Prenatal diagnosis of SMPD4 loss - A neurodevelopmental disorder with microcephaly, arthrogryposis and structural brain anomalies.

ACMG SF v3.0 list for reporting of secondary findings in clinical exome and genome sequencing: a policy statement of the American College of Medical Genetics and Genomics (ACMG)

Genetic counseling in direct-to-consumer exome sequencing: a case report.

Biallelic pathogenic variants in MADD lead to a very rare neurodevelopmental disorder which is phenotypically pleiotropic grossly ranging from severe neonatal hypotonia, failure to thrive, multiple organ dysfunction, and early lethality to a similar but milder phenotype with better survival. Here, we report 5 patients from 3 unrelated Egyptian families in whom 4 patients showed the severe end of the spectrum displaying neonatal respiratory distress, hypotonia and chronic diarrhea while one patient presented with the mild form displaying moderate intellectual disability and myopathy. In addition, we observed distal arthrogryposis and nonspecific structural brain anomalies in all our patients. Interestingly, cerebellar and brainstem hypoplasia were noted in one patient. Whole exome sequencing identified three novel homozygous variants in the MADD gene: two likely pathogenic [c.4321delC p.(Gln1441ArgfsTer46) and c.2620 C > T p.(Arg874Ter)] and one variant of uncertain significance (c.4307 G > A, p.Arg1436Gln). The variants segregated with the disease in all available family members. Our findings confirm that arthrogryposis, genital, cardiac and structural brain anomalies are manifestations of MADD which expand the spectrum of MADD-related neurodevelopmental disorder. Moreover, they further highlight the convergence of MADD variants on different organ systems leading to complex phenotypes.

Whole Exome Sequencing and Extended Thrombophilia Testing in Patients with Venous Thromboembolism

Sphingomyelin phosphodiesterase 4 (SMPD4) encodes a member of the Mg2+-dependent, neutral sphingomyelinase family that catalyzes the hydrolysis of the phosphodiester bond of sphingomyelin to form phosphorylcholine and ceramide. Recent studies have revealed that biallelic loss-of-function variants of SMPD4 cause syndromic neurodevelopmental disorders characterized by microcephaly, congenital arthrogryposis, and structural brain anomalies. In this study, three novel loss-of-function SMPD4 variants were identified using exome sequencing (ES) in two independent patients with developmental delays, microcephaly, seizures, and brain structural abnormalities. Patient 1 had a homozygous c.740_741del, p.(Val247Glufs*21) variant and showed profound intellectual disability, hepatomegaly, a simplified gyral pattern, and a thin corpus callosum without congenital dysmorphic features. Patient 2 had a compound heterozygous nonsense c.2124_2125del, p.(Phe709*) variant and splice site c.1188+2dup variant. RNA analysis revealed that the c.1188+2dup variant caused exon 13 skipping, leading to a frameshift (p.Ala406Ser*6). In vitro transcription analysis using minigene system suggested that mRNA transcribed from mutant allele may be degraded by nonsense-mediated mRNA decay system. He exhibited diverse manifestations, including growth defects, muscle hypotonia, respiratory distress, arthrogryposis, insulin-dependent diabetes mellitus, sensorineural hearing loss, facial dysmorphism, and various brain abnormalities, including cerebral atrophy, hypomyelination, and cerebellar hypoplasia. Here, we review previous literatures and discuss the phenotypic diversity of SMPD4-related disorders.

Simple SummaryInherited predisposition to hematological malignancies is more common than previously perceived. The main objective of our study was to analyze the whole-exome sequencing data for the genomic characterization of sixteen patients with a strong family or personal onco-hematological history. When a duo analysis was performed, we detected pathogenic or likely pathogenic (P/LP) germline variants in four out of the six families studied. In the remaining four individuals, we detected three P/LP germline variants in genes with a potential role in cancer development. Next-generation sequencing strategies lead to the identification of novel candidate genes (NFATC2 and TC2N) potentially involved in the development of these germline syndromes. The recognition of predisposing variants is crucial for disease management and follow-up of affected patients and their relatives.The familial occurrence of hematological malignancies has been underappreciated. Recent studies suggest that up to 15% of adults with myeloid neoplasms carry germline pathogenic variants in cancer-predisposing genes. This study aimed to identify the underlying germline predisposition variant in patients with a strong family or personal onco-hematological history using whole exome sequencing on sixteen uncharacterized individuals. It was carried out in two groups of patients, one with samples available from two affected relatives (Cohort A) and one with available samples from the index case (Cohort B). In Cohort A, six families were characterized. Two families shared variants in genes associated with DNA damage response and involved in cancer development (CHEK2 and RAD54L). Pathogenic or likely pathogenic germline variants were also found in novel candidate genes (NFATC2 and TC2N). In two families, any relevant pathogenic or likely pathogenic genomic variants were identified. In Cohort B, four additional index cases were analyzed. Three of them harbor clinically relevant variants in genes with a probable role in the development of inherited forms of hematological malignancies (GATA1, MSH4 and PRF1). Overall, whole exome sequencing is a useful approach to achieve a further characterization of these patients and their mutational spectra. Moreover, further investigations may help improve optimization for disease management of affected patients and their families.

Whole exome sequencing in a cohort of familial premature ovarian insufficiency cases reveals a broad array of pathogenic or likely pathogenic variants in 50% of families

P3-09-05: Clinical outcome of patients with advanced triple negative breast cancer with germline and somatic variants in homologous recombination gene

Background: Patients with neurodevelopmental and neuromuscular disorders often show overlapping clinical phenotypes. Pathogenic variants in KMT5B, a histone lysine methyltransferase, have been linked to neurodevelopmental disorders, yet their effects on human skeletal muscle remain unexplored. We report on a patient with KMT5B-linked disease who presented to a neuromuscular specialty clinic with significant involvement of skeletal muscle, where a multi-omics approach established the genetic diagnosis and revealed neuromuscular findings relevant for diagnosis, care and rehabilitation. Methods: Whole-exome sequencing was performed from blood and data was analyzed using the RD-Connect Genome Phenome Analysis Platform. Histological analysis and proteomic profiling were performed on muscle tissue. Results: Whole-exome sequencing revealed a pathogenic heterozygous variant (c.554_557del, p.Tyr185Cysfs*27) in KMT5B. Histological examination revealed fiber-type grouping, angular fibers, increased fast-twitch fiber proportion, and lipid droplet accumulation, indicative of muscle denervation. Proteomic profiling identified 77 dysregulated proteins, including upregulation of sarcomeric proteins, mitochondrial and glycolytic enzymes, acute-phase and complement factors, and extracellular matrix components, reflecting structural remodeling, metabolic adaptation, and inflammatory activation. These findings align with the role types observed in Kmt5b mouse models, supporting a role of KMT5B in neuromuscular function. Conclusions: We present the first combined histological and proteomic analysis of quadriceps muscle from a patient carrying a pathogenic KMT5B variant with a neuromuscular phenotype. The convergence of histological and proteomic alterations suggests that KMT5B haploinsufficiency may be associated with fiber-type shifts, denervation, and metabolic stress in human skeletal muscle. Understanding these processes provides mechanistic insight into motor deficits and informs targeted therapeutic strategies, including physiotherapeutic interventions, and early compensatory measures.

In children and adolescents, osteosarcoma (OS) is the most common malignant bone tumor. OS occurs in certain cancer predisposition syndromes at a higher than expected frequency. However, outside of these rare syndromes, OS is significantly more common and its genetic etiology remains poorly understood. We conducted an evaluation of rare exonic variants in 545 unselected OS cases compared with 1061 cancer-free controls using whole-exome sequencing of blood or buccal cell DNA to estimate the prevalence and burden of rare deleterious germline variants. We assessed potentially pathogenic rare variants in 126 established cancer predisposing genes (CPG) and known somatically mutated genes. We also surveyed the exome for genes with a higher burden of rare variants in 342 EUR cases with 994 EUR controls sequenced at the same time. Rare genetic variants, defined by a MAF <0.01 in 1000G, ESP or ExAC, that passed quality control filters were classified based on well-annotated databases (IARC-TP53, Telomerase Database, ARUP, BIC, Insight and COSMIC-hotspots), variant databases (e.g. Clinvar, HGMD and LOVD), and in silico predictions (Sift, Polyphen, Mutation assessor, MutationTaster, FATHMM and LTR). Rare variants matching known pathogenic variants, or inexact matches at the same residue, in selected well-annotated databases were labeled as putative pathogenic variants. High impact variants (frameshift, stop gain/loss, or known splice sites) not matching a previously identified pathogenic variant in the selected databases were categorized as probably pathogenic. Predictions based solely on in silico predictions were categorized as possibly pathogenic, uncertain or likely benign. A total of 14.5% of cases had a predicted pathogenic or high impact variant in an autosomal dominant CPG. A significantly higher pathogenic rare variant burden was present in EUR cases compared with EUR controls (Pburden=3.7x10-05). TP53 had the highest prevalence of pathogenic mutations (5% of EUR cases; Pburden=1.05x10-09). CDKN2A, MEN1, MLH1, MUTYH, PALB2 and VHL had a significantly higher rare variant burden in the EUR cases. 7.5% of cases had a predicted pathogenic or probably pathogenic variant in an autosomal recessive CPG (Pburden=5.4x10-03). Additionally, two males had a pathogenic variant in the X-linked genes, DKC1 and GPC3. In total, 21% of cases had a predicted pathogenic or high impact variant in a CPG (Pburden=2.4x10-07). Exome-wide analysis identified two novel genes with significantly higher rare variant burdens in the cases compared to controls. In conclusion, several CPGs and two novel genes, not previously associated with OS, had an enrichment of rare variants in OS and warrant further follow-up. Our results indicate that a clinically significant fraction of OS cases may harbor one or more mutations worthy of consideration for further investigation and genetic counseling. Citation Format: Roelof Koster, Bin Zhu, Meredith Yeager, Michael Dean, Matthew Gianferante, Lei Song, Joshua Sampson, NCI DCEG Cancer Genomics Research Laboratory, Julie Gastier-Foster, Richard Gorlick, Silvia Regina Caminada de Toledo, Antonio Petrilli, Ana Patiño-Garcia, Fernando Lecanda, Massimo Serra, Claudia Hattinger, Piero Picci, Katia Scotlandi, Adrienne Flanagan, Roberto Tirabosco, Maria Amary, Nilgün Kurucu, Inci Ergurhan Ilhan, Neriman Sari, Mandy Ballinger, David Thomas, Donald Barkauskas, Belynda Hicks, Margaret Tucker, Neil Caporaso, Robert Hoover, Stephen Chanock, Sharon Savage, Lisa Mirabello. Whole-exome sequencing identifies a high frequency of germline deleterious variants in cancer predisposition genes in individuals with osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4871. doi:10.1158/1538-7445.AM2017-4871

BackgroundPathogenic PAK1 variants were described to be causative of neurodevelopmental disorder with macrocephaly, seizures, and speech delay. Herein, we present a de novo PAK1 variant combine with a de novo terminal 1q microdeletion in a Chinese pediatric patient, aiming to provide more insights into the underlying genotype–phenotype relationship.MethodsEnrolled in this study was a 6-year-old girl with clinical features of global developmental delay, severe intellectual disability, speech delay, and seizures from Quanzhou region of China. Karyotype and chromosomal microarray analysis (CMA) were performed to detect chromosome abnormalities in this family. Whole exome sequencing (WES) was performed to investigate additional genetic variants in this family.ResultsNo chromosomal abnormalities were elicited from the entire family by karyotype analysis. Further familial CMA results revealed that the patient had a de novo 2.7-Mb microdeletion (arr[GRCh37] 1q44(246,454,321_249,224,684) × 1]) in 1q44 region, which contains 14 OMIM genes, but did not overlap the reported smallest region of overlap (SRO) responsible for the clinical features in 1q43q44 deletion syndrome. In addition, WES result demonstrated a de novo NM_002576: c.251C > G (p.T84R) variant in PAK1 gene in the patient, which was interpreted as a likely pathogenic variant.ConclusionIn this study, we identify a novel PAK1 variant associated with a terminal 1q microdeletion in a patient with neurodevelopmental disorder. In addition, we believe that the main clinical features may ascribe to the pathogenic variant in PAK1 gene in the patient.

Impact of clinical exomes in neurodevelopmental and neurometabolic disorders.

Autism spectrum disorders (ASD) represent a phenotypically heterogeneous group of patients that strongly intertwine with other neurodevelopmental disorders (NDDs), with genetics playing a significant role in their etiology. Whole exome sequencing (WES) has become predominant in molecular diagnostics for ASD by considerably increasing the diagnostic yield. However, the proportion of undiagnosed patients still remains high due to complex clinical presentation, reduced penetrance, and lack of segregation analysis or clinical information. Thus, reverse phenotyping, where we first identified a possible genetic cause and then determine its clinical relevance, has been shown to be a more efficient approach. WES was performed on 147 Slovenian pediatric patients with suspected ASD. Data analysis was focused on identifying ultrarare or “single event” variants in ASD-associated genes and further expanded to NDD-associated genes. Protein function and gene prioritization were performed on detected clinically relevant variants to determine their role in ASD etiology and phenotype. Reverse phenotyping revealed a pathogenic or likely pathogenic variant in ASD-associated genes in 20.4% of patients, with subsequent segregation analysis indicating that 14 were de novo variants and 1 was presumed compound heterozygous. The diagnostic yield was further increased by 2.7% by the analysis of ultrarare or “single event” variants in all NDD-associated genes. Protein function analysis established that genes in which variants of unknown significance (VUS) were detected were predominantly the cause of intellectual disability (ID), and in most cases, features of ASD as well. Using such an approach, variants in rarely described ASD-associated genes, such as SIN3B, NR4A2, and GRIA1, were detected. By expanding the analysis to include functionally similar NDD genes, variants in KCNK9, GNE, and other genes were identified. These would probably have been missed by classic genotype–phenotype analysis. Our study thus demonstrates that in patients with ASD, analysis of ultrarare or “single event” variants obtained using WES with the inclusion of functionally similar genes and reverse phenotyping obtained a higher diagnostic yield despite limited clinical data. The present study also demonstrates that most of the causative genes in our cohort were involved in the syndromic form of ASD and confirms their comorbidity with other developmental disorders.

Backgrounds and Purpose- Although new methods for genetic analyses are rapidly evolving, there are currently knowledge gaps in how to detect Mendelian forms of stroke. Methods- We performed whole-exome sequencing in 22 probands, under 56 years at their first ischemic stroke episode, from multi-incident stroke families. With the use of a comprehensive stroke-gene panel, we searched for variants in stroke-related genes. The probands' clinical stroke subtype was related to clinical characteristics previously associated with pathogenic variants in these genes. Relatives were genotyped in 7 families to evaluate stroke-gene variants of unknown significance. In 2 larger families with embolic stroke of unknown source, whole-exome sequencing was performed in additional members to examine the possibility of identifying new stroke genes. Results- Six of 22 probands carried pathogenic or possibly pathogenic variants in genes reported to be associated with their stroke subtype. A known pathogenic variant in NOTCH3 and a possibly pathogenic variant in ACAD9 gene were identified. A novel JAK2:c.3188G>A (p.Arg1063His) mutation was seen in a proband with embolic stroke of undetermined source and prothrombotic status. However, penetrance in the family was incomplete. COL4A2:c.3368A>G (p.Glu1123Gly) was detected in 2 probands but did not cosegregate with the disease in their families. Whole-exome sequencing in multiple members of 2 pedigrees with embolic stroke of undetermined source revealed possibly pathogenic variants in genes not previously associated with stroke, GPR142:c.148C>G (p.Leu50Val), and PTPRN2:c.2416A>G (p.Ile806Val); LRRC1 c.808A>G (p.Ile270Val), SLC7A10c.1294dupG (p.Val432fs), IKBKB: c.1070C>T (p.Ala357Val), and OXGR1 c.392G>A (p.Arg131His), respectively. Conclusions- Screening with whole-exome sequencing using a comprehensive stroke-gene panel may identify rare monogenic forms of stroke, but careful evaluation of clinical characteristics and potential pathogenicity of novel variants remain important. In our study, the majority of individuals with familial aggregation of stroke lacked any identified genetic causes.

ObjectiveWe describe the clinical characteristics and genetic etiology of several new cases within the ACO2‐related disease spectrum. Mitochondrial aconitase (ACO2) is a nuclear‐encoded tricarboxylic acid cycle enzyme. Homozygous pathogenic missense variants in the ACO2 gene were initially associated with infantile degeneration of the cerebrum, cerebellum, and retina, resulting in profound intellectual and developmental disability and early death. Subsequent studies have identified a range of homozygous and compound heterozygous pathogenic missense, nonsense, frameshift, and splice‐site ACO2 variants in patients with a spectrum of clinical manifestations and disease severities.MethodsWe describe a cohort of five novel patients with biallelic pathogenic variants in ACO2. We review the clinical histories of these patients as well as the molecular and functional characterization of the associated ACO2 variants and compare with those described previously in the literature.ResultsTwo siblings with relatively mild symptoms presented with episodic ataxia, mild developmental delays, severe dysarthria, and behavioral abnormalities including hyperactivity and depressive symptoms with generalized anxiety. One patient presented with the classic form with cerebellar hypoplasia, ataxia, seizures, optic atrophy, and retinitis pigmentosa. Another unrelated patient presented with ataxia but developed severe progressive spastic quadriplegia. Another patient demonstrated a spinal muscular atrophy‐like presentation with severe neonatal hypotonia, diminished reflexes, and poor respiratory drive, leading to ventilator dependence until death at the age of 9 months.InterpretationIn this study, we highlight the importance of recognizing milder forms of the disorder, which may escape detection due to atypical disease presentation.