An integrated clinical program and crowdsourcing strategy for genomic sequencing and Mendelian disease gene discovery.

Despite major progress in defining the genetic basis of Mendelian disorders, the molecular etiology of many cases remains unknown. Patients with these undiagnosed disorders often have complex presentations and require treatment by multiple health care specialists. Here, we describe an integrated clinical diagnostic and research program using whole-exome and whole-genome sequencing (WES/WGS) for Mendelian disease gene discovery. This program employs specific case ascertainment parameters, a WES/WGS computational analysis pipeline that is optimized for Mendelian disease gene discovery with variant callers tuned to specific inheritance modes, an interdisciplinary crowdsourcing strategy for genomic sequence analysis, matchmaking for additional cases, and integration of the findings regarding gene causality with the clinical management plan. The interdisciplinary gene discovery team includes clinical, computational, and experimental biomedical specialists who interact to identify the genetic etiology of the disease, and when so warranted, to devise improved or novel treatments for affected patients. This program effectively integrates the clinical and research missions of an academic medical center and affords both diagnostic and therapeutic options for patients suffering from genetic disease. It may therefore be germane to other academic medical institutions engaged in implementing genomic medicine programs

Development and Validation of Targeted Next-Generation Sequencing Panels for Detection of Germline Variants in Inherited Diseases.

Context.-The number of targeted next-generation sequencing (NGS) panels for genetic diseases offered by clinical laboratories is rapidly increasing. Before an NGS-based test is implemented in a clinical laboratory, appropriate validation studies are needed to determine the performance characteristics of the test. Objective.-To provide examples of assay design and validation of targeted NGS gene panels for the detection of germline variants associated with inherited disorders. Data Sources.-The approaches used by 2 clinical laboratories for the development and validation of targeted NGS gene panels are described. Important design and validation considerations are examined. Conclusions.-Clinical laboratories must validate performance specifications of each test prior to implementation. Test design specifications and validation data are provided, outlining important steps in validation of targeted NGS panels by clinical diagnostic laboratories

College of American Pathologists\u27 Laboratory Standards for Next-Generation Sequencing Clinical Tests

Context.-The higher throughput and lower per-base cost of next-generation sequencing (NGS) as compared to Sanger sequencing has led to its rapid adoption in clinical testing. The number of laboratories offering NGS-based tests has also grown considerably in the past few years, despite the fact that specific Clinical Laboratory Improvement Amendments of 1988/College of American Pathologists (CAP) laboratory standards had not yet been developed to regulate this technology. Objective.-To develop a checklist for clinical testing using NGS technology that sets standards for the analytic wet bench process and for bioinformatics or \u27\u27 dry bench\u27\u27 analyses. As NGS-based clinical tests are new to diagnostic testing and are of much greater complexity than traditional Sanger sequencing-based tests, there is an urgent need to develop new regulatory standards for laboratories offering these tests. Design.-To develop the necessary regulatory framework for NGS and to facilitate appropriate adoption of this technology for clinical testing, CAP formed a committee in 2011, the NGS Work Group, to deliberate upon the contents to be included in the checklist. Results.-A total of 18 laboratory accreditation checklist requirements for the analytic wet bench process and bioinformatics analysis processes have been included within CAP\u27s molecular pathology checklist (MOL). Conclusions.-This report describes the important issues considered by the CAP committee during the development of the new checklist requirements, which address documentation, validation, quality assurance, confirmatory testing, exception logs, monitoring of upgrades, variant interpretation and reporting, incidental findings, data storage, version traceability, and data transfer confidentiality

Resistance to autosomal dominant Alzheimer's disease in an APOE3 Christchurch homozygote: a case report.

We identified a PSEN1 (presenilin 1) mutation carrier from the world's largest autosomal dominant Alzheimer's disease kindred, who did not develop mild cognitive impairment until her seventies, three decades after the expected age of clinical onset. The individual had two copies of the APOE3 Christchurch (R136S) mutation, unusually high brain amyloid levels and limited tau and neurodegenerative measurements. Our findings have implications for the role of APOE in the pathogenesis, treatment and prevention of Alzheimer's disease

The clinical application of genome-wide sequencing for monogenic diseases in Canada: Position statement of the Canadian College of medical geneticists

Purpose and scope: The aim of this Position Statement is to provide recommendations for Canadian medical geneticists, clinical laboratory geneticists, genetic counsellors and other physicians regarding the use of genome-wide sequencing of germline DNA in the context of clinical genetic diagnosis. This statement has been developed to facilitate the clinical translation and development of best practices for clinical genome-wide sequencing for genetic diagnosis of monogenic diseases in Canada; it does not address the clinical application of this technology in other fields such as molecular investigation of cancer or for population screening of healthy individuals. Methods of statement development: Two multidisciplinary groups consisting of medical geneticists, clinical laboratory geneticists, genetic counsellors, ethicists, lawyers and genetic researchers were assembled to review existing literature and guidelines on genome-wide sequencing for clinical genetic diagnosis in the context of monogenic diseases, and to make recommendations relevant to the Canadian context. The statement was circulated for comment to the Canadian College of Medical Geneticists (CCMG) membership-at-large and, following incorporation of feedback, approved by the CCMG Board of Directors. The CCMG is a Canadian organisation responsible for certifying medical geneticists and clinical laboratory geneticists, and for establishing professional and ethical standards for clinical genetics services in Canada. Results and conclusions: Recommendations include (1) clinical genome-wide sequencing is an appropriate approach in the diagnostic assessment of a patient for whom there is suspicion of a significant monogenic disease that is associated with a high degree of genetic heterogeneity, or where specific genetic tests have failed to provide a diagnosis; (2) until the benefits of reporting incidental findings are established, we do not endorse the intentional clinical analysis of disease-associated genes other than those linked to the primary indication; and (3) clinicians should provide genetic counselling and obtain informed consent prior to undertaking clinical genome-wide sequencing. Counselling should include discussion of the limitations of testing, likelihood and implications of diagnosis and incidental findings, and the potential need for further analysis to facilitate clinical interpretation, including studies performed in a research setting. These recommendations will be routinely reevaluated as knowledge of diagnostic and clinical utility of clinical genome-wide sequencing improves. While the document was developed to direct practice in Canada, the applicability of the statement is broader and will be of interest to clinicians and health jurisdictions internationally

Doctor of Philosophy

dissertationSuccessful molecular diagnosis using an exome sequence hinges on accurate association of damaging variants to the patient's phenotype. Unfortunately, many clinical scenarios (e.g., single affected or small nuclear families) have little power to confidently identify damaging alleles using sequence data alone. Today's diagnostic tools are simply underpowered for accurate diagnosis in these situations, limiting successful diagnoses. In response, clinical genetics relies on candidate-gene and variant lists to limit the search space. Despite their practical utility, these lists suffer from inherent and significant limitations. The impact of false negatives on diagnostic accuracy is considerable because candidate-genes and variants lists are assembled ad hoc, choosing alleles based upon expert knowledge. Alleles not in the list are not considered-ending hope for novel discoveries. Rational alternatives to ad hoc assemblages of candidate lists are thus badly needed. In response, I created Phevor, the Phenotype Driven Variant Ontological Re-ranking tool. Phevor works by combining knowledge resident in biomedical ontologies, like the human phenotype and gene ontologies, with the outputs of variant-interpretation tools such as SIFT, GERP+, Annovar and VAAST. Phevor can then accurately to prioritize candidates identified by third-party variant-interpretation tools in light of knowledge found in the ontologies, effectively bypassing the need for candidate-gene and variant lists. Phevor differs from tools such as Phenomizer and Exomiser, as it does not postulate a set of fixed associations between genes and phenotypes. Rather, Phevor dynamically integrates knowledge resident in multiple bio-ontologies into the prioritization process. This enables Phevor to improve diagnostic accuracy for established diseases and previously undescribed or atypical phenotypes. Inserting known disease-alleles into otherwise healthy exomes benchmarked Phevor. Using the phenotype of the known disease, and the variant interpretation tool VAAST (Variant Annotation, Analysis and Search Tool), Phevor can rank 100% of the known alleles in the top 10 and 80% as the top candidate. Phevor is currently part of the pipeline used to diagnose cases as part the Utah Genome Project. Successful diagnoses of several phenotypes have proven Phevor to be a reliable diagnostic tool that can improve the analysis of any disease-gene search

Cloud-based genomics pipelines for ophthalmology: Reviewed from research to clinical practice

Aim: To familiarize clinicians with clinical genomics, and to describe the potential of cloud computing for enabling the future routine use of genomics in eye hospital settings. Design: Review article exploring the potential for cloud-based genomic pipelines in eye hospitals. Methods: Narrative review of the literature relevant to clinical genomics and cloud computing, using PubMed and Google Scholar. A broad overview of these fields is provided, followed by key examples of their integration. Results: Cloud computing could benefit clinical genomics due to scalability of resources, potentially lower costs, and ease of data sharing between multiple institutions. Challenges include complex pricing of services, costs from mistakes or experimentation, data security, and privacy concerns. Conclusions and future perspectives: Clinical genomics is likely to become more routinely used in clinical practice. Currently this is delivered in highly specialist centers. In the future, cloud computing could enable delivery of clinical genomics services in non-specialist hospital settings, in a fast, cost-effective way, whilst enhancing collaboration between clinical and research teams

The Current Landscape of Genetic Testing in Cardiovascular Malformations: Opportunities and Challenges

Human cardiovascular malformations (CVMs) frequently have a genetic contribution. Through the application of novel technologies, such as next-generation sequencing, DNA sequence variants associated with CVMs are being identified at a rapid pace. While clinicians are now able to offer testing with NGS gene panels or whole exome sequencing to any patient with a CVM, the interpretation of genetic variation remains problematic. Variable phenotypic expression, reduced penetrance, inconsistent phenotyping methods, and the lack of high-throughput functional testing of variants contribute to these challenges. This article elaborates critical issues that impact the decision to broadly implement clinical molecular genetic testing in CVMs. Major benefits of testing include establishing a genetic diagnosis, facilitating cost-effective screening of family members who may have subclinical disease, predicting recurrence risk in offsprings, enabling early diagnosis and anticipatory management of CV and non-CV disease phenotypes, predicting long-term outcomes, and facilitating the development of novel therapies aimed at disease improvement or prevention. Limitations include financial cost, psychosocial cost, and ambiguity of interpretation of results. Multiplex families and patients with syndromic features are two groups where disease causation could potentially be firmly established. However, these account for the minority of the overall CVM population, and there is increasing recognition that genotypes previously associated with syndromes also exist in patients who lack non-CV findings. In all circumstances, ongoing dialog between cardiologists and clinical geneticists will be needed to accurately interpret genetic testing and improve these patients' health. This may be most effectively implemented by the creation and support of CV genetics services at centers committed to pursuing testing for patients