrying genetic variant LMNA p.Arg249Gln.

Human iPSC lines were generated from peripheral blood mononuclear cells of patient carrying LMNA mutation associated with Emery-Dreifuss muscular dystrophy accompanied by atrioventricular block and paroxysmal atrial fibrillation. Reprogramming factors OCT4, KLF4, SOX2, CMYC were delivered using Sendai virus transduction. iPSCs were characterized in order to prove the pluripotency markers expression, normal karyotype, ability to differentiate into three embryonic germ layers. Generated iPSC lines would be useful model to investigate disease development associated with genetic variants in LMNA gene.

Generation of two induced pluripotent stem cell lines (FAMRCi005-A and FAMRCi005-B) from patient carrying genetic variant LMNA p.Asp357Val.

LMNA mutations are often linked to laminopathies characterized by tissue-specific disorders. We generated two induced pluripotent stem cells lines from patient carrying genetic variant LMNA p.Asp357Val associated with paroxysmal ventricular tachycardia and myopathy. Reprogramming of patient's peripheral blood mononuclear cells was performed using Sendai viruses. Characterization of the FAMRCi005-A and FAMRCi005-B lines revealed that generated iPSC lines expressed pluripotent stem cell markers, had normal karyotype and demonstrated triliniage differentiation ability. Generated cell lines can be used to investigate the molecular links between LMNA genetic variants and cardiac disorders.

Generation of two iPSC lines (FAMRCi006-A and FAMRCi006-B) from patient with dilated cardiomyopathy and Emery-Dreifuss muscular dystrophy associated with genetic variant LMNAp.Arg527Pro.

Mutations in LMNA gene are known to cause a broad range of diseases called laminopathies. We have generated two induced pluripotent stem cell lines FAMRCi006-A and FAMRCi006-B from a patient carrying LMNA p. p.Arg527Pro mutation associated with Emery-Dreifuss muscular dystrophy and dilated cardiomyopathy. Patient-specific peripheral blood mononuclear cells were reprogrammed to iPSCs using Sendai virus reprogramming system. Characterization of iPSCs had revealed pluripotency marker expression, normal karyotype, ability to differentiate into three embryonic germ layers. The reported iPSC lines could be a useful tool for in vitro modeling of laminopathies associated with LMNA genetic variants.

Truncating Variant in Myof Gene Is Associated With Limb-Girdle Type Muscular Dystrophy and Cardiomyopathy.

Even though genetic studies of individuals with neuromuscular diseases have uncovered the molecular background of many cardiac disorders such as cardiomyopathies and inherited arrhythmic syndromes, the genetic cause of a proportion of cardiomyopathies associated with neuromuscular phenotype still remains unknown. Here, we present an individual with a combination of cardiomyopathy and limb-girdle type muscular dystrophy where whole exome sequencing identified myoferlin (MYOF)-a member of the Ferlin protein family and close homolog of DYSF-as the most likely candidate gene. The disease-causative role of the identified variant c.[2576delG; 2575G>C], p.G859QfsTer8 is supported by functional studies in vitro using the primary patient's skeletal muscle mesenchymal progenitor cells, including both RNA sequencing and morphological studies, as well as recapitulating the muscle phenotype in vivo in zebrafish. We provide the first evidence supporting a role of MYOF in human muscle disease.