The genomic landscape of balanced cytogenetic abnormalities associated with human congenital anomalies

Despite the clinical significance of balanced chromosomal abnormalities (BCAs), their characterization has largely been restricted to cytogenetic resolution. We explored the landscape of BCAs at nucleotide resolution in 273 subjects with a spectrum of congenital anomalies. Whole-genome sequencing revised 93% of karyotypes and demonstrated complexity that was cryptic to karyotyping in 21% of BCAs, highlighting the limitations of conventional cytogenetic approaches. At least 33.9% of BCAs resulted in gene disruption that likely contributed to the developmental phenotype, 5.2% were associated with pathogenic genomic imbalances, and 7.3% disrupted topologically associated domains (TADs) encompassing known syndromic loci. Remarkably, BCA breakpoints in eight subjects altered a single TAD encompassing MEF2C, a known driver of 5q14.3 microdeletion syndrome, resulting in decreased MEF2C expression. We propose that sequence-level resolution dramatically improves prediction of clinical outcomes for balanced rearrangements and provides insight into new pathogenic mechanisms, such as altered regulation due to changes in chromosome topology

Anaplastic Kaposi Sarcoma: A Clinicopathologic and Molecular Genetic Analysis

Kaposi sarcoma (KS) is an HHV8-associated vascular proliferation that most often involves the skin. Rarely, KS shows marked nuclear atypia or pleomorphism; such examples are known as "anaplastic" KS. This poorly characterized variant often pursues an aggressive course; little is known of its genetic landscape. This study evaluated the clinicopathologic and genomic features of anaplastic KS. We identified 9 anaplastic KS cases from 7 patients and 8 conventional KS cases, including a matched conventional KS and primary-metastasis anaplastic KS pair from a single patient (anaplastic KS diagnosed 9 years after conventional KS). All patients with anaplastic KS were male, aged 51-82 years, who had locally aggressive tumors predominantly affecting the soft tissue and bone of the lower extremities (5/7). Four patients were known to be HIV-positive (all on antiretrovirals), 2 were HIV-negative, and one was of unknown HIV status. The tumors showed angiosarcoma-like or pleomorphic spindle cell sarcoma morphology. Plasma cell-rich chronic inflammation and hemosiderin deposition were commonly present. Single nucleotide polymorphism (SNP) based chromosomal microarray analysis showed the anaplastic KS cohort to demonstrate highly recurrent whole chromosome (chr) gains of chr 7, 11, 19, and 21, which primarily affected olfactory and G-protein coupled receptor (GPCR) signaling, and losses of chr6_q and chrY. Compared to conventional KS, anaplastic KS cases showed significantly more total copy number alterations (CNAs) and more frequent gains of chr7 and chr11_q13.1 (MARK2, RELA, and ESRRA; including high copy number gain in 1 case). Pathway analysis demonstrated these gains preferentially affected genes that facilitate cyclin-dependent cell signaling. Further, anaplastic KS cases were phylogenetically distinct from conventional KS cases, including the patient-matched primary-metastasis anaplastic KS pair and conventional KS. Our study is the first to demonstrate that a more complex genome and distinct CNAs distinguish anaplastic KS from conventional KS. Gains of chr7 and chr11_q13.1 appear central to biological transformation

B9D1 is revealed as a novel Meckel syndrome (MKS) gene by targeted exon-enriched next-generation sequencing and deletion analysis

Meckel syndrome (MKS) is an embryonic lethal, autosomal recessive disorder characterized by polycystic kidney disease, central nervous system defects, polydactyly and liver fibrosis. This disorder is thought to be associated with defects in primary cilia; therefore, it is classed as a ciliopathy. To date, six genes have been commonly associated with MKS (MKS1, TMEM67, TMEM216, CEP290, CC2D2A and RPGRIP1L). However, mutation screening of these genes revealed two mutated alleles in only just over half of our MKS cohort (46 families), suggesting an even greater level of genetic heterogeneity. To explore the full genetic complexity of MKS, we performed exon-enriched next-generation sequencing of 31 ciliopathy genes in 12 MKS pedigrees using RainDance microdroplet-PCR enrichment and IlluminaGAIIx next-generation sequencing. In family M456, we detected a splice-donor site change in a novel MKS gene, B9D1. The B9D1 protein is structurally similar to MKS1 and has been shown to be of importance for ciliogenesis in Caenorhabditis elegans. Reverse transcriptase–PCR analysis of fetal RNA revealed, hemizygously, a single smaller mRNA product with a frameshifting exclusion of B9D1 exon 4. ArrayCGH showed that the second mutation was a 1.713 Mb de novo deletion completely deleting the B9D1 allele. Immunofluorescence analysis highlighted a significantly lower level of ciliated patient cells compared to controls, confirming a role for B9D1 in ciliogenesis. The fetus inherited an additional likely pathogenic novel missense change to a second MKS gene, CEP290; p.R2210C, suggesting oligogenic inheritance in this disorder

Deletion 17q12 is a recurrent copy number variant that confers high risk of autism and schizophrenia

Autism spectrum disorders (ASD) and schizophrenia are neurodevelopmental disorders for which recent evidence indicates an important etiologic role for rare copy number variants (CNVs) and suggests common genetic mechanisms. We performed cytogenomic array analysis in a discovery sample of patients with neurodevelopmental disorders referred for clinical testing. We detected a recurrent 1.4 Mb deletion at 17q12, which harbors HNF1B, the gene responsible for renal cysts and diabetes syndrome (RCAD), in 18/15,749 patients, including several with ASD, but 0/4,519 controls. We identified additional shared phenotypic features among nine patients available for clinical assessment, including macrocephaly, characteristic facial features, renal anomalies, and neurocognitive impairments. In a large follow-up sample, the same deletion was identified in 2/1,182 ASD/neurocognitive impairment and in 4/6,340 schizophrenia patients, but in 0/47,929 controls (corrected p = 7.37 × 10?5). These data demonstrate that deletion 17q12 is a recurrent, pathogenic CNV that confers a very high risk for ASD and schizophrenia and show that one or more of the 15 genes in the deleted interval is dosage sensitive and essential for normal brain development and function. In addition, the phenotypic features of patients with this CNV are consistent with a contiguous gene syndrome that extends beyond RCAD, which is caused by HNF1B mutations only.<br/

An evidence-based approach to establish the functional and clinical significance of copy number variants in intellectual and developmental disabilities

Purpose: Copy number variants have emerged as a major cause of human disease such as autism and intellectual disabilities. Because copy number variants are common in normal individuals, determining the functional and clinical significance of rare copy number variants in patients remains challenging. The adoption of whole-genome chromosomal microarray analysis as a first-tier diagnostic test for individuals with unexplained developmental disabilities provides a unique opportunity to obtain large copy number variant datasets generated through routine patient care.Methods: A consortium of diagnostic laboratories was established (the International Standards for Cytogenomic Arrays consortium) to share copy number variant and phenotypic data in a central, public database. We present the largest copy number variant case-control study to date comprising 15,749 International Standards for Cytogenomic Arrays cases and 10,118 published controls, focusing our initial analysis on recurrent deletions and duplications involving 14 copy number variant regions.Results: Compared with controls, 14 deletions and seven duplications were significantly overrepresented in cases, providing a clinical diagnosis as pathogenic.Conclusion: Given the rapid expansion of clinical chromosomal microarray analysis testing, very large datasets will be available to determine the functional significance of increasingly rare copy number variants. This data will provide an evidence-based guide to clinicians across many disciplines involved in the diagnosis, management, and care of these patients and their families.<br/