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Abstract
Genetic diseases disrupt the functionality of an infant's genome during fetal-neonatal adaptation and represent a leading cause of neonatal and infant mortality in the United States. Due to disease acuity, gene locus and allelic heterogeneity, and overlapping and diverse clinical phenotypes, diagnostic genome sequencing in neonatal intensive care units has required the development of methods to shorten turnaround times and improve genomic interpretation. From 2012 to 2021, 31 clinical studies documented the diagnostic and clinical utility of first-tier rapid or ultrarapid whole-genome sequencing through cost-effective identification of pathogenic genomic variants that change medical management, suggest new therapeutic strategies, and refine prognoses. Genomic diagnosis also permits prediction of reproductive recurrence risk for parents and surviving probands. Using implementation science and quality improvement, deployment of a genomic learning healthcare system will contribute to a reduction of neonatal and infant mortality through the integration of genome sequencing into best-practice neonatal intensive care.
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Affiliation(s)
- Stephen F Kingsmore
- Rady Children's Institute for Genomic Medicine, Rady Children's Hospital, San Diego, California, USA;
| | - F Sessions Cole
- Division of Newborn Medicine, Edward Mallinckrodt Department of Pediatrics, Washington University School of Medicine in St. Louis, St. Louis, Missouri, USA;
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2
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Liu J, Zheng Y, Huang J, Zhu D, Zang P, Luo Z, Yang Y, Peng Y, Xiao Z, Zhu Y, Lu X. Expanding the genotypes and phenotypes for 19 rare diseases by exome sequencing performed in pediatric intensive care unit. Hum Mutat 2021; 42:1443-1460. [PMID: 34298581 PMCID: PMC9292147 DOI: 10.1002/humu.24266] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2021] [Revised: 06/30/2021] [Accepted: 07/21/2021] [Indexed: 12/30/2022]
Abstract
Phenotypes of some rare genetic diseases are atypical and it is a challenge for pediatric intensive care units (PICUs) to diagnose and manage such patients in an emergency. In this study, we investigated 58 PICU patients (39 deceased and 19 surviving) in critical ill status or died shortly without a clear etiology. Whole exome sequencing was performed of 103 DNA samples from their families. Disease-causing single-nucleotide variants (SNVs) and copy number variants (CNVs) were identified to do genotype-phenotypes analysis. In total, 27 (46.6%) patients received a genetic diagnosis. We identified 34 pathogenic or likely pathogenic SNVs from 26 genes, which are related to at least 19 rare diseases. Each rare disease involved an isolated patient except two patients caused by the same gene ACAT1. The genotypic spectrum was expanded by 23 novel SNVs from gene MARS1, PRRT2, TBCK, TOR1A, ECE1, ARX, ZEB2, ACAT1, CPS1, VWF, NBAS, COG4, and INVS. We also identified two novel pathogenic CNVs. Phenotypes associated with respiratory, multiple congenital anomalies, neuromuscular, or metabolic disorders were the most common. Twenty patients (74.1%) accompanied severe infection, 19 patients (70.1%) died. In summary, our findings expanded the genotypes and phenotypes of 19 rare diseases from PICU with complex characteristics.
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Affiliation(s)
- Juan Liu
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
| | - Yu Zheng
- Pediatrics Research Institute of Hunan Province, Hunan Children's Hospital, Changsha, Hunan, China.,Center for Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China
| | - Jiaotian Huang
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
| | - Desheng Zhu
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
| | - Ping Zang
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
| | - Zhenqing Luo
- Pediatrics Research Institute of Hunan Province, Hunan Children's Hospital, Changsha, Hunan, China
| | - Yongjia Yang
- Pediatrics Research Institute of Hunan Province, Hunan Children's Hospital, Changsha, Hunan, China
| | - Yu Peng
- Pediatrics Research Institute of Hunan Province, Hunan Children's Hospital, Changsha, Hunan, China
| | - Zhenghui Xiao
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
| | - Yimin Zhu
- Emergency Medicine Institute of Hunan Province, Changsha, Hunan, China
| | - Xiulan Lu
- Pediatric Intensive Care Unit, Hunan Childrens Hospital, University of South China, Changsha, Hunan, China
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3
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Fang Y, Yu J, Lou J, Peng K, Zhao H, Chen J. Clinical and Genetic Spectra of Inherited Liver Disease in Children in China. Front Pediatr 2021; 9:631620. [PMID: 33763395 PMCID: PMC7982861 DOI: 10.3389/fped.2021.631620] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/20/2020] [Accepted: 01/18/2021] [Indexed: 11/18/2022] Open
Abstract
Background: Children presenting with chronic liver disease or acute liver failure often have an underlying genetic disorder. The aim of this study was to analyze the clinical and genetic spectra of inherited liver disease in children in a tertiary hospital. Methods: A total of 172 patients were classified into three groups according to their clinical presentation: cholestasis (Group A), liver enzyme elevation (Group B), and hepato/splenomegaly (Group C). Next-generation sequencing (NGS) was performed on all patients recruited in this study. The genotypic and phenotypic spectra of disease in these patients were reviewed. Results: The median age at enrollment of the 172 patients was 12.0 months (IQR: 4.9, 42.5 months), with 52.3% males and 47.7% females. The overall diagnostic rate was 55.8% (96/172) in this group. The diagnostic rates of whole-exome sequencing (WES) and targeted gene panel sequencing (TGPS) were 47.2% and 62.0%, respectively (no significant difference, p = 0.054). We identified 25 genes related to different phenotypes, including 46 novel disease-related pathogenic mutations. The diagnostic rates in the three groups were 46.0% (29/63), 48.6% (34/70), and 84.6% (33/39). ATP7B, SLC25A13, and G6PC were the top three genes related to monogenic liver disease in this study. Conclusion: WES and TGPS show similar diagnostic rates in the diagnosis of monogenic liver disease. NGS has an important role in the diagnosis of monogenetic liver disease and can provide more precise medical treatment and predict the prognosis of these diseases.
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Affiliation(s)
| | | | | | | | | | - Jie Chen
- National Clinical Research Center for Child Health, Department of Gastroenterology, The Children's Hospital, Zhejiang University School of Medicine, Hangzhou, China
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Grineski S, Morales DX, Collins T, Wilkes J, Bonkowsky JL. Geographic and Specialty Access Disparities in US Pediatric Leukodystrophy Diagnosis. J Pediatr 2020; 220:193-199. [PMID: 32143930 PMCID: PMC7186149 DOI: 10.1016/j.jpeds.2020.01.063] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/14/2019] [Revised: 01/05/2020] [Accepted: 01/29/2020] [Indexed: 01/01/2023]
Abstract
OBJECTIVE To examine disparities in the diagnosis of leukodystrophies including geographic factors and access to specialty centers. STUDY DESIGN Retrospective cohort study of pediatric patients admitted to Pediatric Health Information System hospitals. Patients with leukodystrophy were identified with International Classification of Diseases, Tenth Revision, Clinical Modification diagnostic codes for any of 4 leukodystrophies (X-linked adrenoleukodystrophy, Hurler disease, Krabbe disease, and metachromatic leukodystrophy). We used 3-level hierarchical generalized logistic modeling to predict diagnosis of a leukodystrophy based on distance traveled for hospital, neighborhood composition, urban/rural context, and access to specialty center. RESULTS We identified 501 patients with leukodystrophy. Patients seen at a leukodystrophy center of excellence hospital were 1.73 times more likely to be diagnosed than patients at non-center of excellence hospitals. Patients who traveled farther were more likely to be diagnosed than those who traveled shorter. Patients living in a Health Professionals Shortage Area zip code were 0.86 times less likely to be diagnosed than those living in a non-Health Professionals Shortage Area zip code. CONCLUSIONS Geographic factors affect the diagnosis of leukodystrophies in pediatric patients, particularly in regard to access to a center with expertise in leukodystrophies. Our findings suggest a need for improving access to pediatric specialists and possibly deploying specialists or diagnostic testing more broadly.
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Affiliation(s)
- Sara Grineski
- Department of Sociology, University of Utah, Salt Lake City, Utah
| | - Danielle X. Morales
- Department of Sociology & Anthropology. University of Texas at El Paso, El Paso, Texas
| | - Timothy Collins
- Department of Geography, University of Utah, Salt Lake City, Utah
| | | | - Joshua L. Bonkowsky
- Division of Pediatric Neurology, Department of Pediatrics, University of Utah School of Medicine, Salt Lake City, Utah,Brain and Spine Center, Primary Children’s Hospital, Salt Lake City, Utah,Primary Children’s Center for Personalized Medicine, Salt Lake City, Utah,Address correspondence to: Josh Bonkowsky, Department of Pediatrics, 295 Chipeta Way/Williams Building, Salt Lake City, Utah 84108, , Phone: 801-581-6756, Fax: 801-581-4233
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Titerlea V, Dembélé D, Mandel JL, Laporte J. Attitudes towards Genetic Information Delivered by High-Throughput Sequencing among Molecular Geneticists, Genetic Counselors, Medical Advisors and Students in France. Eur J Med Genet 2019; 63:103770. [PMID: 31536829 DOI: 10.1016/j.ejmg.2019.103770] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2019] [Revised: 08/10/2019] [Accepted: 09/15/2019] [Indexed: 11/16/2022]
Abstract
High-throughput sequencing technologies performed in the clinical setting have the potential to reveal diverse genetic information. Whether it is initially targeted or unsolicited, strictly medical or not, or even information on a carrier status as part of preconception screening, access to genetic information needs to be managed. The aim of the current study was to gather potential attitudes of various stakeholders towards the sharing of genetic information from next-generation sequencing, and more specifically towards incidental findings, predictive findings, non-medical information and carrier status. Answers from a total number of 1631 individuals belonging to four different groups (45 molecular geneticists, 65 genetic counselors, 56 medical advisors to the state insurance plan, and 1465 university students) were collected through online questionnaires. Overall, the study reflects preferences towards the return of health risks related to serious diseases when effective treatment is available and information on reproductive risks. The importance of the perceived medical utility, both for disease prevention and treatment, was the main distinguishing feature. Attitudes from genetic health professionals were found more reluctant to receive a wide range of information. Hands-on experience with the practice of genetic testing is likely to influence perception of the utility of the genetic information that should be delivered. At the same time, perceptions of preconception genetic carrier screening brought out less differences between participants. Better understanding of the underlying interest in genomic information and thorough education on its value and usage are key elements to the adoption of future guidelines and policy that respect bioethical principles.
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Affiliation(s)
- Vlad Titerlea
- Centre Européen d'Enseignement et de Recherche en Éthique (CEERE), University of Strasbourg, 67000, Strasbourg, France; Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U1258, CNRS UMR7104, University of Strasbourg, 67400, Illkirch, France
| | - Doulaye Dembélé
- Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U1258, CNRS UMR7104, University of Strasbourg, 67400, Illkirch, France
| | - Jean-Louis Mandel
- Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U1258, CNRS UMR7104, University of Strasbourg, 67400, Illkirch, France.
| | - Jocelyn Laporte
- Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U1258, CNRS UMR7104, University of Strasbourg, 67400, Illkirch, France
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6
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The value of diagnostic testing for parents of children with rare genetic diseases. Genet Med 2019; 21:2798-2806. [PMID: 31239560 DOI: 10.1038/s41436-019-0583-1] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2019] [Accepted: 06/01/2019] [Indexed: 02/06/2023] Open
Abstract
PURPOSE Exome sequencing (ES) can rapidly identify disease-causing variants responsible for rare, single-gene diseases, and potentially reduce the duration of the diagnostic odyssey. Our study examines how parents and families value ES. METHODS We developed a discrete choice experiment (DCE) survey that was administered to parents of children with rare diseases. The DCE included 14 choice tasks with 6 attributes and 3 alternatives. A valuation-space model was used to estimate willingness to pay, willingness to wait for test results, and minimum acceptable chance of a diagnosis for changes in each attribute. RESULTS There were n = 319 respondents of whom 89% reported their child had genetic testing, and 66% reported their child had a diagnosis. Twenty-six percent reported that their child had been offered ES. Parents were willing to pay CAD$6590 (US$4943), wait 5.2 years to obtain diagnostic test results, and accept a reduction of 3.1% in the chance of a diagnosis for ES compared with operative procedures. CONCLUSION Timely access to ES could reduce the diagnostic odyssey and associated costs. Before ES is incorporated routinely into care for patients with rare diseases in Canada and more broadly, there must be a clear understanding of its value to patients and families.
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Wang W, Corominas R, Lin GN. De novo Mutations From Whole Exome Sequencing in Neurodevelopmental and Psychiatric Disorders: From Discovery to Application. Front Genet 2019; 10:258. [PMID: 31001316 PMCID: PMC6456656 DOI: 10.3389/fgene.2019.00258] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2018] [Accepted: 03/08/2019] [Indexed: 12/13/2022] Open
Abstract
Neurodevelopmental and psychiatric disorders are a highly disabling and heterogeneous group of developmental and mental disorders, resulting from complex interactions of genetic and environmental risk factors. The nature of multifactorial traits and the presence of comorbidity and polygenicity in these disorders present challenges in both disease risk identification and clinical diagnoses. The genetic component has been firmly established, but the identification of all the causative variants remains elusive. The development of next-generation sequencing, especially whole exome sequencing (WES), has greatly enriched our knowledge of the precise genetic alterations of human diseases, including brain-related disorders. In particular, the extensive usage of WES in research studies has uncovered the important contribution of de novo mutations (DNMs) to these disorders. Trio and quad familial WES are a particularly useful approach to discover DNMs. Here, we review the major WES studies in neurodevelopmental and psychiatric disorders and summarize how genes hit by discovered DNMs are shared among different disorders. Next, we discuss different integrative approaches utilized to interrogate DNMs and to identify biological pathways that may disrupt brain development and shed light on our understanding of the genetic architecture underlying these disorders. Lastly, we discuss the current state of the transition from WES research to its routine clinical application. This review will assist researchers and clinicians in the interpretation of variants obtained from WES studies, and highlights the need to develop consensus analytical protocols and validated lists of genes appropriate for clinical laboratory analysis, in order to reach the growing demands.
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Affiliation(s)
- Weidi Wang
- Shanghai Mental Health Center, School of Biomedical Engineering, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- Shanghai Key Laboratory of Psychotic Disorders, Shanghai Mental Health Center, Shanghai, China
- Brain Science and Technology Research Center, Shanghai Jiao Tong University, Shanghai, China
| | - Roser Corominas
- Departament de Genètica, Microbiologia i Estadística, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Raras, Valencia, Spain
- Institut de Biomedicina de la Universitat de Barcelona, Barcelona, Spain
- Institut de Recerca Sant Joan de Déu, Esplugues de Llobregat, Barcelona, Spain
| | - Guan Ning Lin
- Shanghai Mental Health Center, School of Biomedical Engineering, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- Shanghai Key Laboratory of Psychotic Disorders, Shanghai Mental Health Center, Shanghai, China
- Brain Science and Technology Research Center, Shanghai Jiao Tong University, Shanghai, China
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Zech M, Wagner M, Schormair B, Oexle K, Winkelmann J. [Exome diagnostics in neurology]. DER NERVENARZT 2019; 90:131-137. [PMID: 30645660 DOI: 10.1007/s00115-018-0667-1] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
After an impressively successful application as a research instrument, whole-exome sequencing (WES) now enters the clinical practice due to its high diagnostic, time, and economic efficiency. WES is the diagnostic method of choice for symptoms that may be due to many different monogenic causes. Neurological indications include movement disorders, especially in cases of early symptom onset, familial clustering and complex manifestation. Starting from a blood sample, enrichment and sequencing of the exome enable the examination of all coding DNA regions for point mutations and small insertions/deletions. The identification of variants as the cause of a disease requires a professional evaluation pipeline, variant prioritization schemes and variant classification databases. Whereas many variants can be reliably classified as pathogenic or benign, variants of unclear significance (VUS) remain a challenge for the clinical evaluation and necessitate a periodic reanalysis of WES data. As a genetic examination WES requires adequate patient informed consent which in particular should address possible secondary findings as well as data security. A positive molecular result ends diagnostic odysseys, enables accurate genetic counseling and can point to targeted preventive measures and treatment. A WES significantly contributes to the understanding of the genetic architecture and pathophysiology of neurological diseases, enriching and enabling precision medicine.
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Affiliation(s)
- Michael Zech
- Institut für Neurogenomik, Helmholtz Zentrum München, München, Deutschland. .,Institut für Humangenetik, Klinikum rechts der Isar, Technische Universität München, Ismaninger Straße 22, 81675, München, Deutschland.
| | - Matias Wagner
- Institut für Neurogenomik, Helmholtz Zentrum München, München, Deutschland.,Institut für Humangenetik, Klinikum rechts der Isar, Technische Universität München, Ismaninger Straße 22, 81675, München, Deutschland
| | - Barbara Schormair
- Institut für Neurogenomik, Helmholtz Zentrum München, München, Deutschland
| | - Konrad Oexle
- Institut für Neurogenomik, Helmholtz Zentrum München, München, Deutschland
| | - Juliane Winkelmann
- Institut für Neurogenomik, Helmholtz Zentrum München, München, Deutschland.,Institut für Humangenetik, Klinikum rechts der Isar, Technische Universität München, Ismaninger Straße 22, 81675, München, Deutschland
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Prokop JW, May T, Strong K, Bilinovich SM, Bupp C, Rajasekaran S, Worthey EA, Lazar J. Genome sequencing in the clinic: the past, present, and future of genomic medicine. Physiol Genomics 2018; 50:563-579. [PMID: 29727589 PMCID: PMC6139636 DOI: 10.1152/physiolgenomics.00046.2018] [Citation(s) in RCA: 49] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Genomic sequencing has undergone massive expansion in the past 10 yr, from a rarely used research tool into an approach that has broad applications in a clinical setting. From rare disease to cancer, genomics is transforming our knowledge of biology. The transition from targeted gene sequencing, to whole exome sequencing, to whole genome sequencing has only been made possible due to rapid advancements in technologies and informatics that have plummeted the cost per base of DNA sequencing and analysis. The tools of genomics have resolved the etiology of disease for previously undiagnosable conditions, identified cancer driver gene variants, and have impacted the understanding of pathophysiology for many diseases. However, this expansion of use has also highlighted research's current voids in knowledge. The lack of precise animal models for gene-to-function association, lack of tools for analysis of genomic structural changes, skew in populations used for genetic studies, publication biases, and the "Unknown Proteome" all contribute to voids needing filled for genomics to work in a fast-paced clinical setting. The future will hold the tools to fill in these voids, with new data sets and the continual development of new technologies allowing for expansion of genomic medicine, ushering in the days to come for precision medicine. In this review we highlight these and other points in hopes of advancing and guiding precision medicine into the future for optimal success.
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Affiliation(s)
- Jeremy W Prokop
- HudsonAlpha Institute for Biotechnology , Huntsville, Alabama
- Department of Pediatrics and Human Development, Michigan State University , East Lansing, Michigan
- Department of Pharmacology and Toxicology, Michigan State University , East Lansing, Michigan
| | - Thomas May
- HudsonAlpha Institute for Biotechnology , Huntsville, Alabama
- Institute for Health and Aging, University of California San Francisco , San Francisco, California
- Elson S. Floyd College of Medicine, Washington State University , Spokane, Washington
| | - Kim Strong
- HudsonAlpha Institute for Biotechnology , Huntsville, Alabama
| | - Stephanie M Bilinovich
- Department of Pediatrics and Human Development, Michigan State University , East Lansing, Michigan
| | - Caleb Bupp
- Department of Pediatrics and Human Development, Michigan State University , East Lansing, Michigan
- Department of Genetics, Helen DeVos Children's Hospital, Spectrum Health, Grand Rapids, Michigan
| | - Surender Rajasekaran
- Department of Pediatrics and Human Development, Michigan State University , East Lansing, Michigan
- Department of Pediatric Critical Care Medicine, Helen DeVos Children's Hospital, Spectrum Health, Grand Rapids, Michigan
| | | | - Jozef Lazar
- HudsonAlpha Institute for Biotechnology , Huntsville, Alabama
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Venugopal A, Chandran M, Eruppakotte N, Kizhakkillach S, Breezevilla SC, Vellingiri B. Monogenic diseases in India. MUTATION RESEARCH-REVIEWS IN MUTATION RESEARCH 2018; 776:23-31. [PMID: 29807575 DOI: 10.1016/j.mrrev.2018.03.003] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/28/2017] [Revised: 03/16/2018] [Accepted: 03/16/2018] [Indexed: 12/12/2022]
Abstract
Studies on monogenic diseases are considered valuable because they give insights and expand our knowledge on gene function and regulation. Despite all the current advancement in science and technology, a deep understanding and knowledge as to why only those particular genes are affected in a disease is still vague. We also lack profound illumination as to why only certain mutations are seen in a disease. Though useful from a research perspective, a majority of these diseases are lethal resulting in death of the affected individual. Unfortunately, in the fast - growing land of India, the incidence of monogenic diseases is very high with few counter-measures in place. This article encompasses a list of all monogenic diseases ever to be reported in India with special focus on five diseases which has been stated to have the highest incidence in India. Here, we discuss about the limited research carried out in India on these high incidence monogenic diseases, the other diseases related to those genes, the range of treatments available for these diseases in India in contrast to its availability around the world and the need to develop treatment strategies to reduce the mortality and morbidity due to these rare but daunting diseases.
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Affiliation(s)
- Anila Venugopal
- Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641 046, Tamil Nadu, India.
| | - Manojkumar Chandran
- Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641 046, Tamil Nadu, India
| | - Nimmisha Eruppakotte
- Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641 046, Tamil Nadu, India
| | - Soumya Kizhakkillach
- Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641 046, Tamil Nadu, India
| | - Sanuj C Breezevilla
- Post Graduate & Research Department of Zoology, Sree Narayana College, Cherthala, 688582, Kerala, India
| | - Balachandar Vellingiri
- Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641 046, Tamil Nadu, India.
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11
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Nicastro E, D'Antiga L. Next generation sequencing in pediatric hepatology and liver transplantation. Liver Transpl 2018; 24:282-293. [PMID: 29080241 DOI: 10.1002/lt.24964] [Citation(s) in RCA: 43] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/27/2017] [Revised: 09/04/2017] [Accepted: 10/18/2017] [Indexed: 02/07/2023]
Abstract
Next generation sequencing (NGS) has revolutionized the analysis of human genetic variations, offering a highly cost-effective way to diagnose monogenic diseases (MDs). Because nearly half of the children with chronic liver disorders have a genetic cause and approximately 20% of pediatric liver transplantations are performed in children with MDs, NGS offers the opportunity to significantly improve the diagnostic yield in this field. Among the NGS strategies, the use of targeted gene panels has proven useful to rapidly and reliably confirm a clinical suspicion, whereas the whole exome sequencing (WES) with variants filtering has been adopted to assist the diagnostic workup in unclear clinical scenarios. WES is powerful but challenging because it detects a great number of variants of unknown significance that can be misinterpreted and lead to an incorrect diagnosis. In pediatric hepatology, targeted NGS can be very valuable to discriminate neonatal/infantile cholestatic disorders, disclose genetic causes of acute liver failure, and diagnose the subtype of inborn errors of metabolism presenting with a similar phenotype (such as glycogen storage disorders, mitochondrial cytopathies, or nonalcoholic fatty liver disease). The inclusion of NGS in diagnostic processes will lead to a paradigm shift in medicine, changing our approach to the patient as well as our understanding of factors affecting genotype-phenotype match. In this review, we discuss the opportunities and the challenges offered nowadays by NGS, and we propose a novel algorithm for cholestasis of infancy adopted in our center, including targeted NGS as a pivotal tool for the diagnosis of liver-based MDs. Liver Transplantation 24 282-293 2018 AASLD.
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Affiliation(s)
- Emanuele Nicastro
- Pediatric Hepatology, Gastroenterology and Transplantation, Hospital Papa Giovanni XXIII, Bergamo, Italy
| | - Lorenzo D'Antiga
- Pediatric Hepatology, Gastroenterology and Transplantation, Hospital Papa Giovanni XXIII, Bergamo, Italy
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12
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González-Porras J, Jiménez C, Benito R, Ordoñez GR, Álvarez-Román M, Fontecha ME, Janusz K, Castillo D, Fisac R, García-Frade L, Aguilar C, Martínez P, Bermejo N, Herrero S, Balanzategui A, Martin-Antorán J, Ramos R, Cebeiro M, Pardal E, Aguilera C, Pérez-Gutierrez B, Prieto M, Riesco S, Mendoza M, Benito A, Benito-Sendin A, Jimenez-Yuste V, Hernández-Rivas J, García-Sanz R, González-Díaz M, Sarasquete M, Bastida J. Application of a molecular diagnostic algorithm for haemophilia A and B using next-generation sequencing of entire F8, F9 and VWF genes. Thromb Haemost 2017; 117:66-74. [DOI: 10.1160/th16-05-0375] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2016] [Accepted: 09/17/2016] [Indexed: 12/30/2022]
Abstract
SummaryCurrently, molecular diagnosis of haemophilia A and B (HA and HB) highlights the excess risk-inhibitor development associated with specific mutations, and enables carrier testing of female relatives and prenatal or preimplantation genetic diagnosis. Molecular testing for HA also helps distinguish it from von Willebrand disease (VWD). Next-generation sequencing (NGS) allows simultaneous investigation of several complete genes, even though they may span very extensive regions. This study aimed to evaluate the usefulness of a molecular algorithm employing an NGS approach for sequencing the complete F8, F9 and VWF genes. The proposed algorithm includes the detection of inversions of introns 1 and 22, an NGS custom panel (the entire F8, F9 and VWF genes), and multiplex ligation-dependent probe amplification (MLPA) analysis. A total of 102 samples (97 FVIII- and FIX-deficient patients, and five female carriers) were studied. IVS-22 screening identified 11 out of 20 severe HA patients and one female carrier. IVS-1 analysis did not reveal any alterations. The NGS approach gave positive results in 88 cases, allowing the differential diagnosis of mild/moderate HA and VWD in eight cases. MLPA confirmed one large exon deletion. Only one case did have no pathogenic variants. The proposed algorithm had an overall success rate of 99 %. In conclusion, our evaluation demonstrates that this algorithm can reliably identify pathogenic variants and diagnose patients with HA, HB or VWD.Supplementary Material to this article is available online at www.thrombosis-online.com.
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Dimitriadou E, Melotte C, Debrock S, Esteki MZ, Dierickx K, Voet T, Devriendt K, de Ravel T, Legius E, Peeraer K, Meuleman C, Vermeesch JR. Principles guiding embryo selection following genome-wide haplotyping of preimplantation embryos. Hum Reprod 2017; 32:687-697. [PMID: 28158716 DOI: 10.1093/humrep/dex011] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2016] [Accepted: 01/13/2017] [Indexed: 12/17/2022] Open
Abstract
STUDY QUESTION How to select and prioritize embryos during PGD following genome-wide haplotyping? SUMMARY ANSWER In addition to genetic disease-specific information, the embryo selected for transfer is based on ranking criteria including the existence of mitotic and/or meiotic aneuploidies, but not carriership of mutations causing recessive disorders. WHAT IS KNOWN ALREADY Embryo selection for monogenic diseases has been mainly performed using targeted disease-specific assays. Recently, these targeted approaches are being complemented by generic genome-wide genetic analysis methods such as karyomapping or haplarithmisis, which are based on genomic haplotype reconstruction of cell(s) biopsied from embryos. This provides not only information about the inheritance of Mendelian disease alleles but also about numerical and structural chromosome anomalies and haplotypes genome-wide. Reflections on how to use this information in the diagnostic laboratory are lacking. STUDY DESIGN, SIZE, DURATION We present the results of the first 101 PGD cycles (373 embryos) using haplarithmisis, performed in the Centre for Human Genetics, UZ Leuven. The questions raised were addressed by a multidisciplinary team of clinical geneticist, fertility specialists and ethicists. PARTICIPANTS/MATERIALS, SETTING, METHODS Sixty-three couples enrolled in the genome-wide haplotyping-based PGD program. Families presented with either inherited genetic variants causing known disorders and/or chromosomal rearrangements that could lead to unbalanced translocations in the offspring. MAIN RESULTS AND THE ROLE OF CHANCE Embryos were selected based on the absence or presence of the disease allele, a trisomy or other chromosomal abnormality leading to known developmental disorders. In addition, morphologically normal Day 5 embryos were prioritized for transfer based on the presence of other chromosomal imbalances and/or carrier information. LIMITATIONS, REASONS FOR CAUTION Some of the choices made and principles put forward are specific for cleavage-stage-based genetic testing. The proposed guidelines are subject to continuous update based on the accumulating knowledge from the implementation of genome-wide methods for PGD in many different centers world-wide as well as the results of ongoing scientific research. WIDER IMPLICATIONS OF THE FINDINGS Our embryo selection principles have a profound impact on the organization of PGD operations and on the information that is transferred among the genetic unit, the fertility clinic and the patients. These principles are also important for the organization of pre- and post-counseling and influence the interpretation and reporting of preimplantation genotyping results. As novel genome-wide approaches for embryo selection are revolutionizing the field of reproductive genetics, national and international discussions to set general guidelines are warranted. STUDY FUNDING/COMPETING INTEREST(S) The European Union's Research and Innovation funding programs FP7-PEOPLE-2012-IAPP SARM: 324509 and Horizon 2020 WIDENLIFE: 692065 to J.R.V., T.V., E.D. and M.Z.E. J.R.V., T.V. and M.Z.E. have patents ZL910050-PCT/EP2011/060211-WO/2011/157846 ('Methods for haplotyping single cells') with royalties paid and ZL913096-PCT/EP2014/068315-WO/2015/028576 ('Haplotyping and copy-number typing using polymorphic variant allelic frequencies') with royalties paid, licensed to Cartagenia (Agilent technologies). J.R.V. also has a patent ZL91 2076-PCT/EP20 one 3/070858 ('High throughout genotyping by sequencing') with royalties paid. TRIAL REGISTRATION NUMBER N/A.
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Affiliation(s)
- Eftychia Dimitriadou
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Cindy Melotte
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Sophie Debrock
- University Hospitals Leuven, Leuven University Fertility Center, Herestraat 49, 3000 Leuven, Belgium
| | - Masoud Zamani Esteki
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Kris Dierickx
- Centre for Biomedical Ethics and Law, KU Leuven, 3000 Leuven, Belgium
| | - Thierry Voet
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium.,Single-cell Genomics Centre, Welcome Trust Sanger Institute, Hinxton CB10 1SA, UK
| | - Koen Devriendt
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Thomy de Ravel
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Eric Legius
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
| | - Karen Peeraer
- University Hospitals Leuven, Leuven University Fertility Center, Herestraat 49, 3000 Leuven, Belgium
| | - Christel Meuleman
- University Hospitals Leuven, Leuven University Fertility Center, Herestraat 49, 3000 Leuven, Belgium
| | - Joris Robert Vermeesch
- Department of Human Genetics, Centre for Human Genetics, University Hospitals Leuven, O&N I Herestraat 49 - box 602, KU Leuven, 3000 Leuven, Belgium
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Rabbani B, Nakaoka H, Akhondzadeh S, Tekin M, Mahdieh N. Next generation sequencing: implications in personalized medicine and pharmacogenomics. MOLECULAR BIOSYSTEMS 2017; 12:1818-30. [PMID: 27066891 DOI: 10.1039/c6mb00115g] [Citation(s) in RCA: 64] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
A breakthrough in next generation sequencing (NGS) in the last decade provided an unprecedented opportunity to investigate genetic variations in humans and their roles in health and disease. NGS offers regional genomic sequencing such as whole exome sequencing of coding regions of all genes, as well as whole genome sequencing. RNA-seq offers sequencing of the entire transcriptome and ChIP-seq allows for sequencing the epigenetic architecture of the genome. Identifying genetic variations in individuals can be used to predict disease risk, with the potential to halt or retard disease progression. NGS can also be used to predict the response to or adverse effects of drugs or to calculate appropriate drug dosage. Such a personalized medicine also provides the possibility to treat diseases based on the genetic makeup of the patient. Here, we review the basics of NGS technologies and their application in human diseases to foster human healthcare and personalized medicine.
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Affiliation(s)
- Bahareh Rabbani
- Cardiogenetic Research Center, Rajaie Cardiovascular Medical and Research Center, Iran University of Medical Sciences, Niayesh-Vali asr Intersection, Tehran, Iran.
| | - Hirofumi Nakaoka
- Division of Human Genetics, Department of Integrated Genetics, National Institute of Genetics, Yata 1111, Mishima, Shizuoka 411-8540, Japan
| | - Shahin Akhondzadeh
- Psychiatric Research Center, Roozbeh Psychiatric Hospital, Tehran University of Medical Sciences, Tehran, Iran
| | - Mustafa Tekin
- John P Hussman Institute for Human Genomics, Miller School of Medicine, University of Miami, Miami, FL, USA
| | - Nejat Mahdieh
- Cardiogenetic Research Center, Rajaie Cardiovascular Medical and Research Center, Iran University of Medical Sciences, Niayesh-Vali asr Intersection, Tehran, Iran.
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15
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Sommen M, Wuyts W, Van Camp G. Molecular diagnostics for hereditary hearing loss in children. Expert Rev Mol Diagn 2017; 17:751-760. [PMID: 28593790 DOI: 10.1080/14737159.2017.1340834] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
INTRODUCTION Hearing loss (HL) is the most common birth defect in industrialized countries with far-reaching social, psychological and cognitive implications. It is an extremely heterogeneous disease, complicating molecular testing. The introduction of next-generation sequencing (NGS) has resulted in great progress in diagnostics allowing to study all known HL genes in a single assay. The diagnostic yield is currently still limited, but has the potential to increase substantially. Areas covered: In this review the utility of NGS and the problems for comprehensive molecular testing for HL are evaluated and discussed. Expert commentary: Different publications have proven the appropriateness of NGS for molecular testing of heterogeneous diseases such as HL. However, several problems still exist, such as pseudogenic background of some genes and problematic copy number variant analysis on targeted NGS data. Another main challenge for the future will be the establishment of population specific mutation-spectra to achieve accurate personalized comprehensive molecular testing for HL.
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Affiliation(s)
- Manou Sommen
- a Center of Medical Genetics , University of Antwerp & Antwerp University Hospital , Antwerp , Belgium
| | - Wim Wuyts
- a Center of Medical Genetics , University of Antwerp & Antwerp University Hospital , Antwerp , Belgium
| | - Guy Van Camp
- a Center of Medical Genetics , University of Antwerp & Antwerp University Hospital , Antwerp , Belgium
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16
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Multi-drug resistant Klebsiella pneumoniae strains circulating in hospital setting: whole-genome sequencing and Bayesian phylogenetic analysis for outbreak investigations. Sci Rep 2017; 7:3534. [PMID: 28615687 PMCID: PMC5471223 DOI: 10.1038/s41598-017-03581-4] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2016] [Accepted: 05/09/2017] [Indexed: 01/12/2023] Open
Abstract
Carbapenems resistant Enterobacteriaceae infections are increasing worldwide representing an emerging public health problem. The application of phylogenetic and phylodynamic analyses to bacterial whole genome sequencing (WGS) data have become essential in the epidemiological surveillance of multi-drug resistant nosocomial pathogens. Between January 2012 and February 2013, twenty-one multi-drug resistant K. pneumoniae strains, were collected from patients hospitalized among different wards of the University Hospital Campus Bio-Medico. Epidemiological contact tracing of patients and Bayesian phylogenetic analysis of bacterial WGS data were used to investigate the evolution and spatial dispersion of K. pneumoniae in support of hospital infection control. The epidemic curve of incident K. pneumoniae cases showed a bimodal distribution of cases with two peaks separated by 46 days between November 2012 and January 2013. The time-scaled phylogeny suggested that K. pneumoniae strains isolated during the study period may have been introduced into the hospital setting as early as 2007. Moreover, the phylogeny showed two different epidemic introductions in 2008 and 2009. Bayesian genomic epidemiology is a powerful tool that promises to improve the surveillance and control of multi-drug resistant pathogens in an effort to develop effective infection prevention in healthcare settings or constant strains reintroduction.
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17
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Plöthner M, Frank M, von der Schulenburg JMG. Cost analysis of whole genome sequencing in German clinical practice. THE EUROPEAN JOURNAL OF HEALTH ECONOMICS : HEPAC : HEALTH ECONOMICS IN PREVENTION AND CARE 2017; 18:623-633. [PMID: 27380512 DOI: 10.1007/s10198-016-0815-0] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/06/2015] [Accepted: 06/23/2016] [Indexed: 05/23/2023]
Abstract
OBJECTIVES Whole genome sequencing (WGS) is an emerging tool in clinical diagnostics. However, little has been said about its procedure costs, owing to a dearth of related cost studies. This study helps fill this research gap by analyzing the execution costs of WGS within the setting of German clinical practice. METHODOLOGY First, to estimate costs, a sequencing process related to clinical practice was undertaken. Once relevant resources were identified, a quantification and monetary evaluation was conducted using data and information from expert interviews with clinical geneticists, and personnel at private enterprises and hospitals. This study focuses on identifying the costs associated with the standard sequencing process, and the procedure costs for a single WGS were analyzed on the basis of two sequencing platforms-namely, HiSeq 2500 and HiSeq Xten, both by Illumina, Inc. In addition, sensitivity analyses were performed to assess the influence of various uses of sequencing platforms and various coverage values on a fixed-cost degression. RESULTS In the base case scenario-which features 80 % utilization and 30-times coverage-the cost of a single WGS analysis with the HiSeq 2500 was estimated at €3858.06. The cost of sequencing materials was estimated at €2848.08; related personnel costs of €396.94 and acquisition/maintenance costs (€607.39) were also found. In comparison, the cost of sequencing that uses the latest technology (i.e., HiSeq Xten) was approximately 63 % cheaper, at €1411.20. CONCLUSIONS The estimated costs of WGS currently exceed the prediction of a 'US$1000 per genome', by more than a factor of 3.8. In particular, the material costs in themselves exceed this predicted cost.
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Affiliation(s)
- Marika Plöthner
- Center for Health Economics Research Hannover (CHERH), Leibniz University Hannover, Otto-Brenner-Straße 1, 30159, Hannover, Germany.
| | - Martin Frank
- Center for Health Economics Research Hannover (CHERH), Leibniz University Hannover, Otto-Brenner-Straße 1, 30159, Hannover, Germany
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18
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Smith M. DNA Sequence Analysis in Clinical Medicine, Proceeding Cautiously. Front Mol Biosci 2017; 4:24. [PMID: 28516087 PMCID: PMC5413496 DOI: 10.3389/fmolb.2017.00024] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2017] [Accepted: 04/07/2017] [Indexed: 12/03/2022] Open
Abstract
Delineation of underlying genomic and genetic factors in a specific disease may be valuable in establishing a definitive diagnosis and may guide patient management and counseling. In addition, genetic information may be useful in identification of at risk family members. Gene mapping and initial genome sequencing data enabled the development of microarrays to analyze genomic variants. The goal of this review is to consider different generations of sequencing techniques and their application to exome sequencing and whole genome sequencing and their clinical applications. In recent decades, exome sequencing has primarily been used in patient studies. Discussed in some detail, are important measures that have been developed to standardize variant calling and to assess pathogenicity of variants. Examples of cases where exome sequencing has facilitated diagnosis and led to improved medical management are presented. Whole genome sequencing and its clinical relevance are presented particularly in the context of analysis of nucleotide and structural genomic variants in large population studies and in certain patient cohorts. Applications involving analysis of cell free DNA in maternal blood for prenatal diagnosis of specific autosomal trisomies are reviewed. Applications of DNA sequencing to diagnosis and therapeutics of cancer are presented. Also discussed are important recent diagnostic applications of DNA sequencing in cancer, including analysis of tumor derived cell free DNA and exosomes that are present in body fluids. Insights gained into underlying pathogenetic mechanisms of certain complex common diseases, including schizophrenia, macular degeneration, neurodegenerative disease are presented. The relevance of different types of variants, rare, uncommon, and common to disease pathogenesis, and the continuum of causality, are addressed. Pharmogenetic variants detected by DNA sequence analysis are gaining in importance and are particularly relevant to personalized and precision medicine.
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Affiliation(s)
- Moyra Smith
- Genetics and Genomic Medicine, Pediatrics, School of Medicine, University of CaliforniaIrvine, CA, USA
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19
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Lacoste C, Fabre A, Pécheux C, Lévy N, Krahn M, Malzac P, Bonello-Palot N, Badens C, Bourgeois P. Le séquençage d’ADN à haut débit en pratique clinique. Arch Pediatr 2017; 24:373-383. [DOI: 10.1016/j.arcped.2017.01.008] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2016] [Revised: 01/03/2017] [Accepted: 01/03/2017] [Indexed: 12/22/2022]
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20
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Pediatric Whole Exome Sequencing: an Assessment of Parents’ Perceived and Actual Understanding. J Genet Couns 2016; 26:792-805. [DOI: 10.1007/s10897-016-0052-9] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2016] [Accepted: 11/27/2016] [Indexed: 01/10/2023]
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Yang W, Wu G, Broeckel U, Smith CA, Turner V, Haidar CE, Wang S, Carter R, Karol SE, Neale G, Crews KR, Yang JJ, Mullighan CG, Downing JR, Evans WE, Relling MV. Comparison of genome sequencing and clinical genotyping for pharmacogenes. Clin Pharmacol Ther 2016; 100:380-8. [PMID: 27311679 DOI: 10.1002/cpt.411] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2016] [Revised: 05/23/2016] [Accepted: 06/13/2016] [Indexed: 12/28/2022]
Abstract
We compared whole exome sequencing (WES, n = 176 patients) and whole genome sequencing (WGS, n = 68) and clinical genotyping (DMET array-based approach) for interrogating 13 genes with Clinical Pharmacogenetics Implementation Consortium (CPIC) guidelines. We focused on 127 CPIC important variants: 103 single nucleotide variations (SNV), 21 insertion/deletions (Indel), HLA-B alleles, and two CYP2D6 structural variations. WES and WGS provided interrogation of nonoverlapping sets of 115 SNV/Indels with call rate >98%. Among 68 loci interrogated by both WES and DMET, 64 loci (94.1%, confidence interval [CI]: 85.6-98.4%) showed no discrepant genotyping calls. Among 66 loci interrogated by both WGS and DMET, 63 loci (95.5%, CI: 87.2-99.0%) showed no discrepant genotyping calls. In conclusion, even without optimization to interrogate pharmacogenetic variants, WES and WGS displayed potential to provide reliable interrogation of most pharmacogenes and further validation of genome sequencing in a clinical lab setting is warranted.
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Affiliation(s)
- W Yang
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - G Wu
- Department of Computational Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - U Broeckel
- Human and Molecular Genetics Center, Medical College of Wisconsin, Milwaukee, Wisconsin, USA
| | - C A Smith
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - V Turner
- Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - C E Haidar
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - S Wang
- Department of Computational Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - R Carter
- Department of Computational Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - S E Karol
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - G Neale
- Hartwell Center, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - K R Crews
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - J J Yang
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - C G Mullighan
- Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - J R Downing
- Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - W E Evans
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - M V Relling
- Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA.
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22
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Gohar NA, Rabie WA, Sharaf SA, Elsharkawy MM, Mira MF, Tolba AO, Aly H. Identification of insulin gene variants in neonatal diabetes. J Matern Fetal Neonatal Med 2016; 30:1035-1040. [PMID: 27279137 DOI: 10.1080/14767058.2016.1199674] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022]
Abstract
OBJECTIVES Permanent neonatal diabetes (PNDM) is caused by mutations in the genes responsible for the synthesis of different proteins that are important for the normal behavior of beta cells in the pancreas. Mutations in the insulin gene (INS) are considered as one of the causes of diabetes in neonates. This study aimed to investigate the genetic variations in the INS gene in a group of Egyptian infants diagnosed with PNDM. METHODS We screened exons 2 and 3 with intronic boundaries of the INS gene by direct gene sequencing in 30 PNDM patients and 20 healthy controls. A detailed clinical phenotyping of the patients was carried out to specify the diabetes features in those found to carry an INS variant. RESULTS We identified five variants (four SNPs and one synonymous variant), c(0).187 + 11T > C, c.-17-6T > A, c.*22A > C, c.*9C > T, and c.36G > A (p.A12A), with allelic frequencies of 96.7%, 80%, 75%, 5%, and 1.7%, respectively. All showed no statistically significance difference compared with the controls, with the exception of c.*22A > C. CONCLUSION Genetic screening for the INS gene did not reveal an evident role in the diagnosis of PNDM.
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Affiliation(s)
- Nadida A Gohar
- a Department of Clinical and Chemical Pathology , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt
| | - Walaa A Rabie
- a Department of Clinical and Chemical Pathology , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt
| | - Sahar A Sharaf
- a Department of Clinical and Chemical Pathology , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt
| | - Marwa M Elsharkawy
- a Department of Clinical and Chemical Pathology , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt
| | - Marwa F Mira
- b Department of Pediatrics , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt , and
| | - Aisha O Tolba
- a Department of Clinical and Chemical Pathology , Kasr Al-Aini Hospital, Cairo University , Cairo , Egypt
| | - Hany Aly
- c Division of Newborn Services , The George Washington University and Children's National Medical Center , Washington , DC , USA
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Daoud H, Luco SM, Li R, Bareke E, Beaulieu C, Jarinova O, Carson N, Nikkel SM, Graham GE, Richer J, Armour C, Bulman DE, Chakraborty P, Geraghty M, Lines MA, Lacaze-Masmonteil T, Majewski J, Boycott KM, Dyment DA. Next-generation sequencing for diagnosis of rare diseases in the neonatal intensive care unit. CMAJ 2016; 188:E254-E260. [PMID: 27241786 DOI: 10.1503/cmaj.150823] [Citation(s) in RCA: 80] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2016] [Accepted: 02/23/2016] [Indexed: 02/02/2023] Open
Abstract
BACKGROUND Rare diseases often present in the first days and weeks of life and may require complex management in the setting of a neonatal intensive care unit (NICU). Exhaustive consultations and traditional genetic or metabolic investigations are costly and often fail to arrive at a final diagnosis when no recognizable syndrome is suspected. For this pilot project, we assessed the feasibility of next-generation sequencing as a tool to improve the diagnosis of rare diseases in newborns in the NICU. METHODS We retrospectively identified and prospectively recruited newborns and infants admitted to the NICU of the Children's Hospital of Eastern Ontario and the Ottawa Hospital, General Campus, who had been referred to the medical genetics or metabolics inpatient consult service and had features suggesting an underlying genetic or metabolic condition. DNA from the newborns and parents was enriched for a panel of clinically relevant genes and sequenced on a MiSeq sequencing platform (Illumina Inc.). The data were interpreted with a standard informatics pipeline and reported to care providers, who assessed the importance of genotype-phenotype correlations. RESULTS Of 20 newborns studied, 8 received a diagnosis on the basis of next-generation sequencing (diagnostic rate 40%). The diagnoses were renal tubular dysgenesis, SCN1A-related encephalopathy syndrome, myotubular myopathy, FTO deficiency syndrome, cranioectodermal dysplasia, congenital myasthenic syndrome, autosomal dominant intellectual disability syndrome type 7 and Denys-Drash syndrome. INTERPRETATION This pilot study highlighted the potential of next-generation sequencing to deliver molecular diagnoses rapidly with a high success rate. With broader use, this approach has the potential to alter health care delivery in the NICU.
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Affiliation(s)
- Hussein Daoud
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Stephanie M Luco
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Rui Li
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Eric Bareke
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Chandree Beaulieu
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Olga Jarinova
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Nancy Carson
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Sarah M Nikkel
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Gail E Graham
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Julie Richer
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Christine Armour
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Dennis E Bulman
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Pranesh Chakraborty
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Michael Geraghty
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Matthew A Lines
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Thierry Lacaze-Masmonteil
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Jacek Majewski
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - Kym M Boycott
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que
| | - David A Dyment
- Department of Genetics (Daoud, Luco, Beaulieu, Jarinova, Carson, Nikkel, Graham, Richer, Armour, Boycott, Dyment) and Department of Pediatrics (Bulman, Chakraborty, Geraghty, Lines, Lacaze-Masmonteil), Children's Hospital of Eastern Ontario, Ottawa, Ont.; McGill University (Li, Bareke, Majewski) and Genome Quebec Innovation Centre (Li, Bareke, Majewski), Montréal, Que.
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Whole-genome sequencing overcomes pseudogene homology to diagnose autosomal dominant polycystic kidney disease. Eur J Hum Genet 2016; 24:1584-1590. [PMID: 27165007 DOI: 10.1038/ejhg.2016.48] [Citation(s) in RCA: 57] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2015] [Revised: 03/24/2016] [Accepted: 04/12/2016] [Indexed: 12/16/2022] Open
Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is the most common monogenic kidney disorder and is due to disease-causing variants in PKD1 or PKD2. Strong genotype-phenotype correlation exists although diagnostic sequencing is not part of routine clinical practice. This is because PKD1 bears 97.7% sequence similarity with six pseudogenes, requiring laborious and error-prone long-range PCR and Sanger sequencing to overcome. We hypothesised that whole-genome sequencing (WGS) would be able to overcome the problem of this sequence homology, because of 150 bp, paired-end reads and avoidance of capture bias that arises from targeted sequencing. We prospectively recruited a cohort of 28 unique pedigrees with ADPKD phenotype. Standard DNA extraction, library preparation and WGS were performed using Illumina HiSeq X and variants were classified following standard guidelines. Molecular diagnosis was made in 24 patients (86%), with 100% variant confirmation by current gold standard of long-range PCR and Sanger sequencing. We demonstrated unique alignment of sequencing reads over the pseudogene-homologous region. In addition to identifying function-affecting single-nucleotide variants and indels, we identified single- and multi-exon deletions affecting PKD1 and PKD2, which would have been challenging to identify using exome sequencing. We report the first use of WGS to diagnose ADPKD. This method overcomes pseudogene homology, provides uniform coverage, detects all variant types in a single test and is less labour-intensive than current techniques. This technique is translatable to a diagnostic setting, allows clinicians to make better-informed management decisions and has implications for other disease groups that are challenged by regions of confounding sequence homology.
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Development of a Comprehensive Sequencing Assay for Inherited Cardiac Condition Genes. J Cardiovasc Transl Res 2016; 9:3-11. [PMID: 26888179 PMCID: PMC4767849 DOI: 10.1007/s12265-016-9673-5] [Citation(s) in RCA: 65] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/15/2015] [Accepted: 01/07/2016] [Indexed: 12/15/2022]
Abstract
Inherited cardiac conditions (ICCs) are characterised by marked genetic and allelic heterogeneity and require extensive sequencing for genetic characterisation. We iteratively optimised a targeted gene capture panel for ICCs that includes disease-causing, putatively pathogenic, research and phenocopy genes (n = 174 genes). We achieved high coverage of the target region on both MiSeq (>99.8 % at ≥20× read depth, n = 12) and NextSeq (>99.9 % at ≥20×, n = 48) platforms with 100 % sensitivity and precision for single nucleotide variants and indels across the protein-coding target on the MiSeq. In the final assay, 40 out of 43 established ICC genes informative in clinical practice achieved complete coverage (100 % at ≥20×). By comparison, whole exome sequencing (WES; ∼80×), deep WES (∼500×) and whole genome sequencing (WGS; ∼70×) had poorer performance (88.1, 99.2 and 99.3 % respectively at ≥20×) across the ICC target. The assay described here delivers highly accurate and affordable sequencing of ICC genes, complemented by accessible cloud-based computation and informatics. See Editorial in this issue (DOI: 10.1007/s12265-015-9667-8).
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Bastida JM, del Rey M, Lozano ML, Sarasquete ME, Benito R, Fontecha ME, Fisac R, García-Frade LJ, Aguilar C, Martínez MP, Pardal E, Aguilera C, Pérez B, Ramos R, Cardesa MR, Martin-Antorán JM, Silvestre LA, Cebeira MJ, Bermejo N, Riesco S, Mendoza MC, García-Sanz R, González-Díaz M, Hernández-Rivas JM, González-Porras JR. Design and application of a 23-gene panel by next-generation sequencing for inherited coagulation bleeding disorders. Haemophilia 2016; 22:590-7. [DOI: 10.1111/hae.12908] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/22/2015] [Indexed: 12/19/2022]
Affiliation(s)
- J. M. Bastida
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
| | - M. del Rey
- Instituto de Investigación Biomédica de Salamanca; IBMCC; Centro de Investigación del Cáncer; Universidad de Salamanca-CSIC; Salamanca Spain
| | - M. L. Lozano
- Department of Hematology and Clinical Oncology; Centro Regional de Hemodonación; H. Universitario Morales Meseguer; IMIB-Arrixaca; Murcia Spain
| | - M. E. Sarasquete
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
| | - R. Benito
- Instituto de Investigación Biomédica de Salamanca; IBMCC; Centro de Investigación del Cáncer; Universidad de Salamanca-CSIC; Salamanca Spain
| | - M. E. Fontecha
- Department of Hematology; Hospital Universitario Rio Hortega de Valladolid; Valladolid Spain
| | - R. Fisac
- Department of Hematology; Hospital General de Segovia; Segovia Spain
| | - L. J. García-Frade
- Department of Hematology; Hospital Universitario Rio Hortega de Valladolid; Valladolid Spain
| | - C. Aguilar
- Department of Hematology; Complejo Asistencial de Soria; Soria Spain
| | - M. P. Martínez
- Department of Hematology; Complejo Asistencial de Avila; Avila Spain
| | - E. Pardal
- Department of Hematology; Hospital Virgen del Puerto de Plasencia; Caceres Spain
| | - C. Aguilera
- Department of Hematology; Hospital de El Bierzo; Ponferrada Spain
| | - B. Pérez
- Department of Hematology; Complejo Asistencial de Leon; Leon Spain
| | - R. Ramos
- Department of Hematology; Hospital de Merida; Badajoz Spain
| | - M. R. Cardesa
- Department of Hematology; Hospital de Merida; Badajoz Spain
| | | | - L. A. Silvestre
- Department of Hematology; Hospital Rio Carrion; Palencia Spain
| | - M. J. Cebeira
- Department of Hematology; Hospital Clinico Universitario de Valladolid; Valladolid Spain
| | - N. Bermejo
- Department of Hematology; Hospital San Pedro de Alcantara; Caceres Spain
| | - S. Riesco
- Department of Pediatrics; Hospital Universitario de Salamanca; Salamanca Spain
| | - M. C. Mendoza
- Department of Pediatrics; Hospital Universitario de Salamanca; Salamanca Spain
| | - R. García-Sanz
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
| | - M. González-Díaz
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
| | - J. M. Hernández-Rivas
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
- Instituto de Investigación Biomédica de Salamanca; IBMCC; Centro de Investigación del Cáncer; Universidad de Salamanca-CSIC; Salamanca Spain
| | - J. R. González-Porras
- Department of Hematology; H. Universitario de Salamanca; IBSAL; Instituto de Investigación Biomédica de Salamanca; Salamanca Spain
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Constellation: a tool for rapid, automated phenotype assignment of a highly polymorphic pharmacogene, CYP2D6, from whole-genome sequences. NPJ Genom Med 2016; 1:15007. [PMID: 29263805 PMCID: PMC5685293 DOI: 10.1038/npjgenmed.2015.7] [Citation(s) in RCA: 88] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2015] [Revised: 11/10/2015] [Accepted: 11/10/2015] [Indexed: 12/30/2022] Open
Abstract
An important component of precision medicine-the use of whole-genome sequencing (WGS) to guide lifelong healthcare-is electronic decision support to inform drug choice and dosing. To achieve this, automated identification of genetic variation in genes involved in drug absorption, distribution, metabolism, excretion and response (ADMER) is required. CYP2D6 is a major enzyme for drug bioactivation and elimination. CYP2D6 activity is predominantly governed by genetic variation; however, it is technically arduous to haplotype. Not only is the nucleotide sequence of CYP2D6 highly polymorphic, but the locus also features diverse structural variations, including gene deletion, duplication, multiplication events and rearrangements with the nonfunctional, neighbouring CYP2D7 and CYP2D8 genes. We developed Constellation, a probabilistic scoring system, enabling automated ascertainment of CYP2D6 activity scores from 2×100 paired-end WGS. The consensus reference method included TaqMan genotyping assays, quantitative copy-number variation determination and Sanger sequencing. When compared with the consensus reference Constellation had an analytic sensitivity of 97% (59 of 61 diplotypes) and analytic specificity of 95% (116 of 122 haplotypes). All extreme phenotypes, i.e., poor and ultrarapid metabolisers were accurately identified by Constellation. Constellation is anticipated to be extensible to functional variation in all ADMER genes, and to be performed at marginal incremental financial and computational costs in the setting of diagnostic WGS.
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Deem MJ. Whole-Genome Sequencing and Disability in the NICU: Exploring Practical and Ethical Challenges. Pediatrics 2016; 137 Suppl 1:S47-55. [PMID: 26729703 PMCID: PMC9923973 DOI: 10.1542/peds.2015-3731i] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
Clinical whole-genome sequencing (WGS) promises to deliver faster diagnoses and lead to better management of care in the NICU. However,several disability rights advocates have expressed concern that clinical use of genetic technologies may reinforce and perpetuate stigmatization of and discrimination against disabled persons in medical and social contexts. There is growing need, then, for clinicians and bioethicists to consider how the clinical use of WGS in the newborn period might exacerbate such harms to persons with disabilities. This article explores ways to extend these concerns to clinical WGS in neonatal care. By considering these perspectives during the early phases of expanded use of WGS in the NICU, this article encourages clinicians and bioethicists to continue to reflect on ways to attend to the concerns of disability rights advocates, foster trust and cooperation between the medical and disability communities, and forestall some of the social harms clinical WGS might cause to persons with disabilities and their families.
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Affiliation(s)
- Michael J. Deem
- Address correspondence to Michael J. Deem, Department of Multidisciplinary Studies, Holmstedt Hall 291, Indiana State University, Terre Haute, IN 47809. E-mail:
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Smith LD, Willig LK, Kingsmore SF. Whole-Exome Sequencing and Whole-Genome Sequencing in Critically Ill Neonates Suspected to Have Single-Gene Disorders. Cold Spring Harb Perspect Med 2015; 6:a023168. [PMID: 26684335 DOI: 10.1101/cshperspect.a023168] [Citation(s) in RCA: 83] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
As the ability to identify the contribution of genetic background to human disease continues to advance, there is no discipline of medicine in which this may have a larger impact than in the care of the ill neonate. Newborns with congenital malformations, syndromic conditions, and inherited disorders often undergo an extensive, expensive, and long diagnostic process, often without a final diagnosis resulting in significant health care, societal, and personal costs. Although ethical concerns have been raised about the use of whole-genome sequencing in medical practice, its role in the diagnosis of rare disorders in ill neonates in tertiary care neonatal intensive care units has the potential to augment or modify the care of this vulnerable population of patients.
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Affiliation(s)
- Laurie D Smith
- Department of Pediatrics, The University of Missouri-Kansas City School of Medicine, Kansas City, Missouri 64108 Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, Missouri 64108
| | - Laurel K Willig
- Department of Pediatrics, The University of Missouri-Kansas City School of Medicine, Kansas City, Missouri 64108 Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, Missouri 64108 Division of Pediatric Nephrology, Children's Mercy-Kansas City, Kansas City, Missouri 64108
| | - Stephen F Kingsmore
- Department of Pediatrics, The University of Missouri-Kansas City School of Medicine, Kansas City, Missouri 64108 Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, Missouri 64108
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Abstract
Traditionally, genetic testing has been too slow or perceived to be impractical to initial management of the critically ill neonate. Technological advances have led to the ability to sequence and interpret the entire genome of a neonate in as little as 26 h. As the cost and speed of testing decreases, the utility of whole genome sequencing (WGS) of neonates for acute and latent genetic illness increases. Analyzing the entire genome allows for concomitant evaluation of the currently identified 5588 single gene diseases. When applied to a select population of ill infants in a level IV neonatal intensive care unit, WGS yielded a diagnosis of a causative genetic disease in 57% of patients. These diagnoses may lead to clinical management changes ranging from transition to palliative care for uniformly lethal conditions for alteration or initiation of medical or surgical therapy to improve outcomes in others. Thus, institution of 2-day WGS at time of acute presentation opens the possibility of early implementation of precision medicine. This implementation may create opportunities for early interventional, frequently novel or off-label therapies that may alter disease trajectory in infants with what would otherwise be fatal disease. Widespread deployment of rapid WGS and precision medicine will raise ethical issues pertaining to interpretation of variants of unknown significance, discovery of incidental findings related to adult onset conditions and carrier status, and implementation of medical therapies for which little is known in terms of risks and benefits. Despite these challenges, precision neonatology has significant potential both to decrease infant mortality related to genetic diseases with onset in newborns and to facilitate parental decision making regarding transition to palliative care.
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Affiliation(s)
- Joshua E. Petrikin
- The University of Missouri Kansas City School of Medicine, Department of Pediatrics, Division of Neonatal and Perinatal Medicine, Director of Neonatal Genomics, Center for Pediatric Genomic Medicine, Children's Mercy Hospital Kansas City, Kansas City, Missouri 64108, Phone: 816-701-4806, Fax: 816-802-1111,
| | - Laurel K. Willig
- The University of Missouri, Kansas City School of Medicine, Department of Pediatrics, Division of Pediatric Nephrology, Center for Pediatric Genomic Medicine, Children's Mercy Hospital Kansas City, Kansas City, Missouri 64108 USA, Phone: 816-701-4806, Fax: 816-802-1111,
| | - Laurie D. Smith
- The University of Missouri Kansas City School of Medicine, Department of Pediatrics, Center for Pediatric Genomic Medicine, Children's Mercy Hospital Kansas City, Kansas City, Missouri 64108 USA, Phone: 816-701-4806, Fax: 816-802-111,
| | - Stephen F. Kingsmore
- Dee Lyons/Missouri Endowed Chair in Pediatric Genomic Medicine, Department of Pediatrics, Department of Pathology and Laboratory Medicine, Director, Center for Pediatric Genomic Medicine, Children's Mercy Hospital Kansas City, Kansas City, Missouri, 64108 USA, Phone: 816-701-4806, Fax: 816-802-1111,
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31
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Chiang C, Layer RM, Faust GG, Lindberg MR, Rose DB, Garrison EP, Marth GT, Quinlan AR, Hall IM. SpeedSeq: ultra-fast personal genome analysis and interpretation. Nat Methods 2015; 12:966-8. [PMID: 26258291 PMCID: PMC4589466 DOI: 10.1038/nmeth.3505] [Citation(s) in RCA: 385] [Impact Index Per Article: 38.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2014] [Accepted: 05/28/2015] [Indexed: 12/11/2022]
Abstract
SpeedSeq is an open-source genome analysis platform that accomplishes alignment, variant detection and functional annotation of a 50× human genome in 13 h on a low-cost server and alleviates a bioinformatics bottleneck that typically demands weeks of computation with extensive hands-on expert involvement. SpeedSeq offers performance competitive with or superior to current methods for detecting germline and somatic single-nucleotide variants, structural variants, insertions and deletions, and it includes novel functionality for streamlined interpretation.
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Affiliation(s)
- Colby Chiang
- McDonnell Genome Institute, Washington University School of Medicine, St. Louis, MO, USA
- Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA, USA
| | - Ryan M. Layer
- Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT, USA
- USTAR Center for Genetic Discovery, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Gregory G. Faust
- Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA, USA
| | - Michael R. Lindberg
- Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA, USA
| | - David B. Rose
- Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA, USA
| | - Erik P. Garrison
- USTAR Center for Genetic Discovery, University of Utah School of Medicine, Salt Lake City, UT, USA
- Wellcome Trust Sanger Institute, Hinxton, UK
| | - Gabor T. Marth
- Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT, USA
- USTAR Center for Genetic Discovery, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Aaron R. Quinlan
- Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT, USA
- USTAR Center for Genetic Discovery, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Ira M. Hall
- McDonnell Genome Institute, Washington University School of Medicine, St. Louis, MO, USA
- Department of Medicine, Washington University School of Medicine, St. Louis, MO, USA
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Achermann JC, Domenice S, Bachega TASS, Nishi MY, Mendonca BB. Disorders of sex development: effect of molecular diagnostics. Nat Rev Endocrinol 2015; 11:478-88. [PMID: 25942653 DOI: 10.1038/nrendo.2015.69] [Citation(s) in RCA: 59] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Disorders of sex development (DSDs) are a diverse group of conditions that can be challenging to diagnose accurately using standard phenotypic and biochemical approaches. Obtaining a specific diagnosis can be important for identifying potentially life-threatening associated disorders, as well as providing information to guide parents in deciding on the most appropriate management for their child. Within the past 5 years, advances in molecular methodologies have helped to identify several novel causes of DSDs; molecular tests to aid diagnosis and genetic counselling have now been adopted into clinical practice. Occasionally, genetic profiling of embryos prior to implantation as an adjunct to assisted reproduction, prenatal diagnosis of at-risk pregnancies and confirmatory testing of positive results found during newborn biochemical screening are performed. Of the available genetic tests, the candidate gene approach is the most popular. New high-throughput DNA analysis could enable a genetic diagnosis to be made when the aetiology is unknown or many differential diagnoses are possible. Nonetheless, concerns exist about the use of genetic tests. For instance, a diagnosis is not always possible even using new molecular approaches (which can be worrying for the parents) and incidental information obtained during the test might cause anxiety. Careful selection of the genetic test indicated for each condition remains important for good clinical practice. The purpose of this Review is to describe advances in molecular biological techniques for diagnosing DSDs.
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Affiliation(s)
- John C Achermann
- Developmental Endocrinology Research Group, Genetics and Genomic Medicine, UCL Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK
| | - Sorahia Domenice
- Unidade de Endocrinologia do Desenvolvimento, Laboratório de Hormônios e Genética Molecular LIM/42, Disciplina de Endocrinologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Av Dr Eneas de Carvalho Aguiar, 155, PAMB, 2 andar, Bloco 6, 05403-900 São Paulo, Brazil
| | - Tania A S S Bachega
- Unidade de Endocrinologia do Desenvolvimento, Laboratório de Hormônios e Genética Molecular LIM/42, Disciplina de Endocrinologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Av Dr Eneas de Carvalho Aguiar, 155, PAMB, 2 andar, Bloco 6, 05403-900 São Paulo, Brazil
| | - Mirian Y Nishi
- Unidade de Endocrinologia do Desenvolvimento, Laboratório de Hormônios e Genética Molecular LIM/42, Disciplina de Endocrinologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Av Dr Eneas de Carvalho Aguiar, 155, PAMB, 2 andar, Bloco 6, 05403-900 São Paulo, Brazil
| | - Berenice B Mendonca
- Unidade de Endocrinologia do Desenvolvimento, Laboratório de Hormônios e Genética Molecular LIM/42, Disciplina de Endocrinologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Av Dr Eneas de Carvalho Aguiar, 155, PAMB, 2 andar, Bloco 6, 05403-900 São Paulo, Brazil
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Khurana JK, Reeder JE, Shrimpton AE, Thakar J. GESPA: classifying nsSNPs to predict disease association. BMC Bioinformatics 2015. [PMID: 26206375 PMCID: PMC4513380 DOI: 10.1186/s12859-015-0673-2] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
BACKGROUND Non-synonymous single nucleotide polymorphisms (nsSNPs) are the most common DNA sequence variation associated with disease in humans. Thus determining the clinical significance of each nsSNP is of great importance. Potential detrimental nsSNPs may be identified by genetic association studies or by functional analysis in the laboratory, both of which are expensive and time consuming. Existing computational methods lack accuracy and features to facilitate nsSNP classification for clinical use. We developed the GESPA (GEnomic Single nucleotide Polymorphism Analyzer) program to predict the pathogenicity and disease phenotype of nsSNPs. RESULTS GESPA is a user-friendly software package for classifying disease association of nsSNPs. It allows flexibility in acceptable input formats and predicts the pathogenicity of a given nsSNP by assessing the conservation of amino acids in orthologs and paralogs and supplementing this information with data from medical literature. The development and testing of GESPA was performed using the humsavar, ClinVar and humvar datasets. Additionally, GESPA also predicts the disease phenotype associated with a nsSNP with high accuracy, a feature unavailable in existing software. GESPA's overall accuracy exceeds existing computational methods for predicting nsSNP pathogenicity. The usability of GESPA is enhanced by fast SQL-based cloud storage and retrieval of data. CONCLUSIONS GESPA is a novel bioinformatics tool to determine the pathogenicity and phenotypes of nsSNPs. We anticipate that GESPA will become a useful clinical framework for predicting the disease association of nsSNPs. The program, executable jar file, source code, GPL 3.0 license, user guide, and test data with instructions are available at http://sourceforge.net/projects/gespa.
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Affiliation(s)
- Jay K Khurana
- Department of Urology, SUNY Upstate Medical University, Syracuse, NY, USA.
| | - Jay E Reeder
- Department of Obstetrics and Gynecology, University of Rochester, Rochester, NY, USA.
| | - Antony E Shrimpton
- Department of Pathology, SUNY Upstate Medical University, Syracuse, NY, USA.
| | - Juilee Thakar
- Department of Microbiology and Immunology, University of Rochester, Rochester, NY, USA. .,Department of Biostatistics and Computational Biology, University of Rochester, Rochester, NY, USA.
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Milner LC, Garrison NA, Cho MK, Altman RB, Hudgins L, Galli SJ, Lowe HJ, Schrijver I, Magnus DC. Genomics in the clinic: ethical and policy challenges in clinical next-generation sequencing programs at early adopter USA institutions. Per Med 2015; 12:269-282. [PMID: 29771644 DOI: 10.2217/pme.14.88] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Next-generation sequencing (NGS) technologies are poised to revolutionize clinical diagnosis and treatment, but raise significant ethical and policy challenges. This review examines NGS program challenges through a synthesis of published literature, website and conference presentation content, and interviews at early-adopting institutions in the USA. Institutions are proactively addressing policy challenges related to the management and technical aspects of program development. However, ethical challenges related to patient-related aspects have not been fully addressed. These complex challenges present opportunities to develop comprehensive and standardized regulations across programs. Understanding the strengths, weaknesses and current practices of evolving NGS program approaches are important considerations for institutions developing NGS services, policymakers regulating or funding NGS programs and physicians and patients considering NGS services.
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Affiliation(s)
- Lauren C Milner
- Stanford Center for Biomedical Ethics, Stanford University School of Medicine, Stanford, CA, USA
| | - Nanibaa' A Garrison
- Stanford Center for Biomedical Ethics, Stanford University School of Medicine, Stanford, CA, USA.,Center for Biomedical Ethics & Society, Departments of Pediatrics & Anthropology, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Mildred K Cho
- Stanford Center for Biomedical Ethics, Stanford University School of Medicine, Stanford, CA, USA.,Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA
| | - Russ B Altman
- Department of Bioengineering, Stanford University School of Medicine, Stanford, CA, USA.,Department of Genetics, Stanford University School of Medicine, Stanford, CA, USA.,Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Louanne Hudgins
- Division of Medical Genetics, Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA
| | - Stephen J Galli
- Stanford Center for Genomics & Personalized Medicine, Stanford University School of Medicine, Stanford, CA, USA.,Department of Pathology, Stanford University School of Medicine, Stanford, CA, USA.,Department of Microbiology & Immunology, Stanford University School of Medicine, Stanford, CA, USA
| | - Henry J Lowe
- Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Iris Schrijver
- Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA.,Stanford Center for Genomics & Personalized Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - David C Magnus
- Stanford Center for Biomedical Ethics, Stanford University School of Medicine, Stanford, CA, USA.,Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA
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Willig LK, Petrikin JE, Smith LD, Saunders CJ, Thiffault I, Miller NA, Soden SE, Cakici JA, Herd SM, Twist G, Noll A, Creed M, Alba PM, Carpenter SL, Clements MA, Fischer RT, Hays JA, Kilbride H, McDonough RJ, Rosterman JL, Tsai SL, Zellmer L, Farrow EG, Kingsmore SF. Whole-genome sequencing for identification of Mendelian disorders in critically ill infants: a retrospective analysis of diagnostic and clinical findings. THE LANCET RESPIRATORY MEDICINE 2015; 3:377-87. [PMID: 25937001 DOI: 10.1016/s2213-2600(15)00139-3] [Citation(s) in RCA: 295] [Impact Index Per Article: 29.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/23/2015] [Revised: 03/30/2015] [Accepted: 04/01/2015] [Indexed: 12/28/2022]
Abstract
BACKGROUND Genetic disorders and congenital anomalies are the leading causes of infant mortality. Diagnosis of most genetic diseases in neonatal and paediatric intensive care units (NICU and PICU) is not sufficiently timely to guide acute clinical management. We used rapid whole-genome sequencing (STATseq) in a level 4 NICU and PICU to assess the rate and types of molecular diagnoses, and the prevalence, types, and effect of diagnoses that are likely to change medical management in critically ill infants. METHODS We did a retrospective comparison of STATseq and standard genetic testing in a case series from the NICU and PICU of a large children's hospital between Nov 11, 2011, and Oct 1, 2014. The participants were families with an infant younger than 4 months with an acute illness of suspected genetic cause. The intervention was STATseq of trios (both parents and their affected infant). The main measures were the diagnostic rate, time to diagnosis, and rate of change in management after standard genetic testing and STATseq. FINDINGS 20 (57%) of 35 infants were diagnosed with a genetic disease by use of STATseq and three (9%) of 32 by use of standard genetic testing (p=0·0002). Median time to genome analysis was 5 days (range 3-153) and median time to STATseq report was 23 days (5-912). 13 (65%) of 20 STATseq diagnoses were associated with de-novo mutations. Acute clinical usefulness was noted in 13 (65%) of 20 infants with a STATseq diagnosis, four (20%) had diagnoses with strongly favourable effects on management, and six (30%) were started on palliative care. 120-day mortality was 57% (12 of 21) in infants with a genetic diagnosis. INTERPRETATION In selected acutely ill infants, STATseq had a high rate of diagnosis of genetic disorders. Most diagnoses altered the management of infants in the NICU or PICU. The very high infant mortality rate indicates a substantial need for rapid genomic diagnoses to be allied with a novel framework for precision medicine for infants in NICU and PICU who are diagnosed with genetic diseases to improve outcomes. FUNDING Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Human Genome Research Institute, and National Center for Advancing Translational Sciences.
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Affiliation(s)
- Laurel K Willig
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Josh E Petrikin
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Laurie D Smith
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Carol J Saunders
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pathology, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Isabelle Thiffault
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pathology, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Neil A Miller
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Sarah E Soden
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pathology, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Julie A Cakici
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Suzanne M Herd
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Greyson Twist
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Aaron Noll
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Mitchell Creed
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Patria M Alba
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Shannon L Carpenter
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Mark A Clements
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Ryan T Fischer
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - J Allyson Hays
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Howard Kilbride
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Ryan J McDonough
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Jamie L Rosterman
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Sarah L Tsai
- Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Lee Zellmer
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pathology, Children's Mercy-Kansas City, Kansas City, MO, USA
| | - Emily G Farrow
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA
| | - Stephen F Kingsmore
- Center for Pediatric Genomic Medicine, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pediatrics, Children's Mercy-Kansas City, Kansas City, MO, USA; Department of Pathology, Children's Mercy-Kansas City, Kansas City, MO, USA; School of Medicine, University of Missouri-Kansas City, Kansas City, Missouri 64108, USA.
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Shi J, Qu YP, Hou P. Pathogenetic mechanisms in gastric cancer. World J Gastroenterol 2014; 20:13804-13819. [PMID: 25320518 PMCID: PMC4194564 DOI: 10.3748/wjg.v20.i38.13804] [Citation(s) in RCA: 78] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/24/2013] [Revised: 01/15/2014] [Accepted: 05/29/2014] [Indexed: 02/06/2023] Open
Abstract
Gastric cancer (GC) is a major public health issue as the fourth most common cancer and the second leading cause of cancer-related death. Recent advances have improved our understanding of its molecular pathogenesis, as best exemplified by elucidating the fundamental role of several major signaling pathways and related molecular derangements. Central to these mechanisms are the genetic and epigenetic alterations in these signaling pathways, such as gene mutations, copy number variants, aberrant gene methylation and histone modification, nucleosome positioning, and microRNAs. Some of these genetic/epigenetic alterations represent effective diagnostic and prognostic biomarkers and therapeutic targets for GC. This information has now opened unprecedented opportunities for better understanding of the molecular mechanisms of gastric carcinogenesis and the development of novel therapeutic strategies for this cancer. The pathogenetic mechanisms of GC are the focus of this review.
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Raje N, Soden S, Swanson D, Ciaccio CE, Kingsmore SF, Dinwiddie DL. Utility of next generation sequencing in clinical primary immunodeficiencies. Curr Allergy Asthma Rep 2014; 14:468. [PMID: 25149170 PMCID: PMC4582650 DOI: 10.1007/s11882-014-0468-y] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
Primary immunodeficiencies (PIDs) are a group of genetically heterogeneous disorders that present with very similar symptoms, complicating definitive diagnosis. More than 240 genes have hitherto been associated with PIDs, of which more than 30 have been identified in the last 3 years. Next generation sequencing (NGS) of genomes or exomes of informative families has played a central role in the discovery of novel PID genes. Furthermore, NGS has the potential to transform clinical molecular testing for established PIDs, allowing all PID differential diagnoses to be tested at once, leading to increased diagnostic yield, while decreasing both the time and cost of obtaining a molecular diagnosis. Given that treatment of PID varies by disease gene, early achievement of a molecular diagnosis is likely to enhance treatment decisions and improve patient outcomes.
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Affiliation(s)
- Nikita Raje
- Children's Mercy Hospital, 2401 Gillham Road, Kansas City, MO, 64108, USA,
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Hendriks G, Morolli B, Calléja FMGR, Plomp A, Mesman RLS, Meijers M, Sharan SK, Vreeswijk MPG, Vrieling H. An efficient pipeline for the generation and functional analysis of human BRCA2 variants of uncertain significance. Hum Mutat 2014; 35:1382-91. [PMID: 25146914 DOI: 10.1002/humu.22678] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2014] [Accepted: 08/04/2014] [Indexed: 11/08/2022]
Abstract
The implementation of next-generation sequence analysis of disease-related genes has resulted in an increasing number of genetic variants with an unknown clinical significance. The functional analysis of these so-called "variants of uncertain significance" (VUS) is hampered by the tedious and time-consuming procedures required to generate and test specific sequence variants in genomic DNA. Here, we describe an efficient pipeline for the generation of gene variants in a full-length human gene, BRCA2, using a bacterial artificial chromosome. This method permits the rapid generation of intronic and exonic variants in a complete gene through the use of an exon-replacement strategy based on simple site-directed mutagenesis and an effective positive-negative selection system in E. coli. The functionality of variants can then be assessed through the use of functional assays, such as complementation of gene-deficient mouse-embryonic stem (mES) cells in the case of human BRCA2. Our methodology builds upon an earlier protocol and, through the introduction of a series of major innovations, now represents a practical proposition for the rapid analysis of BRCA2 variants and a blueprint for the analysis of other genes using similar approaches. This method enables rapid generation and reliable classification of VUS in disease-related genes, allowing informed clinical decision-making.
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Affiliation(s)
- Giel Hendriks
- Department of Toxicogenetics, Leiden University Medical Center, Leiden, The Netherlands
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Zemojtel T, Köhler S, Mackenroth L, Jäger M, Hecht J, Krawitz P, Graul-Neumann L, Doelken S, Ehmke N, Spielmann M, Oien NC, Schweiger MR, Krüger U, Frommer G, Fischer B, Kornak U, Flöttmann R, Ardeshirdavani A, Moreau Y, Lewis SE, Haendel M, Smedley D, Horn D, Mundlos S, Robinson PN. Effective diagnosis of genetic disease by computational phenotype analysis of the disease-associated genome. Sci Transl Med 2014; 6:252ra123. [PMID: 25186178 PMCID: PMC4512639 DOI: 10.1126/scitranslmed.3009262] [Citation(s) in RCA: 189] [Impact Index Per Article: 17.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Less than half of patients with suspected genetic disease receive a molecular diagnosis. We have therefore integrated next-generation sequencing (NGS), bioinformatics, and clinical data into an effective diagnostic workflow. We used variants in the 2741 established Mendelian disease genes [the disease-associated genome (DAG)] to develop a targeted enrichment DAG panel (7.1 Mb), which achieves a coverage of 20-fold or better for 98% of bases. Furthermore, we established a computational method [Phenotypic Interpretation of eXomes (PhenIX)] that evaluated and ranked variants based on pathogenicity and semantic similarity of patients' phenotype described by Human Phenotype Ontology (HPO) terms to those of 3991 Mendelian diseases. In computer simulations, ranking genes based on the variant score put the true gene in first place less than 5% of the time; PhenIX placed the correct gene in first place more than 86% of the time. In a retrospective test of PhenIX on 52 patients with previously identified mutations and known diagnoses, the correct gene achieved a mean rank of 2.1. In a prospective study on 40 individuals without a diagnosis, PhenIX analysis enabled a diagnosis in 11 cases (28%, at a mean rank of 2.4). Thus, the NGS of the DAG followed by phenotype-driven bioinformatic analysis allows quick and effective differential diagnostics in medical genetics.
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Affiliation(s)
- Tomasz Zemojtel
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Institute of Bioorganic Chemistry, Polish Academy of Sciences, 61-704 Poznan, Poland. Labor Berlin-Charité Vivantes GmbH, Humangenetik, Föhrer Straße 15, 13353 Berlin, Germany
| | - Sebastian Köhler
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Luisa Mackenroth
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Marten Jäger
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Jochen Hecht
- Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany. Berlin-Brandenburg Center for Regenerative Therapies, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany
| | - Peter Krawitz
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany
| | - Luitgard Graul-Neumann
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Sandra Doelken
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Nadja Ehmke
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Malte Spielmann
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany
| | - Nancy Christine Oien
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Delbrück Center for Molecular Medicine, Robert-Rössle-Str. 10, 13125 Berlin, Germany
| | - Michal R Schweiger
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany. Cologne Center for Genomics, University of Cologne, D-50931 Cologne, Germany
| | - Ulrike Krüger
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Götz Frommer
- Agilent Technologies, Hewlett-Packard-Straße 8, 76337 Waldbronn, Germany
| | - Björn Fischer
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany
| | - Uwe Kornak
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany
| | - Ricarda Flöttmann
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Amin Ardeshirdavani
- Department of Electrical Engineering, STADIUS Center for Dynamical Systems, Signal Processing and Data Analytics, KU Leuven, 3001 Leuven, Belgium
| | - Yves Moreau
- Department of Electrical Engineering, STADIUS Center for Dynamical Systems, Signal Processing and Data Analytics, KU Leuven, 3001 Leuven, Belgium
| | - Suzanna E Lewis
- Genomics Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA
| | - Melissa Haendel
- University Library and Department of Medical Informatics and Clinical Epidemiology, Oregon Health & Sciences University, Portland, OR 97327, USA
| | - Damian Smedley
- Mouse Informatics Group, Wellcome Trust Sanger Institute, CB10 1SA Hinxton, UK
| | - Denise Horn
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Stefan Mundlos
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany. Berlin-Brandenburg Center for Regenerative Therapies, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany
| | - Peter N Robinson
- Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Max Planck Institute for Molecular Genetics, Ihnestr. 63-73, 14195 Berlin, Germany. Berlin-Brandenburg Center for Regenerative Therapies, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany. Institute for Bioinformatics, Department of Mathematics and Computer Science, Freie Universität Berlin, Takustr. 9, 14195 Berlin, Germany.
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40
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Ten years of next-generation sequencing technology. Trends Genet 2014; 30:418-26. [PMID: 25108476 DOI: 10.1016/j.tig.2014.07.001] [Citation(s) in RCA: 897] [Impact Index Per Article: 81.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2014] [Revised: 07/08/2014] [Accepted: 07/09/2014] [Indexed: 02/06/2023]
Abstract
Ten years ago next-generation sequencing (NGS) technologies appeared on the market. During the past decade, tremendous progress has been made in terms of speed, read length, and throughput, along with a sharp reduction in per-base cost. Together, these advances democratized NGS and paved the way for the development of a large number of novel NGS applications in basic science as well as in translational research areas such as clinical diagnostics, agrigenomics, and forensic science. Here we provide an overview of the evolution of NGS and discuss the most significant improvements in sequencing technologies and library preparation protocols. We also explore the current landscape of NGS applications and provide a perspective for future developments.
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Sawyer SL, Schwartzentruber J, Beaulieu CL, Dyment D, Smith A, Chardon JW, Yoon G, Rouleau GA, Suchowersky O, Siu V, Murphy L, Hegele RA, Marshall CR, Bulman DE, Majewski J, Tarnopolsky M, Boycott KM. Exome sequencing as a diagnostic tool for pediatric-onset ataxia. Hum Mutat 2014; 35:45-9. [PMID: 24108619 PMCID: PMC4255313 DOI: 10.1002/humu.22451] [Citation(s) in RCA: 81] [Impact Index Per Article: 7.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2013] [Revised: 09/20/2013] [Accepted: 09/22/2013] [Indexed: 12/30/2022]
Abstract
Ataxia demonstrates substantial phenotypic and genetic heterogeneity. We set out to determine the diagnostic yield of exome sequencing in pediatric patients with ataxia without a molecular diagnosis after standard-of-care assessment in Canada. FORGE (Finding Of Rare disease GEnes) Canada is a nation-wide project focused on identifying novel disease genes for rare pediatric diseases using whole-exome sequencing. We retrospectively selected all FORGE Canada projects that included cerebellar ataxia as a feature. We identified 28 such families and a molecular diagnosis was made in 13; a success rate of 46%. In 11 families, we identified mutations in genes associated with known neurological syndromes and in two we identified novel disease genes. Exome analysis of sib pairs and/or patients born to consanguineous parents was more likely to be successful (9/13) than simplex cases (4/15). Our data suggest that exome sequencing is an effective first line test for pediatric patients with ataxia where a specific single gene is not immediately suspected to be causative.
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Affiliation(s)
- Sarah L Sawyer
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
| | | | - Chandree L Beaulieu
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
| | - David Dyment
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
| | - Amanda Smith
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
| | - Jodi Warman Chardon
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
| | - Grace Yoon
- Divisions of Neurology and Clinical and Metabolic Genetics, Hospital for Sick Children, University of TorontoToronto, Ontario, Canada
| | - Guy A Rouleau
- Montreal Neurological Institute and Hospital, McGill University MontrealQuebec, Canada
| | - Oksana Suchowersky
- Departments of Medicine (Neurology) and Medical Genetics, University of AlbertaEdmonton, Alberta, Canada
| | - Victoria Siu
- Department of Pediatrics, Division of Medical Genetics, Western UniversityLondon, Ontario, Canada
| | - Lisa Murphy
- Department of Pediatrics, Division of Medical Genetics, Western UniversityLondon, Ontario, Canada
| | - Robert A Hegele
- Robarts Research Institute, University of Western OntarioLondon, Canada
| | - Christian R Marshall
- Program in Genetics and Genome Biology, Hospital for Sick Children and McLaughlin Centre, University of TorontoToronto, Ontario, Canada
| | | | - Jacek Majewski
- Department of Human Genetics, McGill UniversityMontréal, Quebec, Canada
| | - Mark Tarnopolsky
- Department of Pediatrics, McMaster Children's HospitalHamilton, Ontario, Canada
| | - Kym M Boycott
- Children's Hospital of Eastern Ontario Research Institute, University of OttawaOttawa, Ontario, Canada
- *Correspondence to: Kym M Boycott, Children's Hospital of Eastern Ontario Research Institute, 401 Smyth Road, Ottawa, ON, K1H 8L1, Canada. E-mail:
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Masino AJ, Dechene ET, Dulik MC, Wilkens A, Spinner NB, Krantz ID, Pennington JW, Robinson PN, White PS. Clinical phenotype-based gene prioritization: an initial study using semantic similarity and the human phenotype ontology. BMC Bioinformatics 2014; 15:248. [PMID: 25047600 PMCID: PMC4117966 DOI: 10.1186/1471-2105-15-248] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2014] [Accepted: 07/16/2014] [Indexed: 12/21/2022] Open
Abstract
Background Exome sequencing is a promising method for diagnosing patients with a complex phenotype. However, variant interpretation relative to patient phenotype can be challenging in some scenarios, particularly clinical assessment of rare complex phenotypes. Each patient’s sequence reveals many possibly damaging variants that must be individually assessed to establish clear association with patient phenotype. To assist interpretation, we implemented an algorithm that ranks a given set of genes relative to patient phenotype. The algorithm orders genes by the semantic similarity computed between phenotypic descriptors associated with each gene and those describing the patient. Phenotypic descriptor terms are taken from the Human Phenotype Ontology (HPO) and semantic similarity is derived from each term’s information content. Results Model validation was performed via simulation and with clinical data. We simulated 33 Mendelian diseases with 100 patients per disease. We modeled clinical conditions by adding noise and imprecision, i.e. phenotypic terms unrelated to the disease and terms less specific than the actual disease terms. We ranked the causative gene against all 2488 HPO annotated genes. The median causative gene rank was 1 for the optimal and noise cases, 12 for the imprecision case, and 60 for the imprecision with noise case. Additionally, we examined a clinical cohort of subjects with hearing impairment. The disease gene median rank was 22. However, when also considering the patient’s exome data and filtering non-exomic and common variants, the median rank improved to 3. Conclusions Semantic similarity can rank a causative gene highly within a gene list relative to patient phenotype characteristics, provided that imprecision is mitigated. The clinical case results suggest that phenotype rank combined with variant analysis provides significant improvement over the individual approaches. We expect that this combined prioritization approach may increase accuracy and decrease effort for clinical genetic diagnosis. Electronic supplementary material The online version of this article (doi:10.1186/1471-2105-15-248) contains supplementary material, which is available to authorized users.
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Affiliation(s)
| | | | | | | | | | | | | | | | - Peter S White
- Department of Pediatrics, Cincinnati Children's Hospital and Medical Center, Cincinnati, OH, USA.
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The business of genomic testing: a survey of early adopters. Genet Med 2014; 16:954-61. [PMID: 25010053 PMCID: PMC4262758 DOI: 10.1038/gim.2014.60] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2014] [Accepted: 04/24/2014] [Indexed: 12/16/2022] Open
Abstract
PURPOSE The practice of "genomic" (or "personalized") medicine requires the availability of appropriate diagnostic testing. Our study objective was to identify the reasons for health systems to bring next-generation sequencing into their clinical laboratories and to understand the process by which such decisions were made. Such information may be of value to other health systems seeking to provide next-generation sequencing testing to their patient populations. METHODS A standardized open-ended interview was conducted with the laboratory medical directors and/or department of pathology chairs of 13 different academic institutions in 10 different states. RESULTS Genomic testing for cancer dominated the institutional decision making, with three primary reasons: more effective delivery of cancer care, the perceived need for institutional leadership in the field of genomics, and the premise that genomics will eventually be cost-effective. Barriers to implementation included implementation cost; the time and effort needed to maintain this newer testing; challenges in interpreting genetic variants; establishing the bioinformatics infrastructure; and curating data from medical, ethical, and legal standpoints. Ultimate success depended on alignment with institutional strengths and priorities and working closely with institutional clinical programs. CONCLUSION These early adopters uniformly viewed genomic analysis as an imperative for developing their expertise in the implementation and practice of genomic medicine.
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Koay PP, Sharp RR. Managing Expectational Language: Translational genetic professionals consider the clinical potential of next-generation sequencing technologies. NEW GENETICS AND SOCIETY 2014; 33:126-148. [PMID: 24883042 PMCID: PMC4038681 DOI: 10.1080/14636778.2014.910448] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/03/2013] [Accepted: 03/13/2014] [Indexed: 05/20/2023]
Affiliation(s)
- Pei P Koay
- Center for Genetic Research Ethics & Law (CGREAL), Department of Bioethics, Case Western Reserve University, School of Medicine, Cleveland, OH, USA
| | - Richard R Sharp
- Director, Biomedical Ethics Program, Mayo Clinic, Rochester, MN, USA
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Swanson A, Ramos E, Snyder H. Next Generation Sequencing is the Impetus for the Next Generation of Laboratory-Based Genetic Counselors. J Genet Couns 2014; 23:647-54. [DOI: 10.1007/s10897-013-9684-1] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2013] [Accepted: 12/12/2013] [Indexed: 02/04/2023]
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Abstract
A rare or orphan disorder is any disease that affects a small percentage of the population. Most genes and pathways underlying these disorders remain unknown. High-throughput techniques are frequently applied to detect disease candidate genes. The speed and affordability of sequencing following recent technological advances while advantageous are accompanied by the problem of data deluge. Furthermore, experimental validation of disease candidate genes is both time-consuming and expensive. Therefore, several computational approaches have been developed to identify the most promising candidates for follow-up studies. Based on the guilt by association principle, most of these approaches use prior knowledge about a disease of interest to discover and rank novel candidate genes. In this chapter, a brief overview of some of the in silico strategies for candidate gene prioritization is provided. To demonstrate their utility in rare disease research, a Web-based computational suite of tools that use integrated heterogeneous data sources for ranking disease candidate genes is used to demonstrate how to run typical queries using this system.
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Affiliation(s)
- Anil G Jegga
- Division of Biomedical Informatics, Cincinnati Children's Hospital Medical Center, 3333 Burnet Avenue, ML 7024, Cincinnati, OH, 45229, USA,
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47
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The promise of whole-exome sequencing in medical genetics. J Hum Genet 2013; 59:5-15. [DOI: 10.1038/jhg.2013.114] [Citation(s) in RCA: 312] [Impact Index Per Article: 26.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2013] [Revised: 09/29/2013] [Accepted: 10/11/2013] [Indexed: 12/14/2022]
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Dotta L, Parolini S, Prandini A, Tabellini G, Antolini M, Kingsmore SF, Badolato R. Clinical, laboratory and molecular signs of immunodeficiency in patients with partial oculo-cutaneous albinism. Orphanet J Rare Dis 2013; 8:168. [PMID: 24134793 PMCID: PMC3856608 DOI: 10.1186/1750-1172-8-168] [Citation(s) in RCA: 51] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2013] [Accepted: 10/11/2013] [Indexed: 01/04/2023] Open
Abstract
Hypopigmentation disorders that are associated with immunodeficiency feature both partial albinism of hair, skin and eyes together with leukocyte defects. These disorders include Chediak Higashi (CHS), Griscelli (GS), Hermansky-Pudlak (HPS) and MAPBP-interacting protein deficiency syndromes. These are heterogeneous autosomal recessive conditions in which the causal genes encode proteins with specific roles in the biogenesis, function and trafficking of secretory lysosomes. In certain specialized cells, these organelles serve as a storage compartment. Impaired secretion of specific effector proteins from that intracellular compartment affects biological activities. In particular, these intracellular granules are essential constituents of melanocytes, platelets, granulocytes, cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. Thus, abnormalities affect pigmentation, primary hemostasis, blood cell counts and lymphocyte cytotoxic activity against microbial pathogens. Among eight genetically distinct types of HPS, only type 2 is characterized by immunodeficiency. Recently, a new subtype, HPS9, was defined in patients presenting with immunodeficiency and oculocutaneous albinism, associated with mutations in the pallidin-encoding gene, PLDN.Hypopigmentation together with recurrent childhood bacterial or viral infections suggests syndromic albinism. T and NK cell cytotoxicity are generally impaired in patients with these disorders. Specific clinical and biochemical phenotypes can allow differential diagnoses among these disorders before molecular testing. Ocular symptoms, including nystagmus, that are usually evident at birth, are common in patients with HPS2 or CHS. Albinism with short stature is unique to MAPBP-interacting protein (MAPBPIP) deficiency, while hemophagocytic lymphohistiocytosis (HLH) mainly suggests a diagnosis of CHS or GS type 2 (GS2). Neurological disease is a long-term complication of CHS, but is uncommon in other syndromic albinism. Chronic neutropenia is a feature of HPS2 and MAPBPIP-deficiency syndrome, whereas it is usually transient in CHS and GS2. In every patient, an accurate diagnosis is required for prompt and appropriate treatment, particularly in patients who develop HLH or in whom bone marrow transplant is required. This review describes the molecular and pathogenetic mechanisms of these diseases, focusing on clinical and biochemical aspects that allow early differential diagnosis.
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Affiliation(s)
- Laura Dotta
- Department of Experimental and Clinical Sciences, Institute of Molecular Medicine “Angelo Nocivelli”, University of Brescia, Brescia, Italy
| | - Silvia Parolini
- Department of Molecular and Translational Medicine, University of Brescia, Brescia 25123, Italy
| | - Alberto Prandini
- Department of Experimental and Clinical Sciences, Institute of Molecular Medicine “Angelo Nocivelli”, University of Brescia, Brescia, Italy
| | - Giovanna Tabellini
- Department of Molecular and Translational Medicine, University of Brescia, Brescia 25123, Italy
| | - Maddalena Antolini
- Department of Experimental and Clinical Sciences, Institute of Molecular Medicine “Angelo Nocivelli”, University of Brescia, Brescia, Italy
| | - Stephen F Kingsmore
- Center for Pediatric Genomic Medicine, Children’s Mercy Hospital, Kansas City, MO 64108, USA
| | - Raffaele Badolato
- Department of Experimental and Clinical Sciences, Institute of Molecular Medicine “Angelo Nocivelli”, University of Brescia, Brescia, Italy
- Istituto di Medicina Molecolare “Angelo Nocivelli”, Universita' di Brescia, c/o Spedali Civili, Brescia 25123, Italy
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Prosperi MCF, Yin L, Nolan DJ, Lowe AD, Goodenow MM, Salemi M. Empirical validation of viral quasispecies assembly algorithms: state-of-the-art and challenges. Sci Rep 2013; 3:2837. [PMID: 24089188 PMCID: PMC3789152 DOI: 10.1038/srep02837] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2013] [Accepted: 09/13/2013] [Indexed: 11/22/2022] Open
Abstract
Next generation sequencing (NGS) is superseding Sanger technology for analysing intra-host viral populations, in terms of genome length and resolution. We introduce two new empirical validation data sets and test the available viral population assembly software. Two intra-host viral population 'quasispecies' samples (type-1 human immunodeficiency and hepatitis C virus) were Sanger-sequenced, and plasmid clone mixtures at controlled proportions were shotgun-sequenced using Roche's 454 sequencing platform. The performance of different assemblers was compared in terms of phylogenetic clustering and recombination with the Sanger clones. Phylogenetic clustering showed that all assemblers captured a proportion of the most divergent lineages, but none were able to provide a high precision/recall tradeoff. Estimated variant frequencies mildly correlated with the original. Given the limitations of currently available algorithms identified by our empirical validation, the development and exploitation of additional data sets is needed, in order to establish an efficient framework for viral population reconstruction using NGS.
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Affiliation(s)
- Mattia C. F. Prosperi
- University of Manchester, Faculty of Medical and Human Sciences, Northwest Institute of Bio-Health Informatics, Centre for Health Informatics, Institute of Population Health, Manchester, UK
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
| | - Li Yin
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
- Florida Center for AIDS Research, Gainesville, Florida, USA
| | - David J. Nolan
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
| | - Amanda D. Lowe
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
- Florida Center for AIDS Research, Gainesville, Florida, USA
| | - Maureen M. Goodenow
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
- Florida Center for AIDS Research, Gainesville, Florida, USA
| | - Marco Salemi
- University of Florida, College of Medicine, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, Florida, USA
- Florida Center for AIDS Research, Gainesville, Florida, USA
- Emerging Pathogens Institute, Gainesville, Florida, USA
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Boycott KM, Vanstone MR, Bulman DE, MacKenzie AE. Rare-disease genetics in the era of next-generation sequencing: discovery to translation. Nat Rev Genet 2013; 14:681-91. [PMID: 23999272 DOI: 10.1038/nrg3555] [Citation(s) in RCA: 523] [Impact Index Per Article: 43.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Work over the past 25 years has resulted in the identification of genes responsible for ~50% of the estimated 7,000 rare monogenic diseases, and it is predicted that most of the remaining disease-causing genes will be identified by the year 2020, and probably sooner. This marked acceleration is the result of dramatic improvements in DNA-sequencing technologies and the associated analyses. We examine the rapid maturation of rare-disease genetic analysis and successful strategies for gene identification. We highlight the impact of discovering rare-disease-causing genes, from clinical diagnostics to insights gained into biological mechanisms and common diseases. Last, we explore the increasing therapeutic opportunities and challenges that the resulting expansion of the 'atlas' of human genetic pathology will bring.
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Affiliation(s)
- Kym M Boycott
- Department of Pediatrics, University of Ottawa and Children's Hospital of Eastern Ontario Research Institute, 401 Smyth Road, Ottawa, Ontario K1H 8L1, Canada
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