The Clinical Variant Analysis Tool: Analyzing the evidence supporting reported genomic variation in clinical practice.

PURPOSE: Genomic test results, regardless of laboratory variant classification, require clinical practitioners to judge the applicability of a variant for medical decisions. Teaching and standardizing clinical interpretation of genomic variation calls for a methodology or tool. METHODS: To generate such a tool, we distilled the Clinical Genome Resource framework of causality and the American College of Medical Genetics/Association of Molecular Pathology and Quest Diagnostic Laboratory scoring of variant deleteriousness into the Clinical Variant Analysis Tool (CVAT). Applying this to 289 clinical exome reports, we compared the performance of junior practitioners with that of experienced medical geneticists and assessed the utility of reported variants. RESULTS: CVAT enabled performance comparable to that of experienced medical geneticists. In total, 124 of 289 (42.9%) exome reports and 146 of 382 (38.2%) reported variants supported a diagnosis. Overall, 10.5% (1 pathogenic [P] or likely pathogenic [LP] variant and 39 variants of uncertain significance [VUS]) of variants were reported in genes without established disease association; 20.2% (23 P/LP and 54 VUS) were in genes without sufficient phenotypic concordance; 7.3% (15 P/LP and 13 VUS) conflicted with the known molecular disease mechanism; and 24% (91 VUS) had insufficient evidence for deleteriousness. CONCLUSION: Implementation of CVAT standardized clinical interpretation of genomic variation and emphasized the need for collaborative and transparent reporting of genomic variation.

Can tandem alternative splicing and evasion of premature termination codon surveillance contribute to attenuated Peutz-Jeghers syndrome?

Alternative use of short distance tandem sites such as NAGNn AG are a common mechanism of alternative splicing; however, single nucleotide variants are rarely reported as likely to generate or to disrupt tandem splice sites. We identify a pathogenic intron 5 STK11 variant (NM_000455.4:c.[735-6A>G];[=]) segregating with the mucocutaneous features but not the hamartomatous polyps of Peutz-Jeghers syndrome in two individuals. By RNAseq analysis of peripheral blood mRNA, this variant was shown to generate a novel and preferentially used tandem proximal splice acceptor (AAGTGAAG). The variant transcript (NM_000455.4:c.734_734 + 1insTGAAG), which encodes a frameshift (p.[Tyr246Glufs*43]) constituted 36%-43% of STK11 transcripts suggesting partial escape from nonsense mediated mRNA decay and translation of a truncated protein. A review of the ClinVar database identified other similar variants. We suggest that nucleotide changes creating or disrupting tandem alternative splice sites are a pertinent disease mechanism and require contextualization for clinical reporting. Additionally, we hypothesize that some pathogenic STK11 variants cause an attenuated phenotype.

Bi-allelic ATG4D variants are associated with a neurodevelopmental disorder characterized by speech and motor impairment.

Autophagy regulates the degradation of damaged organelles and protein aggregates, and is critical for neuronal development, homeostasis, and maintenance, yet few neurodevelopmental disorders have been associated with pathogenic variants in genes encoding autophagy-related proteins. We report three individuals from two unrelated families with a neurodevelopmental disorder characterized by speech and motor impairment, and similar facial characteristics. Rare, conserved, bi-allelic variants were identified in ATG4D, encoding one of four ATG4 cysteine proteases important for autophagosome biogenesis, a hallmark of autophagy. Autophagosome biogenesis and induction of autophagy were intact in cells from affected individuals. However, studies evaluating the predominant substrate of ATG4D, GABARAPL1, demonstrated that three of the four ATG4D patient variants functionally impair ATG4D activity. GABARAPL1 is cleaved or "primed" by ATG4D and an in vitro GABARAPL1 priming assay revealed decreased priming activity for three of the four ATG4D variants. Furthermore, a rescue experiment performed in an ATG4 tetra knockout cell line, in which all four ATG4 isoforms were knocked out by gene editing, showed decreased GABARAPL1 priming activity for the two ATG4D missense variants located in the cysteine protease domain required for priming, suggesting that these variants impair the function of ATG4D. The clinical, bioinformatic, and functional data suggest that bi-allelic loss-of-function variants in ATG4D contribute to the pathogenesis of this syndromic neurodevelopmental disorder.

The practice of genomic medicine: A delineation of the process and its governing principles.

Genomic medicine, an emerging medical discipline, applies the principles of evolution, developmental biology, functional genomics, and structural genomics within clinical care. Enabling widespread adoption and integration of genomic medicine into clinical practice is key to achieving precision medicine. We delineate a biological framework defining diagnostic utility of genomic testing and map the process of genomic medicine to inform integration into clinical practice. This process leverages collaboration and collective cognition of patients, principal care providers, clinical genomic specialists, laboratory geneticists, and payers. We detail considerations for referral, triage, patient intake, phenotyping, testing eligibility, variant analysis and interpretation, counseling, and management within the utilitarian limitations of health care systems. To reduce barriers for clinician engagement in genomic medicine, we provide several decision-making frameworks and tools and describe the implementation of the proposed workflow in a prototyped electronic platform that facilitates genomic care. Finally, we discuss a vision for the future of genomic medicine and comment on areas for continued efforts.

Familial Hypomagnesemia With Secondary Hypocalcemia: A Case Report.

Familial hypomagnesemia with secondary hypocalcemia is a rare genetic disorder of magnesium metabolism that presents with refractory seizures during infancy. It is caused by loss-of-function mutations in the gene encoding transient receptor potential cation channel member 6 (TRPM6). Herein we report an infant who presented with refractory seizures that were brought under control by normalizing the magnesium level. Genetic analysis revealed a nonsense variant in the TRPM6 gene. Our case highlights the importance of evaluation for familial hypomagnesemia in any child with recurrent or refractory seizures.

Recurrent invasive group A streptococcal infection with four-limb amputation in an immunocompetent child.

We report a previously well paediatric patient with two distinct presentations of invasive group A streptococcus (GAS) infection resulting in significant morbidity. The first episode, following GAS pharyngitis, involved multiorgan dysfunction syndrome. This included cardiorespiratory and acute hepatorenal failure and purpura fulminans that eventually necessitated four-limb amputation. The second episode occurred 12 months later, from undetermined aetiology, and resulted in septic shock. Molecular analysis of the emm gene and PCR for Serum Opacity Factor revealed that the initial isolate was M Type 4 and sof gene positive while the second isolate was M Type 1 and sof gene negative. Immunological investigations, including CH50, quantitative IgA, IgM and IgG, and flow cytometry measuring lymphocyte subsets, and vaccine response to measles, mumps, rubella and pneumococcus were normal. This is the first report of recurrent bacteraemia from different strains of Streptococcus pyogenes infection in an apparently immunocompetent child.