The 100 000 Genomes Project: What it means for paediatrics

The 100 000 Genomes Project is a unique, national programme combining research and transformation of clinical care, by undertaking whole genome sequencing (WGS) in patients with rare diseases and cancer. Made possible by technological advances in next-generation sequencing1 and falling costs, this project aims to find the genes which cause a patient's rare disease and identify genetic changes which occur in the tumour of a child or adult with cancer, to understand the mechanism of disease and develop therapies to personalise treatment. Patients are recruited through the National Health Service (NHS) and their medical course is tracked for life through their NHS number with results fed back through routine NHS services. It will also lay the foundations for a new ‘genomic medicine’ service for the NHS.2 The project is coordinated by Genomics England, with participants enrolled through one of 13 NHS Genomic Medicine Centres (figure 1), covering all of England. Unlike genome projects in other countries3 that have yielded information on variants associated with common diseases and ancestry, the scale of the 100 000 Genomes Project is much greater. The ability to track long-term outcomes through the patients' NHS number provides a unique opportunity to link genomic and phenotypic data to hospital admissions (via hospital episode statistics) as well as lifelong response to interventions and treatments

Gentamicin, genetic variation and deafness in preterm children.

Hearing loss in children born before 32 weeks of gestation is more prevalent than in full term infants. Aminoglycoside antibiotics are routinely used to treat bacterial infections in babies on neonatal intensive care units. However, this type of medication can have harmful effects on the auditory system. In order to avoid this blood levels should be maintained in the therapeutic range. However in individuals with a mitochondrial genetic variant (m.1555A > G), permanent hearing loss can occur even when drug levels are within normal limits. The aim of the study is to investigate the burden that the m.1555A > G mutation represents to deafness in very preterm infants

Lack of significant association between mutations of KCNJ10 or FOXI1 and SLC26A4 mutations in pendred syndrome/enlarged vestibular aqueducts

Pendred syndrome is a common autosomal recessive disorder causing deafness. Features include sensorineural hearing impairment, goitre, enlarged vestibular aqueducts (EVA) and occasionally Mondini dysplasia. Hearing impairment and EVA may occur in the absence of goitre or thyroid dyshormonogensis in a condition known as non-syndromic EVA. A significant number of patients with Pendred syndrome and non-syndromic EVA show only one mutation in SLC26A4. Two genes, KCNJ10, encoding an inwardly rectifying potassium channel and FOXI1, a transcriptional factor gene, are thought to play a role in the disease phenotypes

Opening the "black box" of informed consent appointments for genome sequencing: a multisite observational study

PURPOSE: Little is known about how health-care professionals communicate with patients about consenting to genome sequencing. We therefore examined what topics health-care professionals covered and what questions patients asked during consent conversations. METHODS: Twenty-one genome sequencing consent appointments were audio recorded and analyzed. Participants were 35 individuals being invited to participate in the 100,000 Genomes Project (14 participants with rare diseases, 21 relatives), and 10 health-care professionals ("consenters"). RESULTS: Two-thirds of participants' questions were substantive (e.g., genetics and inheritance); one-third administrative (e.g., filling in the consent form). Consenters usually (19/21) emphasized participant choice about secondary findings, but less often (13/21) emphasized the uncertainty about associated disease risks. Consenters primarily used passive statements and closed-ended, rather than open-ended, questions to invite participants' questions and concerns. In two appointments, one parent expressed negative or uncertain views about secondary findings, but after discussion with the other parent opted to receive them. CONCLUSION: Health-care professionals need to be prepared to answer patients' questions about genetics to facilitate genome sequencing consent. Health-care professionals' education also needs to address how to effectively listen and elicit each patient's questions and views, and how to discuss uncertainty around the disease risks associated with secondary findings

Functional Assessment of Variants Associated with Wolfram Syndrome

Wolfram Syndrome (WS) is a heterogeneous multisystem neurodegenerative disorder with two allelic variations in addition to a separate subtype known as WS type 2. The wide phenotypic spectrum of WS includes diabetes mellitus and optic atrophy which is often accompanied by diabetes insipidus, deafness, urological and neurological complications in combination or in isolation. To date, the understanding of the genotype phenotype relationship in this complex syndrome remains poorly understood. In this study we identified and explored the functionality of rare and novel variants in the two causative WS genes WFS1 and CISD2 by assessing the effects of the mutations on the encoded proteins Wolframin and ERIS, in a cohort of 12 patients with autosomal recessive WS, dominant WS and WS type 2. The identified pathogenic variants included missense changes, frameshift deletions and insertions in WFS1 and an exonic deletion in CISD2 which all altered the respective encoded protein in a manner that did not correlate to the phenome previously described. These observations suggest the lack of genotype phenotype correlation in this complex syndrome and the need to explore other molecular genetic mechanisms. Additionally, our findings highlight the importance of functionally assessing variants for their pathogenicity to tackle the problem of increasing variants of unknown significance (VUS) in the public genetic databases

Mitochondrial m.1584A 12S m62A rRNA methylation in families with m.1555A>G associated hearing loss.

The mitochondrial DNA mutation m.1555A>G predisposes to hearing loss following aminoglycoside antibiotic exposure in an idiosyncratic dose-independent manner. However, it may also cause maternally inherited hearing loss in the absence of aminoglycoside exposure or any other clinical features (non-syndromic hearing loss). Although m.1555A>G was identified as a cause of deafness more than twenty years ago, the pathogenic mechanism of this mutation of ribosomal RNA remains controversial. Different mechanistic concepts have been proposed. Most recently, evidence from cell lines and animal models suggested that patients with m.1555A>G may have more 12S rRNA N6, N6-dimethyladenosine (m(6) 2A) methylation than controls, so-called 'hypermethylation'. This has been implicated as a pathogenic mechanism of mitochondrial dysfunction but has yet to be validated in patients. 12S m(6) 2A rRNA methylation, by the mitochondrial transcription factor 1 (TFB1M) enzyme, occurs at two successive nucleotides (m.1584A and m.1583A) in close proximity to m.1555A>G. We examined m(6) 2A methylation in 14 patients with m.1555A>G, and controls, and found all detectable 12S rRNA transcripts to be methylated in both groups. Moreover, different RNA samples derived from the same patient (lymphocyte, fibroblast and lymphoblast) revealed that only transformed cells contained some unmethylated 12S rRNA transcripts, with all detectable 12S rRNA transcripts derived from primary samples m(6) 2A-methylated. Our data indicate that TFB1M 12S m(6) 2A rRNA hypermethylation is unlikely to be a pathogenic mechanism and may be an artefact of previous experimental models studied. We propose that RNA methylation studies in experimental models should be validated in primary clinical samples to ensure that they are applicable to the human situation

Exome sequencing identifies variants in FKBP4 that are associated with recurrent fetal loss in humans

Recurrent pregnancy loss (RPL) is defined as two or more consecutive miscarriages and affects an estimated 1.5% of couples trying to conceive. RPL has been attributed to genetic, endocrine, immune and thrombophilic disorders, But many cases remain unexplained. We investigated a Bangladeshi family where the proband experienced 29 consecutive pregnancy losses with no successful pregnancies from three different marriages. Whole exome sequencing identified rare genetic variants in several candidate genes. These were further investigated in Asian and White European RPL cohorts, and in Bangladeshi controls. FKBP4, encoding the immunophilin FK506 binding protein 4, was identified as a plausible candidate, with three further novel variants identified in Asian patients. None were found in European patients or controls. In silico structural studies predicted damaging effects of the variants in the structure-function properties of the FKBP52 protein. These were located domains reported to be involved in Hsp90 binding and peptidyl-prolyl cic-trans isomerase (PPIase) activity. Profound effects on PPIase activity were demonstrated in transiently transfected HEK293 cells comparing wildtype and mutant FKBP4 constructs. Mice lacking Fkbp4 have been previously reported as infertile through implantation failure. This study therefore strongly implicates FKBP4 as associated with fetal losses in humans, particularly in the Asian population. [Abstract copyright: © The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected].

Clinical and preclinical therapeutic outcome metrics for USH2A-related disease

USH2A variants are the most common cause of Usher syndrome type 2, characterized by congenital sensorineural hearing loss and retinitis pigmentosa (RP), and also contribute to autosomal recessive non-syndromic RP. Several treatment strategies are under development; however, sensitive clinical trial endpoint metrics to determine therapeutic efficacy have not been identified. In the present study, we have performed longitudinal retrospective examination of the retinal and auditory symptoms in (i) 56 biallelic molecularly confirmed USH2A patients and (ii) ush2a mutant zebrafish to identify metrics for the evaluation of future clinical trials and rapid preclinical screening studies. The patient cohort showed a statistically significant correlation between age and both rate of constriction for the ellipsoid zone length and hyperautofluorescent outer retinal ring area. Visual acuity and pure tone audiograms are not suitable outcome measures. Retinal examination of the novel ush2au507 zebrafish mutant revealed a slowly progressive degeneration of predominantly rods, accompanied by rhodopsin and blue cone opsin mislocalization from 6 to 12 months of age with lysosome-like structures observed in the photoreceptors. This was further evaluated in the ush2armc zebrafish model, which revealed similar changes in photopigment mislocalization with elevated autophagy levels at 6 days post fertilization, indicating a more severe genotype-phenotype correlation and providing evidence of new insights into the pathophysiology underlying USH2A-retinal disease