Genomic landscape of human erythroleukemia K562 cell line, as determined by next-generation sequencing and cytogenetics

We have performed detailed analysis of the genomic landscape of commercially available K562 cells, employing targeted enrichment of nearly 1300 cancer-related genes followed by next-generation sequencing (NGS) and also classical cytogenetics. Deep sequencing revealed 88 variants of potentially biological significance. Among them we have detected alterations in genes already known to be mutated in K562, such as TP53 but also in several other genes, which are implicated in tumorigenesis and drug resistance, such as MLH1, ASXL1 and BRCA1 as the most prominent examples. Fluorescence in situ hybridization (FISH) of interphases of K562 cells revealed multiplication of the BCR and ABL1 gene copies, as well as the amplification of the BCR-ABL1 fusion gene. Our results may help to better understand genomic instability of the blastic phase of CML represented by the K562 cell line and can help researchers who want to employ this cell line in various experimental settings

First familial cases of type 2 congenital erythrocytosis (ECYT2) with a Chuvash pathogenic variant in gene in Poland: example of the clinical utility of next-generation sequencing in diagnostics of orphan diseases

In this article, we report familial cases of type 2 congenital erythrocytosis (ECYT2) in two siblings, a 2-year-old boy and his younger sister. Both patients were diagnosed based on laboratory findings including erythrocytosis, elevated hemoglobin levels, and hematocrit. Acquired erythrocytosis was excluded based on the clinical features and genetic analysis of genes. Next-generation sequencing was employed for older brother revealing NM_000551.4: c.598C>T, p.Arg200Trp homozygous variant in the gene, the similar variant was detected in the younger sibling. Sequencing analysis confirmed the c.598C>T heterozygous variant in both parents. To the best of our knowledge, these are the first confirmed cases of familial erythrocytosis type 2, also known as Chuvash type, in Poland

Whole-exome sequencing in patients with protein aggregate myopathies reveals causative mutations associated with novel atypical phenotypes

BACKGROUND: Myofibrillar myopathies (MFM) are a subgroup of protein aggregate myopathies (PAM) characterized by a common histological picture of myofibrillar dissolution, Z-disk disintegration, and accumulation of degradation products into inclusions. Mutations in genes encoding components of the Z-disk or Z-disk-associated proteins occur in some patients whereas in most of the cases, the causative gene defect is still unknown. We aimed to search for pathogenic mutations in genes not previously associated with MFM phenotype.METHODS: We performed whole-exome sequencing in four patients from three unrelated families who were diagnosed with PAM without aberrations in causative genes for MFM.RESULTS: In the first patient and her affected daughter, we identified a heterozygous p.(Arg89Cys) missense mutation in LMNA gene which has not been linked with PAM pathology before. In the second patient, a heterozygous p.(Asn4807Phe) mutation in RYR1 not previously described in PAM represents a novel, candidate gene with a possible causative role in the disease. Finally, in the third patient and his symptomatic daughter, we found a previously reported heterozygous p.(Cys30071Arg) mutation in TTN gene that was clinically associated with cardiac involvement.CONCLUSIONS: Our study identifies a new genetic background in PAM pathology and expands the clinical phenotype of known pathogenic mutations

Results of Polish Adult Leukemia Study Group (PALG) project assessing TP53 mutations with next-generation sequencing technology in relapsed and refractory chronic lymphocytic leukemia patients — an 18-month update

Indtroduction and methods: In chronic lymphocytic leukemia (CLL), molecular and cytogenetic diagnostics are crucial for the determination of accurate prognosis and treatment choice. Among different genetic aberrations, del(17p13) or TP53 mutations constitute high-risk factors, and early identification of such defects is a high priority for CLL patients. While cytogenetic diagnostics is well-established and accessible for the majority of CLL patients in Poland, molecular diagnostics of TP53 mutations is performed only in a few ERIC-certified centers (eight as of September 2020), and only two of these employ next-generation sequencing (NGS) for routine analysis of TP53 status in CLL patients. Here we report the interim results of a project assessing TP53 mutations with NGS technology in relapsed or refractory CLL patients with confirmed negative del(17p13) status. 249 patients from 32 clinical centers were included in the study. Results: NGS analysis revealed TP53 mutations in 42/249 (17%) patients, half of whom (21/249, 8.5%) had subclonal mutations (VAF ≤10%). These results are in line with published data in relapsed/refractory CLL patients. Conclusions: The results of the project demonstrated the feasibility and accuracy of NGS testing in CLL patients despite several initial logistical and technical obstacles. Our study also proved that, with appropriate funding, CLL patients from any hematological center in Poland can have access to state-of-the-art molecular diagnostic

A rare mutation in a rare tumor-SMARCB1-deficient malignant glomus tumor

status: publishe

Variable degree of mosaicism for tetrasomy 18p in phenotypically discordant monozygotic twins-Diagnostic implications

Background: Phenotypically discordant monozygotic twins (PDMZTs) offer a unique opportunity to study post-zygotic genetic variation and provide insights into the linkage between genotype and phenotype. We report a comprehensive analysis of a pair of PDMZTs. Methods: Dysmorphic features and delayed neuro-motor development were observed in the proband, whereas her twin sister was phenotypically normal. Four tissues (blood, skin, hair follicles, and buccal mucosa) from both twins were studied using four complementary methods, including whole-exome sequencing, karyotyping, array CGH, and SNP array. Results: In the proband, tetrasomy 18p affecting all studied tissues except for blood was identified. Karyotyping of fibroblasts revealed isochromosome 18p [i(18p)] in all metaphases. The corresponding analysis of the phenotypically normal sister surprisingly revealed low-level mosaicism (5.4%) for i(18p) in fibroblasts. Conclusion: We emphasize that when mosaicism is suspected, multiple tissues should be studied and we highlight the usefulness of non-invasive sampling of hair follicles and buccal mucosa as a convenient source of non-mesoderm-derived DNA, which complements the analysis of mesoderm using blood. Moreover, low-level mosaic tetrasomy 18p is well tolerated and such low-level mosaicism, readily detected by karyotyping, can be missed by other methods. Finally, mosaicism for low-level tetrasomy 18p might be more common in the general population than it is currently recognized, due to detection limitations

Titin Truncating Variants in Dilated Cardiomyopathy – Prevalence and Genotype-Phenotype Correlations

<div><p><i>TTN</i> gene truncating variants are common in dilated cardiomyopathy (DCM), although data on their clinical significance is still limited. We sought to examine the frequency of truncating variants in <i>TTN</i> in patients with DCM, including familial DCM (FDCM), and to look for genotype-phenotype correlations. Clinical cardiovascular data, family histories and blood samples were collected from 72 DCM probands, mean age of 34 years, 45.8% FDCM. DNA samples were examined by next generation sequencing (NGS) with a focus on the <i>TTN</i> gene. Truncating mutations were followed up by segregation study among family members. We identified 16 <i>TTN</i> truncating variants (<i>TTN</i> trunc) in 17 probands (23.6% of all cases, 30.3% of FDCM, 17.9% of sporadic DCM). During mean 63 months from diagnosis, there was no difference in adverse cardiac events between probands with and without TTN truncating mutations. Among relatives 29 mutation carriers were identified, nine were definitely affected (31%), eight probably affected (27.6%) one possibly affected (3.4%) and eleven were not affected (37.9%). When relatives with all affected statuses were combined, disease penetrance was still incomplete (62.1%) even after exclusion of unaffected relatives under 40 (82%) and was higher in males versus females. In all mutation carriers, during follow-up, 17.4% had major adverse cardiac events, and prognosis was significantly worse in men than in women. In conclusion, <i>TTN</i> truncating variants were observed in nearly one fourth of young DCM patient population, in vast majority without conduction system disease. Incomplete penetrance suggests possible influence of other genetic and/or environmental factors on the course of cardiotitinopathy. Counseling should take into account sex and incomplete penetrance.</p></div

Comparison of clinical data on <i>TTN</i> trunc positive versus <i>TTN</i> trunc negative DCM probands.

<p>Comparison of clinical data on <i>TTN</i> trunc positive versus <i>TTN</i> trunc negative DCM probands.</p