Assessment of the incorporation of CNV surveillance into gene panel next-generation sequencing testing for inherited retinal diseases.

BACKGROUND: Diagnostic use of gene panel next-generation sequencing (NGS) techniques is commonplace for individuals with inherited retinal dystrophies (IRDs), a highly genetically heterogeneous group of disorders. However, these techniques have often failed to capture the complete spectrum of genomic variation causing IRD, including CNVs. This study assessed the applicability of introducing CNV surveillance into first-tier diagnostic gene panel NGS services for IRD. METHODS: Three read-depth algorithms were applied to gene panel NGS data sets for 550 referred individuals, and informatics strategies used for quality assurance and CNV filtering. CNV events were confirmed and reported to referring clinicians through an accredited diagnostic laboratory. RESULTS: We confirmed the presence of 33 deletions and 11 duplications, determining these findings to contribute to the confirmed or provisional molecular diagnosis of IRD for 25 individuals. We show that at least 7% of individuals referred for diagnostic testing for IRD have a CNV within genes relevant to their clinical diagnosis, and determined a positive predictive value of 79% for the employed CNV filtering techniques. CONCLUSION: Incorporation of CNV analysis increases diagnostic yield of gene panel NGS diagnostic tests for IRD, increases clarity in diagnostic reporting and expands the spectrum of known disease-causing mutations

NAA10 polyadenylation signal variants cause syndromic microphthalmia

Background A single variant in NAA10 (c.471+2T>A), the gene encoding N-acetyltransferase 10, has been associated with Lenz microphthalmia syndrome. In this study, we aimed to identify causative variants in families with syndromic X-linked microphthalmia.Methods Three families, including 15 affected individuals with syndromic X-linked microphthalmia, underwent analyses including linkage analysis, exome sequencing and targeted gene sequencing. The consequences of two identified variants in NAA10 were evaluated using quantitative PCR and RNAseq.Results Genetic linkage analysis in family 1 supported a candidate region on Xq27-q28, which included NAA10. Exome sequencing identified a hemizygous NAA10 polyadenylation signal (PAS) variant, chrX:153,195,397T>C, c.*43A>G, which segregated with the disease. Targeted sequencing of affected males from families 2 and 3 identified distinct NAA10 PAS variants, chrX:g.153,195,401T>C, c.*39A>G and chrX:g.153,195,400T>C, c.*40A>G. All three variants were absent from gnomAD. Quantitative PCR and RNAseq showed reduced NAA10 mRNA levels and abnormal 3′ UTRs in affected individuals. Targeted sequencing of NAA10 in 376 additional affected individuals failed to identify variants in the PAS.Conclusion These data show that PAS variants are the most common variant type in NAA10-associated syndromic microphthalmia, suggesting reduced RNA is the molecular mechanism by which these alterations cause microphthalmia/anophthalmia. We reviewed recognised variants in PAS associated with Mendelian disorders and identified only 23 others, indicating that NAA10 harbours more than 10% of all known PAS variants. We hypothesise that PAS in other genes harbour unrecognised pathogenic variants associated with Mendelian disorders. The systematic interrogation of PAS could improve genetic testing yi

CM-Path Molecular Diagnostics Forum-consensus statement on the development and implementation of molecular diagnostic tests in the United Kingdom.

BACKGROUND: Pathology has evolved from a purely morphological description of cellular alterations in disease to our current ability to interrogate tissues with multiple 'omics' technologies. By utilising these techniques and others, 'molecular diagnostics' acts as the cornerstone of precision/personalised medicine by attempting to match the underlying disease mechanisms to the most appropriate targeted therapy. METHODS: Despite the promises of molecular diagnostics, significant barriers have impeded its widespread clinical adoption. Thus, the National Cancer Research Institute (NCRI) Cellular Molecular Pathology (CM-Path) initiative convened a national Molecular Diagnostics Forum to facilitate closer collaboration between clinicians, academia, industry, regulators and other key stakeholders in an attempt to overcome these. RESULTS: We agreed on a consensus 'roadmap' that should be followed during development and implementation of new molecular diagnostic tests. We identified key barriers to efficient implementation and propose possible solutions to these. In addition, we discussed the recent reconfiguration of molecular diagnostic services in NHS England and its likely impacts. CONCLUSIONS: We anticipate that this consensus statement will provide practical advice to those involved in the development of novel molecular diagnostic tests. Although primarily focusing on test adoption within the United Kingdom, we also refer to international guidelines to maximise the applicability of our recommendations

Variation in health care services for specialist genetic testing and implications for planning genetic services: The example of inherited retinal dystrophy in the English NHS

This study aims to identify and quantify the extent of current variation in service provision of a genetic testing service for dominant and X-linked retinal dystrophies in the English National Health Service (NHS). National audit data (all test requests and results (n = 1839) issued between 2003 and 2011) and survey of English regional genetic testing services were used. Age- and gender-adjusted standardised testing rates were calculated using indirect standardisation, and survey responses were transcribed verbatim and data collated and summarised. The cumulative incidence rate of testing in England was 4.5 per 100,000 population for males and 2.6 per 100,000 population for females. The standardised testing rate (STR) varied widely between regions of England, being particularly low in the North-east (STR 0.485), with half as many tests as expected based on the size and demographic distribution of the population and high in the South-east (STR 1.355), with 36\ua0% more tests than expected. Substantial and significantly different rates of testing were found between regional populations. Specific policy mechanisms to promote, monitor and evaluate the regional distribution of access to genetic and genomic testing are required. However, commissioners will require information on the scope and role of genetic services and the population at risk of the conditions for which patients are tested

Phenome-wide Mendelian randomisation analysis identifies causal factors for age-related macular degeneration

Background: Age-related macular degeneration (AMD) is a leading cause of blindness in the industrialised world and is projected to affect >280 million people worldwide by 2040. Aiming to identify causal factors and potential therapeutic targets for this common condition, we designed and undertook a phenome-wide Mendelian randomisation (MR) study. Methods: We evaluated the effect of 4591 exposure traits on early AMD using univariable MR. Statistically significant results were explored further using: validation in an advanced AMD cohort; MR Bayesian model averaging (MR-BMA); and multivariable MR. Results: Overall, 44 traits were found to be putatively causal for early AMD in univariable analysis. Serum proteins that were found to have significant relationships with AMD included S100-A5 (odds ratio [OR] = 1.07, p-value = 6.80E−06), cathepsin F (OR = 1.10, p-value = 7.16E−05), and serine palmitoyltransferase 2 (OR = 0.86, p-value = 1.00E−03). Univariable MR analysis also supported roles for complement and immune cell traits. Although numerous lipid traits were found to be significantly related to AMD, MR-BMA suggested a driving causal role for serum sphingomyelin (marginal inclusion probability [MIP] = 0.76; model-averaged causal estimate [MACE] = 0.29). Conclusions: The results of this MR study support several putative causal factors for AMD and highlight avenues for future translational research. Funding: This project was funded by the Wellcome Trust (224643/Z/21/Z; 200990/Z/16/Z); the University of Manchester’s Wellcome Institutional Strategic Support Fund (Wellcome ISSF) grant (204796/Z/16/Z); the UK National Institute for Health Research (NIHR) Academic Clinical Fellow and Clinical Lecturer Programmes; Retina UK and Fight for Sight (GR586); the Australian National Health and Medical Research Council (NHMRC) (1150144)

High penetrance of myeloid neoplasia with diverse clinical and cytogenetic features in three siblings with a familial GATA2 deficiency.

From PubMed via Jisc Publications RouterHistory: received 2020-12-21, revised 2021-03-25, accepted 2021-04-06Publication status: aheadofprintPathogenic germ-line variants in GATA2 (GATA2-deficiency) can cause childhood myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML), and can be associated with distinct clinical syndromic features. However, penetrance and genotype-phenotype correlations are incompletely understood. Here we report on the clinically diverse features of three siblings affected by GATA2c.1021_1031del over an 18-year period, all initially presenting in childhood and adolescence with MDS and AML with monosomy 7 (-7), and one also with trisomy 8 (+8). The siblings inherited a GATA2c.1021_1031del from their father who remains asymptomatic in his sixth decade. The two younger sisters are well after unrelated haematopoietic stem cell transplantation (HSCT), while the first boy died of severe chronic lung disease after sibling HSCT from his youngest sister, who subsequently also developed GATA2-deficiency associated MDS. This family illustrates high penetrance with variable genotype/phenotype correlation within one generation with GATA2-deficiency. We surmise that the lung disease post sibling HSCT was also caused by the GATA2-deficiency. The experience with this family underlines the necessity for GATA2 analysis in all apparently sporadic childhood and teenage MDS and AML with -7 also in the absence of a family history or other clinical features, and rigorous genetic testing in siblings. Moreover, our findings support the arguments for pre-emptive HSCT in variant-carrying siblings. [Abstract copyright: Copyright © 2021 Elsevier Inc. All rights reserved.

Mast cell infiltration of the choroid and protease release are early events in age-related macular degeneration associated with genetic risk at both chromosomes 1q32 and 10q26.

Age-related macular degeneration (AMD) is a leading cause of visual loss. It has a strong genetic basis, and common haplotypes on chromosome (Chr) 1 (CFH Y402H variant) and on Chr10 (near HTRA1/ARMS2) contribute the most risk. Little is known about the early molecular and cellular processes in AMD, and we hypothesized that analyzing submacular tissue from older donors with genetic risk but without clinical features of AMD would provide biological insights. Therefore, we used mass spectrometry–based quantitative proteomics to compare the proteins in human submacular stromal tissue punches from donors who were homozygous for high-risk alleles at either Chr1 or Chr10 with those from donors who had protective haplotypes at these loci, all without clinical features of AMD. Additional comparisons were made with tissue from donors who were homozygous for high-risk Chr1 alleles and had early AMD. The Chr1 and Chr10 risk groups shared common changes compared with the low-risk group, particularly increased levels of mast cell–specific proteases, including tryptase, chymase, and carboxypeptidase A3. Histological analyses of submacular tissue from donors with genetic risk of AMD but without clinical features of AMD and from donors with Chr1 risk and AMD demonstrated increased mast cells, particularly the tryptase-positive/chymase-negative cells variety, along with increased levels of denatured collagen compared with tissue from low–genetic risk donors. We conclude that increased mast cell infiltration of the inner choroid, degranulation, and subsequent extracellular matrix remodeling are early events in AMD pathogenesis and represent a unifying mechanistic link between Chr1- and Chr10-mediated AMD