Bekim Sadikovic

NLRP7 affects trophoblast lineage differentiation, binds to overexpressed YY1 and alters CpG methylation

Maternal-effect mutations in NLRP7 cause rare biparentally inherited hydatidiform moles (BiHMs), abnormal pregnancies containing hypertrophic vesicular trophoblast but no embryo. BiHM trophoblasts display abnormal DNA methylation patterns affecting maternally methylated germline differentially methylated regions (gDMRs), suggesting that NLRP7 plays an important role in reprogramming imprinted gDMRs. How NLRP7-a component of the CATERPILLAR family of proteins involved in innate immunity and apoptosis-causes these specific DNA methylation and trophoblast defects is unknown. Because rodents lack NLRP7, we used human embryonic stem cells to study its function and demonstrate that NLRP7 interacts with YY1, an important chromatin-binding factor. Reduced NLRP7 levels alter DNA methylation and accelerate trophoblast lineage differentiation. NLRP7 thus appears to function in chromatin reprogramming and DNA methylation in the germline or early embryonic development, functions not previously associated with members of the NLRP family.

Recurrent genomic alterations in sequential progressive leukoplakia and oral cancer: drivers of oral tumorigenesis?

A significant proportion (up to 62%) of oral squamous cell carcinomas (OSCCs) may arise from oral potential malignant lesions (OPMLs), such as leukoplakia. Patient outcomes may thus be improved through detection of lesions at a risk for malignant transformation, by identifying and categorizing genetic changes in sequential, progressive OPMLs. We conducted array comparative genomic hybridization analysis of 25 sequential, progressive OPMLs and same-site OSCCs from five patients. Recurrent DNA copy number gains were identified on 1p in 20/25 cases (80%) with minimal, high-level amplification regions on 1p35 and 1p36. Other regions of gains were frequently observed: 11q13.4 (68%), 9q34.13 (64%), 21q22.3 (60%), 6p21 and 6q25 (56%) and 10q24, 19q13.2, 22q12, 5q31.2, 7p13, 10q24 and 14q22 (48%). DNA losses were observed in >20% of samples and mainly detected on 5q31.2 (35%), 16p13.2 (30%), 9q33.1 and 9q33.29 (25%) and 17q11.2, 3p26.2, 18q21.1, 4q34.1 and 8p23.2 (20%). Such copy number alterations (CNAs) were mapped in all grades of dysplasia that progressed, and their corresponding OSCCs, in 70% of patients, indicating that these CNAs may be associated with disease progression. Amplified genes mapping within recurrent CNAs (KHDRBS1, PARP1, RAB1A, HBEGF, PAIP2, BTBD7) were selected for validation, by quantitative real-time PCR, in an independent set of 32 progressive leukoplakia, 32 OSSCs and 21 non-progressive leukoplakia samples. Amplification of BTBD7, KHDRBS1, PARP1 and RAB1A was exclusively detected in progressive leukoplakia and corresponding OSCC. BTBD7, KHDRBS1, PARP1 and RAB1A may be associated with OSCC progression. Protein–protein interaction networks were created to identify possible pathways associated with OSCC progression.

Identification of a methylation profile for DNMT1-associated autosomal dominant cerebellar ataxia, deafness, and narcolepsy

BackgroundDNA methylation is an essential epigenetic mark, controlled by DNA methyltransferase (DNMT) proteins, which regulates chromatin structure and gene expression throughout the genome. In this study, we describe a family with adult-onset autosomal dominant cerebellar ataxia with deafness and narcolepsy (ADCA-DN) caused by mutations in the maintenance methyltransferase DNMT1 and assess the DNA methylation profile of these individuals.ResultsWe report a family with six individuals affected with ADCA-DN; specifically, patients first developed hearing loss and ataxia, followed by narcolepsy, and cognitive decline. We identified a heterozygous DNMT1 variant, c.1709C>T [p.Ala570Val] by Sanger sequencing, which had been previously reported as pathogenic for ADCA-DN and segregated with disease in the family. DNA methylation analysis by high-resolution genome-wide DNA methylation array identified a decrease in CpGs with 0–10xa0% methylation and 80–95xa0% methylation and a concomitant increase in sites with 10–30xa0% methylation and >95xa0% methylation. This pattern suggests an increase in methylation of normally unmethylated regions, such as promoters and CpG islands, as well as further methylation of highly methylated gene bodies and intergenic regions. Furthermore, a regional analysis identified 82 hypermethylated loci with consistent robust differences across ≥5 consecutive probes compared to our large reference cohort.ConclusionsThis report identifies robust changes in the DNA methylation patterns in ADCA-DN patients, which is an important step towards elucidating disease pathogenesis.

Mutation Update for UBE3A variants in Angelman syndrome.

Angelman syndrome is a neurodevelopmental disorder caused by a deficiency of the imprinted and maternally expressed UBE3A gene. Although de novo genetic and epigenetic imprinting defects of UBE3A genomic locus account for majority of Angelman diagnoses, approximately 10% of individuals affected with Angelman syndrome are a result of UBE3A loss‐of‐function mutations occurring on the expressed maternal chromosome. The variants described in this manuscript represent the analysis of 2,515 patients referred for UBE3A gene sequencing at our institution, along with a comprehensive review of the UBE3A mutation literature. Of these, 267 (10.62%) patients had a report issued for detection of a UBE3A gene nucleotide variant, which in many cases involved family studies resulting in reclassification of variants of unknown clinical significance (VUS). Overall, 111 (4.41%) probands had a nucleotide change classified as pathogenic or strongly favored to be pathogenic, 29 (1.15%) had a VUS, and 126 (5.0%) had a nucleotide change classified as benign or strongly favored to be benign. All variants and their clinical interpretations are submitted to NCBI ClinVar, a freely accessible human variation and phenotype database.

DNA methylation analysis in constitutional disorders: Clinical implications of the epigenome.

Abstract Genomic, chromosomal, and gene-specific changes in the DNA sequence underpin both phenotypic variations in populations as well as disease associations, and the application of genomic technologies for the identification of constitutional (inherited) or somatic (acquired) alterations in DNA sequence forms a cornerstone of clinical and molecular genetics. In addition to the disruption of primary DNA sequence, the modulation of DNA function by epigenetic phenomena, in particular by DNA methylation, has long been known to play a role in the regulation of gene expression and consequent pathogenesis. However, these epigenetic factors have been identified only in a handful of pediatric conditions, including imprinting disorders. Technological advances in the past decade that have revolutionized clinical genomics are now rapidly being applied to the emerging discipline of clinical epigenomics. Here, we present an overview of epigenetic mechanisms with a focus on DNA modifications, including the molecular mechanisms of DNA methylation and subtypes of DNA modifications, and we describe the classic and emerging genomic technologies that are being applied to this study. This review focuses primarily on constitutional epigenomic conditions associated with a spectrum of developmental and intellectual disabilities. Epigenomic disorders are discussed in the context of global genomic disorders, imprinting disorders, and single gene disorders. We include a section focused on integration of genetic and epigenetic mechanisms together with their effect on clinical phenotypes. Finally, we summarize emerging epigenomic technologies and their impact on diagnostic aspects of constitutional genetic and epigenetic disorders.

Gender and BCR‐ABL transcript type are correlated with molecular response to imatinib treatment in patients with chronic myeloid leukemia

Achieving a major molecular response (MMR) is the goal of imatinib therapy for chronic myeloid leukemia. However, the association between gender, BCR‐ABL transcript type, and age with MMR is not well understood and often controversial.

The defining DNA methylation signature of Kabuki syndrome enables functional assessment of genetic variants of unknown clinical significance

ABSTRACT Kabuki syndrome (KS) is caused by mutations in KMT2D, which is a histone methyltransferase involved in methylation of H3K4, a histone marker associated with DNA methylation. Analysis of >450,000 CpGs in 24 KS patients with pathogenic mutations in KMT2D and 216 controls, identified 24 genomic regions, along with 1,504 CpG sites with significant DNA methylation changes including a number of Hox genes and the MYO1F gene. Using the most differentiating and significant probes and regions we developed a “methylation variant pathogenicity (MVP) score,” which enables 100% sensitive and specific identification of individuals with KS, which was confirmed using multiple public and internal patient DNA methylation databases. We also demonstrated the ability of the MVP score to accurately reclassify variants of unknown significance in subjects with apparent clinical features of KS, enabling its potential use in molecular diagnostics. These findings provide novel insights into the molecular etiology of KS and illustrate that DNA methylation patterns can be interpreted as ‘epigenetic echoes’ in certain clinical disorders.

Prader–Willi syndrome and Tay–Sachs disease in association with mixed maternal uniparental isodisomy and heterodisomy 15 in a girl who also had isochromosome Xq

Malsegregation of chromosomes during reproduction can result in uniparental disomy when associated with trisomy rescue, monosomy rescue or gamete complementation. Pathogenicity stemming from uniparental disomy in liveborns results from imprinting disorders or autozygosity for autosomal recessive disorders. We report on a girl with Prader–Willi syndrome and Tay–Sachs disease resulting from maternal uniparental disomy of chromosome 15. The child also had an isochromosome Xq. To further characterize the etiology of the aberrant chromosome 15 and the isochromosome Xq, SNP loci from both chromosomes were assessed in the proband and parents, and genome‐wide DNA methylation analysis was performed. SNP and DNA methylation analysis confirmed maternal uniparental heterodisomy around the Prader–Willi locus, while the region around the HEXA locus showed maternal uniparental isodisomy. This result is consistent with trisomy rescue of a maternal meiosis l error in a chromosome 15 with two meiotic recombinations. SNP analysis of the X chromosomes is consistent with a maternal origin for the isochromosome.

MG-143 Age is significantly associated with the bone marrow engraftment in patients with allogeneic stem cell transplantation

Objectives Allogeneic stem cell transplantation is a curative therapy for patients with haematological malignancies and nonmalignant haematological disorders. Measuring the bone marrow engraftment via donor chimerism following transplantation is often used as a predictor for the overall survival. However, prognostic factors associated with bone marrow engraftment are poorly understood. The objective of this study is to evaluate the effect of age, gender and the recipient-donor relationship on the extent of bone marrow engraftment following transplantation and thereby provide more precise counselling to patients. Methods The bone marrow engraftment was routinely monitored using the multiplex fluorescent short tandem repeat analysis. Complete engraftment is defined as ≥95% of both donor-derived white blood cells and T cells. Data from a cohort of 194 patients with various conditions and up to 10 years of follow up was retrospectively analysed. Logistic regression and survival analyses were performed using the R statistic software. Results 64.9% of patients reached complete engraftment, ˜70% out of which achieved this milestone within 2 years post-transplantation; but 6.3% of these patients relapsed. Gender and recipient-donor relationship (related or not, same sex or not) did not have significant effect on the probability and the time for reaching complete engraftment. Age was inversely related to the probability of complete engraftment. The older the patient was, the less likely he/she achieved complete engraftment (p value 0.0015), and the longer time took for reaching complete engraftment (p value 0.0009). Conclusion Age is significantly associated with the likelihood and time for reaching complete engraftment.

A 42-year-old man with elevated ferritin

A 42-year-old businessman of South Asian descent presents with a prolonged history of fatigue and bothersome joint pain in his hands and wrists. The patient takes metformin for type 2 diabetes and ramipril for hypertension. He consumes 3 martinis per night and does not smoke. A physical examination