Germline pathogenic variants in CDKN2A predispose to various cancers, including melanoma, pancreatic cancer, and neural system tumors, whereas CDKN2B variants are associated with renal cell carcinoma. A few case reports have described heterozygous germline deletions spanning both CDKN2A and CDKN2B associated with a cancer predisposition syndrome (CPS) that constitutes a risk of cancer beyond those associated with haploinsufficiency of each gene individually, indicating an additive effect or a contiguous gene deletion syndrome. We report a young woman with a de novo germline 9p21 microdeletion involving the CDKN2A/CDKN2B genes, who developed six primary cancers since childhood, including a very rare extraskeletal osteosarcoma (eOS) at the age of 8. To our knowledge this is the first report of eOS in a patient with CDKN2A/CDKN2B deletion.

Here we describe for the first time double paternal uniparental isodisomy (iUPD) 7 and 15 in a baby boy with features in the Beckwith–Wiedemann syndrome spectrum (BWSp) (placentomegaly, hyperinsulinism, enlarged viscera, hemangiomas, and earlobe creases) in addition to conjugated hyperbilirubinemia. His phenotype was also reminiscent of genome-wide paternal uniparental isodisomy. We discuss the most likely origin of the UPDs: a maternal double monosomy 7 and 15 rescued by duplication of the paternal chromosomes after fertilization. So far, paternal UPD7 is not associated with an abnormal phenotype, whereas paternal UPD15 causes Angelman syndrome. Methylation analysis for other clinically relevant imprinting disorders, including BWSp, was normal. Therefore, we hypothesized that the double UPD affected other imprinted genes. To look for such effects, patient fibroblast RNA was isolated and analyzed for differential expression compared to six controls. We did not find apparent transcription differences in imprinted genes outside Chromosomes 7 and 15 in patient fibroblast. PEG10 (7q21.3) was the only paternally imprinted gene on these chromosomes up-regulated beyond double-dose expectation (sixfold). We speculate that a high PEG10 level could have a growth-promoting effect as his phenotype was not related to aberrations in BWS locus on 11p15.5 after DNA, RNA, and methylation testing. However, many genes in gene sets associated with growth were up-regulated. This case broadens the phenotypic spectrum of UPDs but does not show evidence of involvement of an imprinted gene network.

Sclerosing epithelioid fibrosarcoma (SEF) is a rare and aggressive soft-tissue sarcoma thought to originate in fibroblasts of the tissues comprising tendons, ligaments, and muscles. Minimally responsive to conventional cytotoxic chemotherapies, >50% of SEF patients experience local recurrence and/or metastatic disease. SEF is most commonly discovered in middle-aged and elderly adults, but also rarely in children. A common gene fusion occurring between the EWSR1 and CREB3L1 genes has been observed in 80%–90% of SEF cases. We describe here the youngest SEF patient reported to date (a 3-yr-old Caucasian male) who presented with numerous bony and lung metastases. Additionally, we perform a comprehensive literature review of all SEF-related articles published since the disease was first characterized. Finally, we describe the generation of an SEF primary cell line, the first such culture to be reported. The patient described here experienced persistent disease progression despite aggressive treatment including multiple resections, radiotherapy, and numerous chemotherapies and targeted therapeutics. Untreated and locally recurrent tumor and metastatic tissue were sequenced by whole-genome, whole-exome, and deep-transcriptome next-generation sequencing with comparison to a patient-matched normal blood sample. Consistent across all sequencing analyses was the disease-defining EWSR1–CREB3L1 fusion as a single feature consensus. We provide an analysis of our genomic findings and discuss potential therapeutic strategies for SEF.

Somatic mutations in hotspot regions of the cytosolic or mitochondrial isoforms of the isocitrate dehydrogenase gene (IDH1 and IDH2, respectively) contribute to the pathogenesis of acute myeloid leukemia (AML) by producing the oncometabolite 2-hydroxyglutarate (2-HG). The allosteric IDH1 inhibitor, ivosidenib, suppresses 2-HG production and induces clinical responses in relapsed/refractory IDH1-mutant AML. Herein, we describe a clinical case of AML in which we detected the neomorphic IDH1 p.R132C mutation in consecutive patient samples with a mutational hotspot targeted next-generation sequencing (NGS) assay. The patient had a clinical response to ivosidenib, followed by relapse and disease progression. Subsequent sequencing of the relapsed sample using a newly developed all-exon, hybrid-capture-based NGS panel identified an additional IDH1 p.S280F mutation known to cause renewed 2-HG production and drug resistance. Structural modeling confirmed that serine-to-phenylalanine substitution at this codon sterically hinders ivosidenib from binding to the mutant IDH1 dimer interface and predicted a similar effect on the pan-IDH inhibitor AG-881. Joint full-exon NGS and structural modeling enables monitoring IDH1 inhibitor-treated AML patients for acquired drug resistance and choosing follow-up therapy.

Wilson disease is a medically actionable rare autosomal recessive disorder of defective copper excretion caused by mutations in ATP7B, one of two highly evolutionarily conserved copper-transporting ATPases. Hundreds of disease-causing variants in ATP7B have been reported to public databases; more than half of these are missense changes, and a significant proportion are presumed unequivocal loss-of-function variants (nonsense, frameshift, and canonical splice site). Current molecular genetic testing includes sequencing all coding exons (±10 bp) as well as deletion/duplication testing, with reported sensitivity of >98%. We report a proband from a consanguineous family with a biochemical phenotype consistent with early-onset Wilson disease who tested negative on conventional molecular genetic testing. Using a combination of whole-genome sequencing and transcriptome sequencing, we found that the proband's disease is due to skipping of exons 6–7 of the ATP7B gene associated with a novel intronic variant (NM_000053.4:c.1947-19T > A) that alters a putative splicing enhancer element. This variant was also homozygous in the proband's younger sister, whose subsequent clinical evaluations revealed biochemical evidence of Wilson disease. Our work adds to emerging evidence that ATP7B exon skipping from deep intronic variants outside typical splice junctions is an important mechanism of Wilson disease; the variants responsible may elude standard genetic testing.

Patients harboring germline pathogenic biallelic variants in genes involved in the recognition and repair of DNA damage are known to have a substantially increased cancer risk. Emerging evidence suggests that individuals harboring heterozygous variants in these same genes may also be at heightened, albeit lesser, risk for cancer. Herein, we sought to determine whether heterozygous variants in RECQL4, the gene encoding an essential DNA helicase that is defective in children with the autosomal recessive cancer-predisposing condition Rothmund–Thomson syndrome (RTS), are associated with increased risk for childhood cancer. To address this question, we interrogated germline sequence data from 4435 pediatric cancer patients at St. Jude Children's Research Hospital and 1127 from the National Cancer Institute Therapeutically Applicable Research to Generate Effective Treatment (TARGET) database and identified 24 (0.43%) who harbored loss-of-function (LOF) RECQL4 variants, including five of 249 (2.0%) with osteosarcoma (OS). These RECQL4 variants were significantly overrepresented in children with OS, the cancer most frequently observed in patients with RTS, as compared to 134,187 noncancer controls in the Genome Aggregation Database (gnomAD v2.1; P = 0.00087, odds ratio [OR] = 7.1, 95% CI, 2.9–17). Nine of the 24 (38%) individuals possessed the same c.1573delT (p.Cys525Alafs) variant located in the highly conserved DNA helicase domain, suggesting that disruption of this domain is central to oncogenesis. Altogether these data expand our understanding of the genetic factors predisposing to childhood cancer and reveal a novel association between heterozygous RECQL4 LOF variants and development of pediatric OS.

Gliomatosis peritonei is a rare pathologic finding that is associated with ovarian teratomas and malignant mixed germ cell tumors. The occurrence of gliomatosis as a mature glial implant can impart an improved prognosis to patients with immature ovarian teratoma, making prompt and accurate diagnosis important. We describe a case of recurrent immature teratoma in a 10-yr-old female patient, in which comparative analysis of the RNA sequencing gene expression data from the patient's tumor was used effectively to aid in the diagnosis of gliomatosis peritonei.

Exome sequencing is increasingly being used to help diagnose pediatric neurology cases when clinical presentations are not specific. However, interpretation of equivocal results that include variants of uncertain significance remains a challenge. In those cases, follow-up testing and clinical correlation can help clarify the clinical relevance of the molecular findings. In this report, we describe the diagnostic odyssey of a 4-year-old girl who presented with global developmental delay and seizures, with leukodystrophy seen on MRI. Clinical evaluation, MRI, and comprehensive metabolic testing were performed, followed by whole-exome sequencing (WES), parental testing, follow-up testing, and retrospective detailed clinical evaluation. WES identified two candidate causative pathogenic variants in SAMHD1, a gene associated with the recessive condition Aicardi–Goutières syndrome (AGS) type 5 (OMIM 612952): a previously reported pathogenic variant NM_015474 c.602T>A (p.I201N), maternally inherited, and a rare missense variant of uncertain significance, c.1293A>T(p.L431F). Analysis of type I interferon-related biomarkers demonstrated that the patient has an interferon signature characteristic of AGS. Retrospective detailed clinical evaluation showed that the girl has a phenotype consistent with AGS5, a rare neurological condition. These results further define the phenotypic spectrum associated with specific SAMHD1 variants, including heterozygous variants in AGS carriers, and support the idea that autoinflammatory dysregulation is part of the disease pathophysiology. More broadly, this work highlights the issues and methodology involved in ascribing clinical relevance to interpretation of variants detected by WES.

FGFR2 is recurrently amplified in 5% of gastric cancers and 1%–4% of breast cancers; however, this molecular alteration has never been reported in a primary colorectal cancer specimen. Preclinical studies indicate that several FGFR tyrosine-kinase inhibitors (TKIs), such as AZD4547, have in vitro activity against the FGFR2-amplified colorectal cell line, NCI-H716. The efficacy of these inhibitors is currently under investigation in clinical trials for breast and gastric cancer. Thus, better characterizing colorectal tumors for FGFR2 amplification could identify a subset of patients who may benefit from FGFR TKI therapies. Here, we describe a novel FGFR2 amplification identified by clinical next-generation sequencing in a primary colorectal cancer. Further characterization of the tumor by immunohistochemistry showed neuroendocrine differentiation, similar to the reported properties of the NCI-H716 cell line. These findings demonstrate that the spectrum of potentially clinically actionable mutations detected by targeted clinical sequencing panels is not limited to only single-nucleotide polymorphisms and insertions/deletions but also to copy-number alterations.

Pathogenic variants in EBF3 were recently described in three back-to-back publications in association with a novel neurodevelopmental disorder characterized by intellectual disability, speech delay, ataxia, and facial dysmorphisms. In this report, we describe an additional patient carrying a de novo missense variant in EBF3 (c.487C>T, p.(Arg163Trp)) that falls within a conserved residue in the zinc knuckle motif of the DNA binding domain. Without a solved structure of the DNA binding domain, we generated a homology-based atomic model and performed molecular dynamics simulations for EBF3, which predicted decreased DNA affinity for p.(Arg163Trp) compared with wild-type protein and control variants. These data are in agreement with previous experimental studies of EBF1 showing the paralogous residue is essential for DNA binding. The conservation and experimental evidence existing for EBF1 and in silico modeling and dynamics simulations to validate comparable behavior of multiple variants in EBF3 demonstrates strong support for the pathogenicity of p.(Arg163Trp). We show that our patient presents with phenotypes consistent with previously reported patients harboring EBF3 variants and expands the phenotypic spectrum of this newly identified disorder with the additional feature of a bicornuate uterus.