Genetic testing for mitochondrial disease: the United Kingdom best practice guidelines.

Primary mitochondrial disease describes a diverse group of neuro-metabolic disorders characterised by impaired oxidative phosphorylation. Diagnosis is challenging; >350 genes, both nuclear and mitochondrial DNA (mtDNA) encoded, are known to cause mitochondrial disease, leading to all possible inheritance patterns and further complicated by heteroplasmy of the multicopy mitochondrial genome. Technological advances, particularly next-generation sequencing, have driven a shift in diagnostic practice from 'biopsy first' to genome-wide analyses of blood and/or urine DNA. This has led to the need for a reference framework for laboratories involved in mitochondrial genetic testing to facilitate a consistent high-quality service. In the United Kingdom, consensus guidelines have been prepared by a working group of Clinical Scientists from the NHS Highly Specialised Service followed by national laboratory consultation. These guidelines summarise current recommended technologies and methodologies for the analysis of mtDNA and nuclear-encoded genes in patients with suspected mitochondrial disease. Genetic testing strategies for diagnosis, family testing and reproductive options including prenatal diagnosis are outlined. Importantly, recommendations for the minimum levels of mtDNA testing for the most common referral reasons are included, as well as guidance on appropriate referrals and information on the minimal appropriate gene content of panels when analysing nuclear mitochondrial genes. Finally, variant interpretation and recommendations for reporting of results are discussed, focussing particularly on the challenges of interpreting and reporting mtDNA variants.

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.

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.