An international effort towards developing standards for best practices in analysis, interpretation and reporting of clinical genome sequencing results in the CLARITY Challenge

BACKGROUND: There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data was donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance. RESULTS: A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization. CONCLUSIONS: The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups

Requirement of argininosuccinate lyase for systemic nitric oxide production

Nitric oxide (NO) is crucial in diverse physiological and pathological processes. We show that a hypomorphic mouse model of argininosuccinate lyase (encoded by Asl) deficiency has a distinct phenotype of multiorgan dysfunction and NO deficiency. Loss of Asl in both humans and mice leads to reduced NO synthesis, owing to both decreased endogenous arginine synthesis and an impaired ability to use extracellular arginine for NO production. Administration of nitrite, which can be converted into NO in vivo, rescued the manifestations of NO deficiency in hypomorphic Asl mice, and a nitric oxide synthase (NOS)-independent NO donor restored NO-dependent vascular reactivity in humans with ASL deficiency. Mechanistic studies showed that ASL has a structural function in addition to its catalytic activity, by which it contributes to the formation of a multiprotein complex required for NO production. Our data demonstrate a previously unappreciated role for ASL in NOS function and NO homeostasis. Hence, ASL may serve as a target for manipulating NO production in experimental models, as well as for the treatment of NO-related diseases

An international effort towards developing standards for best practices in analysis, interpretation and reporting of clinical genome sequencing results in the CLARITY Challenge

There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data were donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease-causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance. RESULTS: A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization. CONCLUSIONS: The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups

Argininosuccinate lyase is an essential regulator of nictric oxide homeostatis

Nitric Oxide (NO) is an essential signaling molecule for diverse physiological and disease processes. The regulation of NO flux has focused on the study of the three NO synthases (NOS), but their respective genetic deficiencies exhibit only modest phenotypes. In humans, the urea cycle disorder argininosuccinic aciduria (ASA) caused by deficiency of argininosuccinic acid lyase (ASL) shows systemic features that may reflect global dysregulation of NO homeostasis due to deficiency of intracellular arginine synthesis and/or inability to utilize extracellular arginine. A hypomorphic mouse model of Asl deficiency shows multi-system dysregulation of NO synthesis that is manifest at physiological, histological, and biochemical levels. Endothelial nitric oxide synthase (eNos) and Asl epistasis further supports the specificity of NO dysregulation in this model. Importantly, ASA patients have decreased biochemical markers and dynamic measures of NO production, while ASA fibroblasts cannot utilize extracellular arginine for NO synthesis. NO synthesis from arginine requires formation of a protein complex that includes HSP90, ASL, ASS, and NOS that is decreased in hypomorphic Asl mice. Together, these data show that channeling of both intracellular and extracellular arginine by ASL regulates systemic NO production within a novel metabolomic compartment that explains the arginine paradox

Genomic alterations that contribute to the development of isolated and non-isolated congenital diaphragmatic hernia

Background Congenital diaphragmatic hernia (CDH) is a life threatening birth defect. Most of the genetic factors that contribute to the development of CDH remain unidentified. Objective To identify genomic alterations that contribute to the development of diaphragmatic defects. Methods A cohort of 45 unrelated patients with CDH or diaphragmatic eventrations was screened for genomic alterations by array comparative genomic hybridisation or single nucleotide polymorphism based copy number analysis. Results Genomic alterations that were likely to have contributed to the development of CDH were identified in 8 patients. Inherited deletions of ZFPM2 were identified in 2 patients with isolated diaphragmatic defects and a large de novo 8q deletion overlapping the same gene was found in a patient with non-isolated CDH. A de novo microdeletion of chromosome 1q41q42 and two de novo microdeletions on chromosome 16p11.2 were identified in patients with non-isolated CDH. Duplications of distal 11q and proximal 13q were found in a patient with non-isolated CDH and a de novo single gene deletion of FZD2 was identified in a patient with a partial pentalogy of Cantrell phenotype. Conclusions Haploinsufficiency of ZFPM2 can cause dominantly inherited isolated diaphragmatic defects with incomplete penetrance. These data define a new minimal deleted region for CDH on 1q41q42, provide evidence for the existence of CDH related genes on chromosomes 16p11.2, 11q23-24 and 13q12, and suggest a possible role for FZD2 and Wnt signalling in pentalogy of Cantrell phenotypes. These results demonstrate the clinical utility of screening for genomic alterations in individuals with both isolated and non-isolated diaphragmatic defects

B: Requirement of argininosuccinate lyase for systemic nitric oxide production. Nat Med 2011, 17:1619–1626. doi:10.1186/2049-3002-1-12 Cite this article as: Zheng et al.: Reversed argininosuccinate lyase activity in fumarate hydratase-deficient cancer cel

a r t i c l e s nature medicine advance online publication l-arginine is the natural substrate of NOSs for generating NO. As a by-product of the NOS reaction, l-citrulline is formed from l-arginine. Within the cell, citrulline can be recycled back to arginine by the cytoplasmic enzymes argininosuccinate synthase (ASS1) and ASL, constituting the citrulline-NO cycle Many tissues and cell types contain ASS1 and ASL, providing a cell-autonomous mechanism for generating arginine ( Argininosuccinic aciduria (ASA) (Mendelian Inheritance of Man (MIM) 207900) is the second most common human urea cycle disorder (UCD) and is caused by deficiency of ASL. Individuals with ASA cannot generate arginine from citrulline. Despite early treatment and adequate metabolic control of hyperammonemia, people with ASA can still have persistent intellectual impairment, delayed motor skills 4,5 and progressive hepatic disease We hypothesized that ASL deficiency would lead to systemic NO deficiency. In addition, because of the inability of supplemental l-arginine to prevent long-term complications in patients with ASA, we investigated whether ASL has a more central role in cellular Nitric oxide (NO) is crucial in diverse physiological and pathological processes. We show that a hypomorphic mouse model of argininosuccinate lyase (encoded by Asl) deficiency has a distinct phenotype of multiorgan dysfunction and NO deficiency. Loss of Asl in both humans and mice leads to reduced NO synthesis, owing to both decreased endogenous arginine synthesis and an impaired ability to use extracellular arginine for NO production. Administration of nitrite, which can be converted into NO in vivo, rescued the manifestations of NO deficiency in hypomorphic Asl mice, and a nitric oxide synthase (NOS)-independent NO donor restored NO-dependent vascular reactivity in humans with ASL deficiency. Mechanistic studies showed that ASL has a structural function in addition to its catalytic activity, by which it contributes to the formation of a multiprotein complex required for NO production. Our data demonstrate a previously unappreciated role for ASL in NOS function and NO homeostasis. Hence, ASL may serve as a target for manipulating NO production in experimental models, as well as for the treatment of NO-related diseases