A retrospective cohort analysis of the Yale pediatric genomics discovery program.

The Pediatric Genomics Discovery Program (PGDP) at Yale uses next-generation sequencing (NGS) and translational research to evaluate complex patients with a wide range of phenotypes suspected to have rare genetic diseases. We conducted a retrospective cohort analysis of 356 PGDP probands evaluated between June 2015 and July 2020, querying our database for participant demographics, clinical characteristics, NGS results, and diagnostic and research findings. The three most common phenotypes among the entire studied cohort (n = 356) were immune system abnormalities (n = 105, 29%), syndromic or multisystem disease (n = 103, 29%), and cardiovascular system abnormalities (n = 62, 17%). Of 216 patients with final classifications, 77 (36%) received new diagnoses and 139 (64%) were undiagnosed; the remaining 140 patients were still actively being investigated. Monogenetic diagnoses were found in 67 (89%); the largest group had variants in known disease genes but with new contributions such as novel variants (n = 31, 40%) or expanded phenotypes (n = 14, 18%). Finally, five PGDP diagnoses (8%) were suggestive of novel gene-to-phenotype relationships. A broad range of patients can benefit from single subject studies combining NGS and functional molecular analyses. All pediatric providers should consider further genetics evaluations for patients lacking precise molecular diagnoses.

DYNC1H1-related disorders: A description of four new unrelated patients and a comprehensive review of previously reported variants.

Heterozygous variants in the DYNC1H1 gene have been associated chiefly with intellectual disability (ID), malformations in cortical development (MCD), spinal muscular atrophy (SMA), and Charcot-Marie-Tooth axonal type 20 (CMT), with fewer reports describing other intersecting phenotypes. To better characterize the variable syndromes associated with DYNC1H1, we undertook a detailed analysis of reported patients in the medical literature through June 30, 2019. In sum we identified 200 patients from 143 families harboring 103 different DYNC1H1 variants, and added reports for four unrelated patients identified at our center, three with novel variants. The most common features associated with DYNC1H1 were neuromuscular (NM) disease (largely associated with variants in the stem domain), ID with MCD (largely associated with variants in the motor domain), or a combination of these phenotypes. Despite these trends, exceptions are noted throughout. Overall, DYNC1H1 is associated with variable neurodevelopmental and/or neuromuscular phenotypes that overlap. To avoid confusion DYNC1H1 disorders may be best categorized at this time by more general descriptions rather than phenotype-specific nomenclature such as SMA or CMT. We therefore propose the terms: DYNC1H1-related NM disorder, DYNC1H1-related CNS disorder, and DYNC1H1-related combined disorder. Our single center's experience may be evidence that disease-causing variants in this gene are more prevalent than currently recognized.

Y265C DNA polymerase beta knockin mice survive past birth and accumulate base excision repair intermediate substrates.

DNA is susceptible to damage by a wide variety of chemical agents that are generated either as byproducts of cellular metabolism or exposure to man-made and harmful environments. Therefore, to maintain genomic integrity, having reliable DNA repair systems is important. DNA polymerase ß is known to be a key player in the base excision repair pathway, and mice devoid of DNA polymerase beta do not live beyond a few hours after birth. In this study, we characterized mice harboring an impaired pol ß variant. This Y265C pol ß variant exhibits slow DNA polymerase activity but WT lyase activity and has been shown to be a mutator polymerase. Mice expressing Y265C pol ß are born at normal Mendelian ratios. However, they are small, and 60% die within a few hours after birth. Slow proliferation and significantly increased levels of cell death are observed in many organs of the E14 homozygous embryos compared with WT littermates. Mouse embryo fibroblasts prepared from the Y265C pol ß embryos proliferate at a rate slower than WT cells and exhibit a gap-filling deficiency during base excision repair. As a result of this, chromosomal aberrations and single- and double-strand breaks are present at significantly higher levels in the homozygous mutant versus WT mouse embryo fibroblasts. This is study in mice is unique in that two enzymatic activities of pol ß have been separated; the data clearly demonstrate that the DNA polymerase activity of pol ß is essential for survival and genome stability.

A novel variant in MAP3K7 associated with an expanded cardiospondylocarpofacial syndrome phenotype.

The transforming growth factor-ß-activated kinase 1 (TAK1) encoded by mitogen-activated protein kinase kinase kinase 7 (MAP3K7) is widely expressed and participates in multiple molecular and cellular processes, including growth, differentiation, inflammation, and apoptosis. Pathogenic variants in MAP3K7 have recently been associated with two disorders: cardiospondylocarpofacial syndrome (CSCFS) and frontometaphyseal dysplasia 2 (FMD2). To date, all small in-frame deletions and splice variants in MAP3K7 have been associated with CSCFS, whereas missense variants have been reported in both CSCFS and FMD2. Here, we present a patient with a novel heterozygous likely pathogenic variant, c.125_127del, p.(Val42del), in MAP3K7, only the sixth variant associated with CSCFS to be described in the literature. Although this patient has a phenotype that is most consistent with that of CSCFS, including valvular heart disease, short stature, fusions of the spine and bones of the hands and feet, and certain facial dysmorphisms, he interestingly has some features reported previously in FMD2 but not CSCFS. These include flexion contractures of the elbow and widely spaced first and second toes, highlighting new points of overlap between these two syndromes. We additionally point out features in the patient presented here that are rare but recurrent among CSCFS patients previously reported in the literature, as well as a new distinctive cutaneous finding not previously reported.

Identification of a novel MYOC variant in a Hispanic family with early-onset primary open-angle glaucoma with elevated intraocular pressure.

Primary open-angle glaucoma (POAG) is the leading cause of irreversible blindness worldwide. Most cases are multifactorial in etiology, but some are associated with variants in the myocilin gene, MYOC Here, we report the identification of a novel MYOC variant, c.1153G>A, in a 24-yr-old female patient with a personal and family history of juvenile/early-onset POAG. Further genetic testing within her family demonstrated that this variant segregates with the POAG phenotype in an autosomal dominant pattern. Identification of this MYOC variant in multiple affected relatives provides evidence for its pathogenicity, supporting previous findings linking MYOC mutations, in particular in the third exon's olfactomedin domain, to juvenile-onset POAG. This case also emphasizes the potential value of genetic testing in families with histories of eye disorders.

A multiplex assay for the detection and mapping of complex glycerol kinase deficiency.

BACKGROUND: Glycerol kinase deficiency (GKD) is an X-linked recessive disorder that presents in both isolated and complex forms. The contiguous deletion that leads to GKD also commonly affects NR0B1 (DAX1), the gene associated with adrenal hypoplasia congenita, and DMD, the Duchenne muscular dystrophy gene. Molecular testing to delineate this deletion is expensive and has only limited availability. METHODS: We designed a multiplex PCR assay for the detection and mapping of a contiguous deletion potentially affecting the IL1RAPL1, NR0B1, GK, and DMD genes in a 29-month-old male patient with GKD. RESULTS: Multiplex PCR detected a contiguous deletion that involved the IL1RAPL1, NR0B1, GK, and DMD genes. Although the patient had a creatine kinase concentration within the reference interval, further mapping with PCR revealed that exon 74 was the last intact exon at the 3' end of the DMD gene. CONCLUSIONS: Multiplex PCR is an effective and inexpensive way to detect and map the contiguous deletion in cases of complex GKD. The extension of a deletion to include DMD exon 75 in a patient with a creatine kinase concentration within the reference interval suggests that this region of the gene may not be essential for protein function.

Cardiac and CNS defects in a mouse with targeted disruption of suppressor of fused.

The hedgehog (Hh) pathway is conserved from Drosophila to humans and plays a key role in embryonic development. In addition, activation of the pathway in somatic cells contributes to cancer development in several tissues. Suppressor of fused is a negative regulator of Hh signaling. Targeted disruption of the murine suppressor of fused gene (Sufu) led to a phenotype that included neural tube defects and lethality at mid-gestation (9.0-10.5 dpc). This phenotype resembled that caused by loss of patched (Ptch1), another negative regulator of the Hh pathway. Consistent with this finding, Ptch1 and Sufu mutants displayed excess Hh signaling and resultant altered dorsoventral patterning of the neural tube. Sufu mutants also had abnormal cardiac looping, indicating a defect in the determination of left-right asymmetry. Marked expansion of nodal expression in 7.5 dpc embryos and variable degrees of node dysmorphology in 7.75 dpc embryos suggested that the pathogenesis of the cardiac developmental abnormalities was related to node development. Other mutants of the Hh pathway, such as Shh, Smo and Shh/Ihh compound mutants, also have laterality defects. In contrast to Ptch1 heterozygous mice, Sufu heterozygotes had no developmental defects and no apparent tumor predisposition. The resemblance of Sufu homozygotes to Ptch1 homozygotes is consistent with mouse Sufu being a conserved negative modulator of Hh signaling.