Novel methodologies for detection of chromosomal abnormalities have been made available in the recent years but their clinical utility in prenatal settings is still unknown. We have conducted a ...comparative study of currently available methodologies for detection of chromosomal abnormalities after invasive prenatal sampling. A multicentric collection of a 1-year series of fetal samples with indication for prenatal invasive sampling was simultaneously evaluated using three screening methodologies: (1) karyotype and quantitative fluorescent polymerase chain reaction (QF-PCR), (2) two panels of multiplex ligation-dependent probe amplification (MLPA), and (3) chromosomal microarray-based analysis (CMA) with a targeted BAC microarray. A total of 900 pregnant women provided informed consent to participate (94% acceptance rate). Technical performance was excellent for karyotype, QF-PCR, and CMA (~1% failure rate), but relatively poor for MLPA (10% failure). Mean turn-around time (TAT) was 7 days for CMA or MLPA, 25 for karyotype, and two for QF-PCR, with similar combined costs for the different approaches. A total of 57 clinically significant chromosomal aberrations were found (6.3%), with CMA yielding the highest detection rate (32% above other methods). The identification of variants of uncertain clinical significance by CMA (17, 1.9%) tripled that of karyotype and MLPA, but most alterations could be classified as likely benign after proving they all were inherited. High acceptability, significantly higher detection rate and lower TAT, could justify the higher cost of CMA and favor targeted CMA as the best method for detection of chromosomal abnormalities in at-risk pregnancies after invasive prenatal sampling.
Nuclear transfer experiments undertaken in the mid-80's revealed that both maternal and paternal genomes are necessary for normal development. This is due to genomic imprinting, an epigenetic ...mechanism that results in parent-of-origin monoallelic expression of genes regulated by germline-derived allelic methylation. To date, ∼100 imprinted transcripts have been identified in mouse, with approximately two-thirds showing conservation in humans. It is currently unknown how many imprinted genes are present in humans, and to what extent these transcripts exhibit human-specific imprinted expression. This is mainly due to the fact that the majority of screens for imprinted genes have been undertaken in mouse, with subsequent analysis of the human orthologues. Utilizing extremely rare reciprocal genome-wide uniparental disomy samples presenting with Beckwith-Wiedemann and Silver-Russell syndrome-like phenotypes, we analyzed ∼0.1% of CpG dinculeotides present in the human genome for imprinted differentially methylated regions (DMRs) using the Illumina Infinium methylation27 BeadChip microarray. This approach identified 15 imprinted DMRs associated with characterized imprinted domains, and confirmed the maternal methylation of the RB1 DMR. In addition, we discovered two novel DMRs, first, one maternally methylated region overlapping the FAM50B promoter CpG island, which results in paternal expression of this retrotransposon. Secondly, we found a paternally methylated, bidirectional repressor located between maternally expressed ZNF597 and NAT15 genes. These three genes are biallelically expressed in mice due to lack of differential methylation, suggesting that these genes have become imprinted after the divergence of mouse and humans.
Abstract
Background
Imprinting disorders, which affect growth, development, metabolism and neoplasia risk, are caused by genetic or epigenetic changes to genes that are expressed from only one ...parental allele. Disease may result from changes in coding sequences, copy number changes, uniparental disomy or imprinting defects. Some imprinting disorders are clinically heterogeneous, some are associated with more than one imprinted locus, and some patients have alterations affecting multiple loci. Most imprinting disorders are diagnosed by stepwise analysis of gene dosage and methylation of single loci, but some laboratories assay a panel of loci associated with different imprinting disorders. We looked into the experience of several laboratories using single-locus and/or multi-locus diagnostic testing to explore how different testing strategies affect diagnostic outcomes and whether multi-locus testing has the potential to increase the diagnostic efficiency or reveal unforeseen diagnoses.
Results
We collected data from 11 laboratories in seven countries, involving 16,364 individuals and eight imprinting disorders. Among the 4721 individuals tested for the growth restriction disorder Silver–Russell syndrome, 731 had changes on chromosomes 7 and 11 classically associated with the disorder, but 115 had unexpected diagnoses that involved atypical molecular changes, imprinted loci on chromosomes other than 7 or 11 or multi-locus imprinting disorder. In a similar way, the molecular changes detected in Beckwith–Wiedemann syndrome and other imprinting disorders depended on the testing strategies employed by the different laboratories.
Conclusions
Based on our findings, we discuss how multi-locus testing might optimise diagnosis for patients with classical and less familiar clinical imprinting disorders. Additionally, our compiled data reflect the daily life experiences of diagnostic laboratories, with a lower diagnostic yield than in clinically well-characterised cohorts, and illustrate the need for systematising clinical and molecular data.
The aim in this study was to translate and cross-culturally adapt the Genetic Counseling Outcome Scale (GCOS-24) for use in Spain and to carry out a preliminary psychometric validation in a sample of ...Spanish patients. With oversight by an expert panel, forward and backward translations were conducted to create the draft Spanish GCOS-24. Fourteen patients were recruited from a clinical genetics service in Madrid, Spain, to participate in cognitive interviews designed to explore readability and interpretability of the draft. Following qualitative analysis of interview transcripts, a final version of the Spanish GCOS-24 was agreed with the expert panel. No significant cross-cultural differences were identified. The Spanish GCOS-24 was then completed prior to and 2–4 weeks after genetic counseling by 59 patients attending the service, and data were analysed using analysis of variance. Preliminary psychometric validation of the Spanish GCOS-24 showed significantly higher GCOS-24 scores after genetic counseling (
p
< 0.0001), with good internal consistency (α = 0.84) and sensitivity to change over time, with a medium-to-large size effect (Cohen’s d = 0.70). This compares well with the original English language GCOS-24. Findings demonstrate that the Spanish GCOS-24 has potential for use in evaluating clinical genetics services in Spain, but would benefit from assessment of test-retest reliability as well as structural and construct validity.
Pulmonary Arterial Hypertension (PAH) is a severe complication of Connective Tissue Disease (CTD), with remarkable morbidity and mortality. However, the molecular and genetic basis of CTD-PAH remains ...incompletely understood. This study aimed to screen for genetic defects in a cohort of patients with CTD-PAH, using a PAH-specific panel of 35 genes. During recruitment, 79 patients were studied, including 59 Systemic Sclerosis patients (SSc) and 69 females. Disease-associated variants were observed in nine patients: 4 pathogenic/likely pathogenic variants in 4 different genes (TBX4, ABCC8, KCNA5 and GDF2/BMP9) and 5 Variants of Unknown Significance (VUS) in 4 genes (ABCC8, NOTCH3, TOPBP1 and CTCFL). One patient with mixed CTD had a frameshift pathogenic variant in TBX4. Two patients with SSc-PAH carried variants in ABCC8. A patient diagnosed with Systemic Lupus Erythematous (SLE) presented a pathogenic nonsense variant in GDF2/BMP9. Another patient with SSc-PAH presented a pathogenic variant in KCNA5. Four patients with SSc-PAH carried a VUS in NOTCH1, CTCFL, CTCFL and TOPBP1, respectively. These findings suggest that genetic factors may contribute to Pulmonary Vascular Disease (PVD) in CTD patients.
We designate a novel term “isolated lateralized overgrowth” (ILO) for the findings previously described as “isolated hemihypertrophy” and “isolated hemihyperplasia.” ILO is defined as lateralized ...overgrowth in the absence of a recognized pattern of malformations, dysplasia, or morphologic variants. ILO is likely genetically heterogeneous. Further study is required to determine more of the underlying genetic etiologies and potential associations with currently unrecognized patterns of malformation.
Pulmonary arterial hypertension (PAH) is a severe clinical condition characterized by an increase in mean pulmonary artery pressure, which leads to a right ventricular hypertrophy and potentially ...heart failure and death. In the last several years, many genes have been associated with PAH, particularly in idiopathic and heritable forms but also in associated forms. Here we described the identification of two unrelated families in which the
variant was found from a cohort of 300 patients. The variants were identified by whole exome sequencing (WES). In the first family, the variant was detected in three affected members from a hereditary PAH, and in the second family the proband had PAH associated with scleroderma. In addition, we have reviewed all cases published in the literature thus far of patients with PAH and
variants. Functional studies have led to some contradictory conclusions, and the evidence of the relationship of
and PAH is still limited. However, we describe two further families with PAH and variants in
, expanding both the number of cases and the clinically associated phenotype. We provide further evidence of the association of
and the development of hereditary and associated forms of PAH.
Autism spectrum disorders (ASD) comprise a group of neurodevelopmental disorders (NDD) characterized by deficits in communication and social interaction, as well as repetitive and restrictive ...behaviors, etc. The genetic implications of ASD have been widely documented, and numerous genes have been associated with it. The use of chromosomal microarray analysis (CMA) has proven to be a rapid and effective method for detecting both small and large deletions and duplications associated with ASD. In this article, we present the implementation of CMA as a first-tier test in our clinical laboratory for patients with primary ASD over a prospective period of four years. The cohort was composed of 212 individuals over 3 years of age, who met DSM-5 diagnostic criteria for ASD. The use of a customized array-CGH (comparative genomic hybridization) design (KaryoArray
) found 99 individuals (45.20%) with copy number variants (CNVs); 34 of them carried deletions (34.34%) and 65 duplications (65.65%). A total of 28 of 212 patients had pathogenic or likely pathogenic CNVs, representing approximately 13% of the cohort. In turn, 28 out of 212 (approximately 12%) had variants of uncertain clinical significance (VUS). Our findings involve clinically significant CNVs, known to cause ASD (syndromic and non-syndromic), and other CNVs previously related to other comorbidities such as epilepsy or intellectual disability (ID). Lastly, we observed new rearrangements that will enhance the information available and the collection of genes associated with this disorder. Our data also highlight that CMA could be very useful in diagnosing patients with essential/primary autism, and demonstrate the existence of substantial genetic and clinical heterogeneity in non-syndromic ASD individuals, underscoring the continued challenge for genetic laboratories in terms of its molecular diagnosis.
Multiple Osteochondromatosis (MO, MIM 133700 & 133701), an autosomal dominant O-glycosylation disorder (EXT1/EXT2-CDG), can be associated with a reduction in skeletal growth, bony deformity, ...restricted joint motion, shortened stature and pathogenic variants in two tumor suppressor genes, EXT1 and EXT2. In this work, we report a cross-sectional study including 35 index patients and 20 affected family members. Clinical phenotyping of all 55 affected cases was obtained, but genetic studies were performed only in 35 indexes. Of these, a total of 40% (n = 14) had a family history of MO. Clinical severity scores were class I in 34% (n:18), class II in 24.5% (n:13) and class III in 41.5% (n:22). Pathogenic variants were identified in 83% (29/35) probands. We detected 18 (62%) in EXT1 and 11 (38%) in EXT2. Patients with EXT1 variants showed a height z-score of 1.03 SD lower than those with EXT2 variants and greater clinical severity (II–III vs. I). Interestingly, three patients showed intellectual impairment, two patients showed a dual diagnosis, one Turner Syndrome and one hypochondroplasia. This study improves knowledge of MO, reporting new pathogenic variants and forwarding the worldwide collaboration necessary to promote the inclusion of patients into future biologically based therapeutics.
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