Pontocerebellar hypoplasia with spinal muscular atrophy, also known as PCH1, is a group of autosomal recessive disorders characterized by generalized muscle weakness and global developmental delay ...commonly resulting in early death. Gene defects had been discovered only in single patients until the recent identification of EXOSC3 mutations in several families with relatively mild course of PCH1. We aim to genetically stratify subjects in a large and well-defined cohort to define the clinical spectrum and genotype-phenotype correlation.
We documented clinical, neuroimaging, and morphologic data of 37 subjects from 27 families with PCH1. EXOSC3 gene sequencing was performed in 27 unrelated index patients of mixed ethnicity.
Biallelic mutations in EXOSC3 were detected in 10 of 27 families (37%). The most common mutation among all ethnic groups was c.395A>C, p.D132A, responsible for 11 (55%) of the 20 mutated alleles and ancestral in origin. The mutation-positive subjects typically presented with normal pregnancy, normal birth measurements, and relative preservation of brainstem and cortical structures. Psychomotor retardation was profound in all patients but lifespan was variable, with 3 subjects surviving beyond the late teens. Abnormal oculomotor function was commonly observed in patients surviving beyond the first year. Major clinical features previously reported in PCH1, including intrauterine abnormalities, postnatal hypoventilation and feeding difficulties, joint contractures, and neonatal death, were rarely observed in mutation-positive infants but were typical among the mutation-negative subjects.
EXOSC3 mutations account for 30%-40% of patients with PCH1 with variability in survival and clinical severity that is correlated with the genotype.
High-throughput DNA sequencing has become a mainstay for the discovery of genomic variants that may cause disease or affect phenotype. A next-generation sequencing pipeline typically identifies ...thousands of variants in each sample. A particular challenge is the annotation of each variant in a way that is useful to downstream consumers of the data, such as clinical sequencing centers or researchers. These users may require that all data storage and analysis remain on secure local servers to protect patient confidentiality or intellectual property, may have unique and changing needs to draw on a variety of annotation data sets and may prefer not to rely on closed-source applications beyond their control. Here we describe scalable methods for using the plugin capability of the Ensembl Variant Effect Predictor to enrich its basic set of variant annotations with additional data on genes, function, conservation, expression, diseases, pathways and protein structure, and describe an extensible framework for easily adding additional custom data sets.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
ABSTRACT
Objectives:
Congenital diarrhea disorders are a group of genetically diverse and typically autosomal recessive disorders that have yet to be well characterized phenotypically or molecularly. ...Diagnostic assessments are generally limited to nutritional challenges and histologic evaluation, and many subjects eventually require a prolonged course of intravenous nutrition. Here we describe next‐generation sequencing techniques to investigate a child with perplexing congenital malabsorptive diarrhea and other presumably unrelated clinical problems; this method provides an alternative approach to molecular diagnosis.
Methods:
We screened the diploid genome of an affected individual, using exome sequencing, for uncommon variants that have observed protein‐coding consequences. We assessed the functional activity of the mutant protein, as well as its lack of expression using immunohistochemistry.
Results:
Among several rare variants detected was a homozygous nonsense mutation in the catalytic domain of the proprotein convertase subtilisin/kexin type 1 gene. The mutation abolishes prohormone convertase 1/3 endoprotease activity as well as expression in the intestine. These primary genetic findings prompted a careful endocrine reevaluation of the child at 4.5 years of age, and multiple significant problems were subsequently identified consistent with the known phenotypic consequences of proprotein convertase subtilisin/kexin type 1 (PCSK1) gene mutations. Based on the molecular diagnosis, alternate medical and dietary management was implemented for diabetes insipidus, polyphagia, and micropenis.
Conclusions:
Whole‐exome sequencing provides a powerful diagnostic tool to clinicians managing rare genetic disorders with multiple perplexing clinical manifestations.
Next-generation sequencing (NGS) diagnostic assays increasingly are becoming the standard of care in oncology practice. As the scale of an NGS laboratory grows, management of these assays requires ...organizing large amounts of information, including patient data, laboratory processes, genomic data, as well as variant interpretation and reporting. Although several Laboratory Information Systems and/or Laboratory Information Management Systems are commercially available, they may not meet all of the needs of a given laboratory, in addition to being frequently cost-prohibitive. Herein, we present the System for Informatics in the Molecular Pathology Laboratory (SIMPL), a free and open-source Laboratory Information System/Laboratory Information Management System for academic and nonprofit molecular pathology NGS laboratories, developed at the Genomic and Molecular Pathology Division at the University of Chicago Medicine. SIMPL was designed as a modular end-to-end information system to handle all stages of the NGS laboratory workload from test order to reporting. We describe the features of SIMPL, its clinical validation at University of Chicago Medicine, and its installation and testing within a different academic center laboratory (University of Colorado), and we propose a platform for future community co-development and interlaboratory data sharing.
Next-generation sequencing (NGS) diagnostic assays increasingly are becoming the standard of care in oncology practice. As the scale of an NGS laboratory grows, management of these assays requires ...organizing large amounts of information, including patient data, laboratory processes, genomic data, as well as variant interpretation and reporting. Although several Laboratory Information Systems and/or Laboratory Information Management Systems are commercially available, they may not meet all of the needs of a given laboratory, in addition to being frequently cost-prohibitive. Herein, we present the System for Informatics in the Molecular Pathology Laboratory (SIMPL), a free and open-source Laboratory Information System/Laboratory Information Management System for academic and nonprofit molecular pathology NGS laboratories, developed at the Genomic and Molecular Pathology Division at the University of Chicago Medicine. SIMPL was designed as a modular end-to-end information system to handle all stages of the NGS laboratory workload from test order to reporting. We describe the features of SIMPL, its clinical validation at University of Chicago Medicine, and its installation and testing within a different academic center laboratory (University of Colorado), and we propose a platform for future community co-development and interlaboratory data sharing.
High throughput, massively parallel DNA sequencing provides a powerful technology to study the human genome and to identify variations in DNA that cause disease. Sequencing the protein coding region ...of the genome (`whole-exome sequencing') is a cost effective method to search the part of the genome that is most likely to harbor disease related mutations. We developed software methods to process sequencing data and to annotate variants with data on genes, function, conservation, expression, diseases, pathways, and protein structure. We applied whole-exome sequencing to search for the molecular basis of disease in three projects: 1) a cohort of patients with congenital diarrheal disorders (CDDs); 2) a cohort of patients with congenital chronic intestinal pseudo-obstruction (CIPO) or the related disease, megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIH); and 3) four siblings with infantile pontocerebellar hypoplasia and spinal motor neuron degeneration. We sequenced 45 probands from diverse ethnic backgrounds who were diagnosed with a variety of CDDs of probable, but unknown genetic cause. Patients had been diagnosed with generalized malabsorptive diarrhea, selective nutrient malabsorption, secretory diarrhea, and infantile IBD. We found homozygous or compound heterozygous mutations, 25 of them novel, in genes known to be associated with CDDs in 27 cases (60%). The genes implicated were ADAM17, DGAT1, EPCAM, IL10RA, MALT1, MYO5B, NEUROG3, PCSK1, SI, SKIV2L, SLC26A3, and SLC5A. With whole-exome sequencing in a cohort of 20 patients with congenital CIPO or MMIH, we identified a subset of 10 cases with potentially damaging de-novo dominant acting mutations at highly conserved loci in the ACTG2 gene, encoding actin, gamma-enteric smooth muscle precursor, a protein essential to the functioning of muscle cells in the intestinal wall. By exome sequencing, we discovered rare recessive mutations in EXOSC3 (encoding exosome component 3) that were responsible for pontocerebellar hypoplasia and spinal motor neuron degeneration in the four probands, and identified identical and additional novel mutations in a large percentage of other children with the same disorder. In conclusion, we demonstrated that whole-exome sequencing is an effective approach for the identification of casual mutations in that may escape detection with standard practice involving a complex diagnostic workup and targeted gene sequencing.
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