Comparing the Diagnostic Yield of Germline Exome Versus Panel Sequencing in the Diverse Population of the Texas KidsCanSeq Pediatric Cancer Study.

PURPOSE: To evaluate the relative diagnostic yield of clinical germline genomic tests in a diverse pediatric cancer population. PATIENTS AND METHODS: The KidsCanSeq study enrolled pediatric cancer patients across six sites in Texas. Germline analysis included both exome sequencing and a therapy-focused pediatric cancer gene panel. The results were categorized by participants demographics, the presence of pathogenic or likely pathogenic (P/LP) variants, and variants of uncertain significance (VUS) in cancer predisposition genes (CPGs). Pediatric actionable CPGs were defined as those with cancer surveillance recommendations during childhood. RESULTS: Cancer P/LP variants were reported by at least one platform in 103 of 578 (17.8%) participants of which 76 were dominant cancer genes (13.1%) with no significant differences by self-described race or Hispanic ethnicity. However, the proportion of participants with VUS was greater in Asian and African American participants (P = .0029). Diagnostic yield was 16.6% for exome versus 8.5% for panel (P < .0001) with 42 participants with concordant germline results. Exome-only results included P/LP variants in 30 different CPGs in 54 participants, whereas panel-only results included seven participants with a copy number or structural P/LP variants in CPGs. There was no significant difference in diagnostic yield limited to pediatric actionable CPGs (P = .6171). CONCLUSION: Approximately 18% of a diverse pediatric cancer population had germline diagnostic findings with 50% of P/LP variants reported by only one platform because of CPGs not on the targeted panel and copy number variants (CNVs)/rearrangements not reported by exome. Although diagnostic yields were similar in this diverse population, increases in VUS results were observed in Asian and African American populations. Given the clinical significance of CNVs/rearrangements in this cohort, detection is critical to optimize germline analysis of pediatric cancer populations.

Clinical and molecular features of pediatric cancer patients with Lynch syndrome.

BACKGROUND: The association of childhood cancer with Lynch syndrome is not established compared with the significant pediatric cancer risk in recessive constitutional mismatch repair deficiency syndrome (CMMRD). PROCEDURE: We describe the clinical features, germline analysis, and tumor genomic profiling of patients with Lynch syndrome among patients enrolled in pediatric cancer genomic studies. RESULTS: There were six of 773 (0.8%) pediatric patients with solid tumors identified with Lynch syndrome, defined as a germline heterozygous pathogenic variant in one of the mismatch repair (MMR) genes (three with MSH6, two with MLH1, and one with MSH2). Tumor analysis demonstrated evidence for somatic second hits and/or increased tumor mutation burden in three of four patients with available tumor with potential implications for therapy and identification of at-risk family members. Only one patient met current guidelines for pediatric cancer genetics evaluation at the time of tumor diagnosis. CONCLUSION: Approximately 1% of children with cancer have Lynch syndrome, which is missed with current referral guidelines, suggesting the importance of adding MMR genes to tumor and hereditary pediatric cancer panels. Tumor analysis may provide the first suggestion of an underlying cancer predisposition syndrome and is useful in distinguishing between Lynch syndrome and CMMRD.

Single-cell and bulk transcriptome sequencing identifies two epithelial tumor cell states and refines the consensus molecular classification of colorectal cancer.

The consensus molecular subtype (CMS) classification of colorectal cancer is based on bulk transcriptomics. The underlying epithelial cell diversity remains unclear. We analyzed 373,058 single-cell transcriptomes from 63 patients, focusing on 49,155 epithelial cells. We identified a pervasive genetic and transcriptomic dichotomy of malignant cells, based on distinct gene expression, DNA copy number and gene regulatory network. We recapitulated these subtypes in bulk transcriptomes from 3,614 patients. The two intrinsic subtypes, iCMS2 and iCMS3, refine CMS. iCMS3 comprises microsatellite unstable (MSI-H) cancers and one-third of microsatellite-stable (MSS) tumors. iCMS3 MSS cancers are transcriptomically more similar to MSI-H cancers than to other MSS cancers. CMS4 cancers had either iCMS2 or iCMS3 epithelium; the latter had the worst prognosis. We defined the intrinsic epithelial axis of colorectal cancer and propose a refined 'IMF' classification with five subtypes, combining intrinsic epithelial subtype (I), microsatellite instability status (M) and fibrosis (F).

De novo variants in H3-3A and H3-3B are associated with neurodevelopmental delay, dysmorphic features, and structural brain abnormalities.

The histone H3 variant H3.3, encoded by two genes H3-3A and H3-3B, can replace canonical isoforms H3.1 and H3.2. H3.3 is important in chromatin compaction, early embryonic development, and lineage commitment. The role of H3.3 in somatic cancers has been studied extensively, but its association with a congenital disorder has emerged just recently. Here we report eleven de novo missense variants and one de novo stop-loss variant in H3-3A (n = 6) and H3-3B (n = 6) from Baylor Genetics exome cohort (n = 11) and Matchmaker Exchange (n = 1), of which detailed phenotyping was conducted for 10 individuals (H3-3A = 4 and H3-3B = 6) that showed major phenotypes including global developmental delay, short stature, failure to thrive, dysmorphic facial features, structural brain abnormalities, hypotonia, and visual impairment. Three variant constructs (p.R129H, p.M121I, and p.I52N) showed significant decrease in protein expression, while one variant (p.R41C) accumulated at greater levels than wild-type control. One H3.3 variant construct (p.R129H) was found to have stronger interaction with the chaperone death domain-associated protein 6.

Compound heterozygous KCTD7 variants in progressive myoclonus epilepsy.

KCTD7 is a member of the potassium channel tetramerization domain-containing protein family and has been associated with progressive myoclonic epilepsy (PME), characterized by myoclonus, epilepsy, and neurological deterioration. Here we report four affected individuals from two unrelated families in which we identified KCTD7 compound heterozygous single nucleotide variants through exome sequencing. RNAseq was used to detect a non-annotated splicing junction created by a synonymous variant in the second family. Whole-cell patch-clamp analysis of neuroblastoma cells overexpressing the patients' variant alleles demonstrated aberrant potassium regulation. While all four patients experienced many of the common clinical features of PME, they also showed variable phenotypes not previously reported, including dysautonomia, brain pathology findings including a significantly reduced thalamus, and the lack of myoclonic seizures. To gain further insight into the pathogenesis of the disorder, zinc finger nucleases were used to generate kctd7 knockout zebrafish. Kctd7 homozygous mutants showed global dysregulation of gene expression and increased transcription of c-fos, which has previously been correlated with seizure activity in animal models. Together these findings expand the known phenotypic spectrum of KCTD7-associated PME, report a new animal model for future studies, and contribute valuable insights into the disease.

Functional and structural analysis of cytokine-selective IL6ST defects that cause recessive hyper-IgE syndrome.

BACKGROUND: Biallelic variants in IL6ST, encoding GP130, cause a recessive form of hyper-IgE syndrome (HIES) characterized by high IgE level, eosinophilia, defective acute phase response, susceptibility to bacterial infections, and skeletal abnormalities due to cytokine-selective loss of function in GP130, with defective IL-6 and IL-11 and variable oncostatin M (OSM) and IL-27 levels but sparing leukemia inhibitory factor (LIF) signaling. OBJECTIVE: Our aim was to understand the functional and structural impact of recessive HIES-associated IL6ST variants. METHODS: We investigated a patient with HIES by using exome, genome, and RNA sequencing. Functional assays assessed IL-6, IL-11, IL-27, OSM, LIF, CT-1, CLC, and CNTF signaling. Molecular dynamics simulations and structural modeling of GP130 cytokine receptor complexes were performed. RESULTS: We identified a patient with compound heterozygous novel missense variants in IL6ST (p.Ala517Pro and the exon-skipping null variant p.Gly484_Pro518delinsArg). The p.Ala517Pro variant resulted in a more profound IL-6- and IL-11-dominated signaling defect than did the previously identified recessive HIES IL6ST variants p.Asn404Tyr and p.Pro498Leu. Molecular dynamics simulations suggested that the p.Ala517Pro and p.Asn404Tyr variants result in increased flexibility of the extracellular membrane-proximal domains of GP130. We propose a structural model that explains the cytokine selectivity of pathogenic IL6ST variants that result in recessive HIES. The variants destabilized the conformation of the hexameric cytokine receptor complexes, whereas the trimeric LIF-GP130-LIFR complex remained stable through an additional membrane-proximal interaction. Deletion of this membrane-proximal interaction site in GP130 consequently caused additional defective LIF signaling and Stüve-Wiedemann syndrome. CONCLUSION: Our data provide a structural basis to understand clinical phenotypes in patients with IL6ST variants.

Clinical experience with carrier screening in a general population: support for a comprehensive pan-ethnic approach.

PURPOSE: To present results from a large cohort of individuals receiving expanded carrier screening (CS) in the United States. METHODS: Single-gene disorder carrier status for 381,014 individuals was determined using next-generation sequencing (NGS) based CS for up to 274 genes. Detection rates were compared with literature-reported values derived from disease prevalence and carrier frequencies. Combined theoretical affected pregnancy rates for the 274 screened disorders were calculated. RESULTS: For Ashkenazi Jewish (AJ) diseases, 81.6% (4434/5435) of carriers identified did not report AJ ancestry. For cystic fibrosis, 44.0% (6260/14,229) of carriers identified had a variant not on the standard genotyping panel. Individuals at risk of being a silent spinal muscular atrophy carrier, not detectable by standard screening, comprised 1/39 (8763/344,407) individuals. For fragile X syndrome, compared with standard premutation screening, AGG interruption analysis modified risk in 83.2% (1128/1356) premutation carriers. Assuming random pairing across the study population, approximately 1/175 pregnancies would be affected by a disorder in the 274-gene screening panel. CONCLUSION: Compared with standard screening, NGS-based CS provides additional information that may impact reproductive choices. Pan-ethnic CS leads to substantially increased identification of at-risk couples. These data support offering NGS-based CS to all reproductive-aged women.

Multiple Myeloma DREAM Challenge reveals epigenetic regulator PHF19 as marker of aggressive disease.

While the past decade has seen meaningful improvements in clinical outcomes for multiple myeloma patients, a subset of patients does not benefit from current therapeutics for unclear reasons. Many gene expression-based models of risk have been developed, but each model uses a different combination of genes and often involves assaying many genes making them difficult to implement. We organized the Multiple Myeloma DREAM Challenge, a crowdsourced effort to develop models of rapid progression in newly diagnosed myeloma patients and to benchmark these against previously published models. This effort lead to more robust predictors and found that incorporating specific demographic and clinical features improved gene expression-based models of high risk. Furthermore, post-challenge analysis identified a novel expression-based risk marker, PHF19, which has recently been found to have an important biological role in multiple myeloma. Lastly, we show that a simple four feature predictor composed of age, ISS, and expression of PHF19 and MMSET performs similarly to more complex models with many more gene expression features included.

Characterization of the renal phenotype in RMND1-related mitochondrial disease.

BACKGROUND: The nuclear encoded gene RMND1 (Required for Meiotic Nuclear Division 1 homolog) has recently been linked to RMND1-related mitochondrial disease (RRMD). This autosomal recessive condition characteristically presents with an infantile-onset multisystem disease characterized by severe hypotonia, global developmental delay, failure to thrive, sensorineural hearing loss, and lactic acidosis. Renal disease, however, appears to be one of the more prominent features of RRMD, affecting patients at significantly higher numbers compared to other mitochondrial diseases. We report the clinical, histological, and molecular findings of four RRMD patients across three academic institutions with a focus on the renal manifestations. METHODS: Four patients were identified for the purpose of this study, all of whom had molecular confirmation at the time of inclusion, which included the common pathogenic variant c.713A>G (p.N238S) as well as the three rare variants: c.485delC (p.P162fs), c.533C>T (p.T178M), and c.1317 + 1G>C splice donor variant. Medical history and laboratory findings were collected from the medical records and medical providers. RESULTS: In this study, all four patients developed renal disease characterized as tubulopathy (3/4), renal tubular acidosis (2/4), interstitial nephritis (1/4), and/or end-stage renal disease (4/4) necessitating renal transplantation (2/4). Histological evaluation of renal biopsy specimens revealed generalized tubular atrophy and on electron microscopy, abundant mitochondria with pleomorphism and abnormal cristae. CONCLUSION: Our experience with RRMD demonstrates a specific pattern of renal disease manifestations and clinical course. Patients are unlikely to respond to traditional chronic kidney disease (CKD) treatments, making early diagnosis and consideration of renal transplantation paramount to the management of RRMD.

Extracutaneous manifestations in phacomatosis cesioflammea and cesiomarmorata: Case series and literature review.

Phacomatosis pigmentovascularis (PPV) comprises a family of rare conditions that feature vascular abnormalities and melanocytic lesions that can be solely cutaneous or multisystem in nature. Recently published work has demonstrated that both vascular and melanocytic abnormalities in PPV of the cesioflammea and cesiomarmorata subtypes can result from identical somatic mosaic activating mutations in the genes GNAQ and GNA11. Here, we present three new cases of PPV with features of the cesioflammea and/or cesiomarmorata subtypes and mosaic mutations in GNAQ or GNA11. To better understand the risk of potentially occult complications faced by such patients we additionally reviewed 176 cases published in the literature. We report the frequency of clinical findings, their patterns of co-occurrence as well as published recommendations for surveillance after diagnosis. Features assessed include: capillary malformation; dermal and ocular melanocytosis; glaucoma; limb asymmetry; venous malformations; and central nervous system (CNS) anomalies, such as ventriculomegaly and calcifications. We found that ocular findings are common in patients with phacomatosis cesioflammea and cesiomarmorata. Facial vascular involvement correlates with a higher risk of seizures (p = .0066). Our genetic results confirm the role of mosaic somatic mutations in GNAQ and GNA11 in phacomatosis cesioflammea and cesiomarmorata. Their clinical and molecular findings place these conditions on a clinical spectrum encompassing other GNAQ and GNA11 related disorders and inform recommendations for their management.

Bi-allelic Variants in TONSL Cause SPONASTRIME Dysplasia and a Spectrum of Skeletal Dysplasia Phenotypes.

SPONASTRIME dysplasia is an autosomal-recessive spondyloepimetaphyseal dysplasia characterized by spine (spondylar) abnormalities, midface hypoplasia with a depressed nasal bridge, metaphyseal striations, and disproportionate short stature. Scoliosis, coxa vara, childhood cataracts, short dental roots, and hypogammaglobulinemia have also been reported in this disorder. Although an autosomal-recessive inheritance pattern has been hypothesized, pathogenic variants in a specific gene have not been discovered in individuals with SPONASTRIME dysplasia. Here, we identified bi-allelic variants in TONSL, which encodes the Tonsoku-like DNA repair protein, in nine subjects (from eight families) with SPONASTRIME dysplasia, and four subjects (from three families) with short stature of varied severity and spondylometaphyseal dysplasia with or without immunologic and hematologic abnormalities, but no definitive metaphyseal striations at diagnosis. The finding of early embryonic lethality in a Tonsl-/- murine model and the discovery of reduced length, spinal abnormalities, reduced numbers of neutrophils, and early lethality in a tonsl-/- zebrafish model both support the hypomorphic nature of the identified TONSL variants. Moreover, functional studies revealed increased amounts of spontaneous replication fork stalling and chromosomal aberrations, as well as fewer camptothecin (CPT)-induced RAD51 foci in subject-derived cell lines. Importantly, these cellular defects were rescued upon re-expression of wild-type (WT) TONSL; this rescue is consistent with the hypothesis that hypomorphic TONSL variants are pathogenic. Overall, our studies in humans, mice, zebrafish, and subject-derived cell lines confirm that pathogenic variants in TONSL impair DNA replication and homologous recombination-dependent repair processes, and they lead to a spectrum of skeletal dysplasia phenotypes with numerous extra-skeletal manifestations.

Non-invasive prenatal sequencing for multiple Mendelian monogenic disorders using circulating cell-free fetal DNA.

Current non-invasive prenatal screening is targeted toward the detection of chromosomal abnormalities in the fetus1,2. However, screening for many dominant monogenic disorders associated with de novo mutations is not available, despite their relatively high incidence3. Here we report on the development and validation of, and early clinical experience with, a new approach for non-invasive prenatal sequencing for a panel of causative genes for frequent dominant monogenic diseases. Cell-free DNA (cfDNA) extracted from maternal plasma was barcoded, enriched, and then analyzed by next-generation sequencing (NGS) for targeted regions. Low-level fetal variants were identified by a statistical analysis adjusted for NGS read count and fetal fraction. Pathogenic or likely pathogenic variants were confirmed by a secondary amplicon-based test on cfDNA. Clinical tests were performed on 422 pregnancies with or without abnormal ultrasound findings or family history. Follow-up studies on cases with available outcome results confirmed 20 true-positive, 127 true-negative, zero false-positive, and zero-false negative results. The initial clinical study demonstrated that this non-invasive test can provide valuable molecular information for the detection of a wide spectrum of dominant monogenic diseases, complementing current screening for aneuploidies or carrier screening for recessive disorders.

Clinical exome sequencing reveals locus heterogeneity and phenotypic variability of cohesinopathies.

PURPOSE: Defects in the cohesin pathway are associated with cohesinopathies, notably Cornelia de Lange syndrome (CdLS). We aimed to delineate pathogenic variants in known and candidate cohesinopathy genes from a clinical exome perspective. METHODS: We retrospectively studied patients referred for clinical exome sequencing (CES, N = 10,698). Patients with causative variants in novel or recently described cohesinopathy genes were enrolled for phenotypic characterization. RESULTS: Pathogenic or likely pathogenic single-nucleotide and insertion/deletion variants (SNVs/indels) were identified in established disease genes including NIPBL (N = 5), SMC1A (N = 14), SMC3 (N = 4), RAD21 (N = 2), and HDAC8 (N = 8). The phenotypes in this genetically defined cohort skew towards the mild end of CdLS spectrum as compared with phenotype-driven cohorts. Candidate or recently reported cohesinopathy genes were supported by de novo SNVs/indels in STAG1 (N = 3), STAG2 (N = 5), PDS5A (N = 1), and WAPL (N = 1), and one inherited SNV in PDS5A. We also identified copy-number deletions affecting STAG1 (two de novo, one of unknown inheritance) and STAG2 (one of unknown inheritance). Patients with STAG1 and STAG2 variants presented with overlapping features yet without characteristic facial features of CdLS. CONCLUSION: CES effectively identified disease-causing alleles at the mild end of the cohensinopathy spectrum and enabled characterization of candidate disease genes.

De Novo Missense Variants in TRAF7 Cause Developmental Delay, Congenital Anomalies, and Dysmorphic Features.

TRAF7 is a multi-functional protein involved in diverse signaling pathways and cellular processes. The phenotypic consequence of germline TRAF7 variants remains unclear. Here we report missense variants in TRAF7 in seven unrelated individuals referred for clinical exome sequencing. The seven individuals share substantial phenotypic overlap, with developmental delay, congenital heart defects, limb and digital anomalies, and dysmorphic features emerging as key unifying features. The identified variants are de novo in six individuals and comprise four distinct missense changes, including a c.1964G>A (p.Arg655Gln) variant that is recurrent in four individuals. These variants affect evolutionarily conserved amino acids and are located in key functional domains. Gene-specific mutation rate analysis showed that the occurrence of the de novo variants in TRAF7 (p = 2.6 × 10-3) and the recurrent de novo c.1964G>A (p.Arg655Gln) variant (p = 1.9 × 10-8) in our exome cohort was unlikely to have occurred by chance. In vitro analyses of the observed TRAF7 mutations showed reduced ERK1/2 phosphorylation. Our findings suggest that missense mutations in TRAF7 are associated with a multisystem disorder and provide evidence of a role for TRAF7 in human development.

Fabry disease revisited: Management and treatment recommendations for adult patients.

Fabry disease is an X-linked lysosomal storage disorder caused by mutations in the GLA gene leading to deficient α-galactosidase A activity, glycosphingolipid accumulation, and life-threatening complications. Phenotypes vary from the "classic" phenotype, with pediatric onset and multi-organ involvement, to later-onset, a predominantly cardiac phenotype. Manifestations are diverse in female patients in part due to variations in residual enzyme activity and X chromosome inactivation patterns. Enzyme replacement therapy (ERT) and adjunctive treatments can provide significant clinical benefit. However, much of the current literature reports outcomes after late initiation of ERT, once substantial organ damage has already occurred. Updated monitoring and treatment guidelines for pediatric patients with Fabry disease have recently been published. Expert physician panels were convened to develop updated, specific guidelines for adult patients. Management of adult patients depends on 1) a personalized approach to care, reflecting the natural history of the specific disease phenotype; 2) comprehensive evaluation of disease involvement prior to ERT initiation; 3) early ERT initiation; 4) thorough routine monitoring for evidence of organ involvement in non-classic asymptomatic patients and response to therapy in treated patients; 5) use of adjuvant treatments for specific disease manifestations; and 6) management by an experienced multidisciplinary team.

Biallelic Variants in OTUD6B Cause an Intellectual Disability Syndrome Associated with Seizures and Dysmorphic Features.

Ubiquitination is a posttranslational modification that regulates many cellular processes including protein degradation, intracellular trafficking, cell signaling, and protein-protein interactions. Deubiquitinating enzymes (DUBs), which reverse the process of ubiquitination, are important regulators of the ubiquitin system. OTUD6B encodes a member of the ovarian tumor domain (OTU)-containing subfamily of deubiquitinating enzymes. Herein, we report biallelic pathogenic variants in OTUD6B in 12 individuals from 6 independent families with an intellectual disability syndrome associated with seizures and dysmorphic features. In subjects with predicted loss-of-function alleles, additional features include global developmental delay, microcephaly, absent speech, hypotonia, growth retardation with prenatal onset, feeding difficulties, structural brain abnormalities, congenital malformations including congenital heart disease, and musculoskeletal features. Homozygous Otud6b knockout mice were subviable, smaller in size, and had congenital heart defects, consistent with the severity of loss-of-function variants in humans. Analysis of peripheral blood mononuclear cells from an affected subject showed reduced incorporation of 19S subunits into 26S proteasomes, decreased chymotrypsin-like activity, and accumulation of ubiquitin-protein conjugates. Our findings suggest a role for OTUD6B in proteasome function, establish that defective OTUD6B function underlies a multisystemic human disorder, and provide additional evidence for the emerging relationship between the ubiquitin system and human disease.

Germline mutations in ABL1 cause an autosomal dominant syndrome characterized by congenital heart defects and skeletal malformations.

ABL1 is a proto-oncogene well known as part of the fusion gene BCR-ABL1 in the Philadelphia chromosome of leukemia cancer cells. Inherited germline ABL1 changes have not been associated with genetic disorders. Here we report ABL1 germline variants cosegregating with an autosomal dominant disorder characterized by congenital heart disease, skeletal abnormalities, and failure to thrive. The variant c.734A>G (p.Tyr245Cys) was found to occur de novo or cosegregate with disease in five individuals (families 1-3). Additionally, a de novo c.1066G>A (p.Ala356Thr) variant was identified in a sixth individual (family 4). We overexpressed the mutant constructs in HEK 293T cells and observed increased tyrosine phosphorylation, suggesting increased ABL1 kinase activities associated with both the p.Tyr245Cys and p.Ala356Thr substitutions. Our clinical and experimental findings, together with previously reported teratogenic effects of selective BCR-ABL inhibitors in humans and developmental defects in Abl1 knockout mice, suggest that ABL1 has an important role during organismal development.

Lessons learned from additional research analyses of unsolved clinical exome cases.

BACKGROUND: Given the rarity of most single-gene Mendelian disorders, concerted efforts of data exchange between clinical and scientific communities are critical to optimize molecular diagnosis and novel disease gene discovery. METHODS: We designed and implemented protocols for the study of cases for which a plausible molecular diagnosis was not achieved in a clinical genomics diagnostic laboratory (i.e. unsolved clinical exomes). Such cases were recruited to a research laboratory for further analyses, in order to potentially: (1) accelerate novel disease gene discovery; (2) increase the molecular diagnostic yield of whole exome sequencing (WES); and (3) gain insight into the genetic mechanisms of disease. Pilot project data included 74 families, consisting mostly of parent-offspring trios. Analyses performed on a research basis employed both WES from additional family members and complementary bioinformatics approaches and protocols. RESULTS: Analysis of all possible modes of Mendelian inheritance, focusing on both single nucleotide variants (SNV) and copy number variant (CNV) alleles, yielded a likely contributory variant in 36% (27/74) of cases. If one includes candidate genes with variants identified within a single family, a potential contributory variant was identified in a total of ~51% (38/74) of cases enrolled in this pilot study. The molecular diagnosis was achieved in 30/63 trios (47.6%). Besides this, the analysis workflow yielded evidence for pathogenic variants in disease-associated genes in 4/6 singleton cases (66.6%), 1/1 multiplex family involving three affected siblings, and 3/4 (75%) quartet families. Both the analytical pipeline and the collaborative efforts between the diagnostic and research laboratories provided insights that allowed recent disease gene discoveries (PURA, TANGO2, EMC1, GNB5, ATAD3A, and MIPEP) and increased the number of novel genes, defined in this study as genes identified in more than one family (DHX30 and EBF3). CONCLUSION: An efficient genomics pipeline in which clinical sequencing in a diagnostic laboratory is followed by the detailed reanalysis of unsolved cases in a research environment, supplemented with WES data from additional family members, and subject to adjuvant bioinformatics analyses including relaxed variant filtering parameters in informatics pipelines, can enhance the molecular diagnostic yield and provide mechanistic insights into Mendelian disorders. Implementing these approaches requires collaborative clinical molecular diagnostic and research efforts.

Renal cell carcinoma harboring somatic TSC2 mutations in a child with methylmalonic acidemia.

Pediatric renal cell carcinoma (RCC) is a rare cancer that can be associated with inherited diseases including tuberous sclerosis complex (TSC) caused by germline mutations in TSC1 or TSC2. Somatic mutations in TSC1 and TSC2 have also been reported in adult RCC, which predict response to mTOR inhibitors. Here, we present the first case of RCC in a child with methylmalonic acidemia (MMA). Clinical whole exome sequencing of blood and tumor samples confirmed the diagnosis of MMA and revealed two somatic inactivating mutations in TSC2, suggesting the potential consideration of an mTOR inhibitor in the event of tumor recurrence.