Diagnostic yield of rare skeletal dysplasia conditions in the radiogenomics era.

BACKGROUND: Skeletal dysplasia (SD) conditions are rare genetic diseases of the skeleton, encompassing a heterogeneous group of over 400 disorders, and represent approximately 5% of all congenital anomalies. Developments in genetic and treatment technologies are leading to unparalleled therapeutic advances; thus, it is more important than ever to molecularly confirm SD conditions. Data on 'rates-of-molecular yields' in SD conditions, through exome sequencing approaches, is limited. Figures of 39% and 52.5% have been reported in the USA (n = 54) and South Korea (n = 185) respectively. METHODS: We discuss a single-centre (in the UK) experience of whole-exome sequencing (WES) in a cohort of 15 paediatric patients (aged 5 months to 12 years) with SD disorders previously molecularly unconfirmed. Our cohort included patients with known clinical diagnoses and undiagnosed skeletal syndromes. Extensive phenotyping and expert radiological review by a panel of international SD radiology experts, coupled with a complex bioinformatics pipeline, allowed for both gene-targeted and gene-agnostic approaches. RESULTS: Significant variants leading to a likely or confirmed diagnosis were identified in 53.3% (n = 8/15) of patients; 46.7% (n = 7/15) having a definite molecular diagnosis and 6.7% (n = 1/15) having a likely molecular diagnosis. We discuss this in the context of a rare disease in general and specifically SD presentations. Of patients with known diagnoses pre-WES (n = 10), molecular confirmation occurred in 7/10 cases, as opposed to 1/5 where a diagnosis was unknown pre-test. Thus, diagnostic return is greatest where the diagnosis is known pre-test. For WGS (whole genome sequencing, the next iteration of WES), careful case selection (ideally of known diagnoses pre-test) will yield highest returns. CONCLUSIONS: Our results highlight the cost-effective use of WES-targeted bioinformatic analysis as a diagnostic tool for SD, particularly patients with presumed SD, where detailed phenotyping is essential. Thorough co-ordinated clinical evaluation between clinical, radiological, and molecular teams is essential for improved yield and clinical care. WES (and WGS) yields will increase with time, allowing faster diagnoses, avoiding needless investigations, ensuring individualised patient care and patient reassurance. Further diagnoses will lead to increased information on natural history/mechanistic details, and likely increased therapies and clinical trials.

Nebulin (NEB) mutations in a childhood onset distal myopathy with rods and cores uncovered by next generation sequencing.

Recessive nebulin (NEB) mutations are a common cause of nemaline myopathy (NM), typically characterized by generalized weakness of early-onset and nemaline rods on muscle biopsy. Exceptional adult cases with additional cores and an isolated distal weakness have been reported. The large NEB gene with 183 exons has been an obstacle for the genetic work-up. Here we report a childhood-onset case with distal weakness and a core-rod myopathy, associated with recessive NEB mutations identified by next generation sequencing (NGS). This 6-year-old boy presented with a history of gross-motor difficulties following a normal early development. He had distal leg weakness with bilateral foot drop, as well as axial muscle weakness, scoliosis and spinal rigidity; additionally he required nocturnal respiratory support. Muscle magnetic resonance (MR) imaging showed distal involvement in the medial and anterior compartment of the lower leg. A muscle biopsy featured both rods and cores. Initial targeted testing identified a heterozygous Nebulin exon 55 deletion. Further analysis using NGS revealed a frameshifting 4 bp duplication, c.24372_24375dup (P.Val8126fs), on the opposite allele. This case illustrates that NEB mutations can cause childhood onset distal NM, with additional cores on muscle biopsy and proves the diagnostic utility of NGS for myopathies, particularly when large genes are implicated.

Recessive mutations in EPG5 cause Vici syndrome, a multisystem disorder with defective autophagy.

Vici syndrome is a recessively inherited multisystem disorder characterized by callosal agenesis, cataracts, cardiomyopathy, combined immunodeficiency and hypopigmentation. To investigate the molecular basis of Vici syndrome, we carried out exome and Sanger sequence analysis in a cohort of 18 affected individuals. We identified recessive mutations in EPG5 (previously KIAA1632), indicating a causative role in Vici syndrome. EPG5 is the human homolog of the metazoan-specific autophagy gene epg-5, encoding a key autophagy regulator (ectopic P-granules autophagy protein 5) implicated in the formation of autolysosomes. Further studies showed a severe block in autophagosomal clearance in muscle and fibroblasts from individuals with mutant EPG5, resulting in the accumulation of autophagic cargo in autophagosomes. These findings position Vici syndrome as a paradigm of human multisystem disorders associated with defective autophagy and suggest a fundamental role of the autophagy pathway in the immune system and the anatomical and functional formation of organs such as the brain and heart.

Low prevalence of BRCA1 exon rearrangements in familial and young sporadic breast cancer patients.

BRCA1 exon deletions and duplications have been reported in a number of studies, and in order to design an effective mutation screening strategy in a diagnostic setting it is import to determine the frequency of this type of mutation in breast and ovarian cancer patients. We have designed and applied quantitative fluorescent PCR (QF-PCR) assays to screen for BRCA1 exon rearrangements in breast cancer patients both with and without a family history. A panel of 182 familial patients was screened, and an exon 3-7 deletion mutation was detected in a patient with a family history of breast and ovarian cancer. Additionally, we detected a duplication of exons 18-19 in an early onset sporadic breast cancer patient from a panel of 100 patients tested. These data indicate that in the absence of any founder mutations, screening for BRCA1 exon rearrangements does not significantly increase the overall BRCA1 mutation detection rate in patients referred to a genetics clinic because of either a family history and/or an early onset of disease.