Notch signaling determines and reinforces cell fate in bilaterally symmetric multicellular eukaryotes. Despite the involvement of Notch in many key developmental systems, human mutations in Notch signaling components have mainly been described in disorders with vascular and bone effects. Here, we report five heterozygous NOTCH1 variants in unrelated individuals with Adams-Oliver syndrome (AOS), a rare disease with major features of aplasia cutis of the scalp and terminal transverse limb defects. Using whole-genome sequencing in a cohort of 11 families lacking mutations in the four genes with known roles in AOS pathology (ARHGAP31, RBPJ, DOCK6, and EOGT), we found a heterozygous de novo 85 kb deletion spanning the NOTCH1 5' region and three coding variants (c.1285T>C [p.Cys429Arg], c.4487G>A [p.Cys1496Tyr], and c.5965G>A [p.Asp1989Asn]), two of which are de novo, in four unrelated probands. In a fifth family, we identified a heterozygous canonical splice-site variant (c.743-1 G>T) in an affected father and daughter. These variants were not present in 5,077 in-house control genomes or in public databases. In keeping with the prominent developmental role described for Notch1 in mouse vasculature, we observed cardiac and multiple vascular defects in four of the five families. We propose that the limb and scalp defects might also be due to a vasculopathy in NOTCH1-related AOS. Our results suggest that mutations in NOTCH1 are the most common cause of AOS and add to a growing list of human diseases that have a vascular and/or bony component and are caused by alterations in the Notch signaling pathway.

Purpose:To assess the potential of whole-genome sequencing (WGS) to replicate and augment results from conventional blood-based newborn screening (NBS). Methods:Research-generated WGS data from an ancestrally diverse cohort of 1,696 infants and both parents of each infant were analyzed for variants in 163 genes involved in disorders included or under discussion for inclusion in US NBS programs. WGS results were compared with results from state NBS and related follow-up testing.Results: NBS genes are generally well covered by WGS. There is a median of one (range: 0-6) database-annotated pathogenic variant in the NBS genes per infant. Results of WGS and NBS in detecting 28 state-screened disorders and four hemoglobin traits were concordant for 88.6% of true positives (n = 35) and 98.9% of true negatives (n = 45,757). Of the five infants affected with a state-screened disorder, WGS identified two whereas NBS detected four. WGS yielded fewer false positives than NBS (0.037 vs. 0.17%) but more results of uncertain significance (0.90 vs. 0.013%). Conclusion:WGS may help rule in and rule out NBS disorders, pinpoint molecular diagnoses, and detect conditions not amenable to current NBS assays.

Rubinstein-Taybi syndrome (RSTS) can be caused by heterozygous mutations or deletions involving CREBBP or, less commonly, EP300. To date, only 15 patients with EP300 mutations have been clinically described. Frequently reported manifestations in these patients include characteristic facial and limb features, varying degrees of neurocognitive dysfunction, and maternal preeclampsia. Other congenital anomalies are less frequently reported. We describe a child found to have a de novo EP300 mutation (c.4933C>T, predicted to result in p.Arg1645X) through research-based whole-genome sequencing of the family trio. The child's presentation involved dysmorphic features as well as unilateral renal agenesis, a myelomeningocele, and minor genitourinary anomalies. The involvement of congenital anomalies in all 16 clinically described patients with EP300 mutations (25% of which have been identified by "hypothesis free" methods, including microarray, exome, and whole-genome sequencing) is reviewed. In summary, genitourinary anomalies have been identified in 38%, cardiovascular anomalies in 25%, spinal/vertebral anomalies in 19%, other skeletal anomalies in 19%, brain anomalies in 13%, and renal anomalies in 6%. Our patient expands the phenotypic spectrum in EP300-related RSTS; this case demonstrates the evolving practice of clinical genomics related to increasing availability of genomic sequencing methods.

PurposeImmunodeficiency screening has been added to many state-directed newborn screening programs. The current methodology is limited to screening for severe T-cell lymphopenia disorders. We evaluated the potential of genomic sequencing to augment current newborn screening for immunodeficiency, including identification of non-T cell disorders.MethodsWe analyzed whole-genome sequencing (WGS) and clinical data from a cohort of 1,349 newborn-parent trios by genotype-first and phenotype-first approaches. For the genotype-first approach, we analyzed predicted protein-impacting variants in 329 immunodeficiency-related genes in the WGS data. As a phenotype-first approach, electronic health records were used to identify children with clinical features suggestive of immunodeficiency. Genomes of these children and their parents were analyzed using a separate pipeline for identification of candidate pathogenic variants for rare Mendelian disorders.ResultsWGS provides adequate coverage for most known immunodeficiency-related genes. 13,476 distinct variants and 8,502 distinct predicted protein-impacting variants were identified in this cohort; five individuals carried potentially pathogenic variants requiring expert clinical correlation. One clinically asymptomatic individual was found genomically to have complement component 9 deficiency. Of the symptomatic children, one was molecularly identified as having an immunodeficiency condition and two were found to have other molecular diagnoses.ConclusionNeonatal genomic sequencing can potentially augment newborn screening for immunodeficiency.

We describe a case of an infant presenting with intractable diarrhea who subsequently developed dilated cardiomyopathy, for whom a diagnosis was not initially achieved despite extensive clinical testing, including panel-based genetic testing. Research-based whole-genome sequences of the proband and both parents were analyzed by the SAVANNA pipeline, a variant prioritization strategy integrating features of variants, genes, and phenotypes, which was implemented using publicly available tools. Although the intestinal morphological abnormalities characteristic of congenital tufting enteropathy (CTE) were not observed in the initial clinical gastrointestinal tract biopsies of the proband, an intronic variant, EPCAM c.556-14A>G, previously identified as pathogenic for CTE, was found in the homozygous state. A newborn cousin of the proband also presenting with intractable diarrhea was found to carry the same homozygous EPCAM variant, and clinical testing revealed intestinal tufting and loss of EPCAM staining. This variant, however, was considered nonexplanatory for the proband's dilated cardiomyopathy, which could be a sequela of the child's condition and/or related to other genetic variants, which include de novo mutations in the genes NEDD4L and GSK3A and a maternally inherited SCN5A variant. This study illustrates three ways in which genomic sequencing can aid in the diagnosis of clinically challenging patients: differential diagnosis despite atypical clinical presentation, distinguishing the possibilities of a syndromic condition versus multiple conditions, and generating hypotheses for novel contributory genes.