Utility of whole exome sequencing for the early diagnosis of pediatric-onset cerebellar atrophy associated with developmental delay in an inbred population

International audienceAbstractBackgroundCerebellar atrophy and developmental delay are commonly associated features in large numbers of genetic diseases that frequently also include epilepsy. These defects are highly heterogeneous on both the genetic and clinical levels. Patients with these signs also typically present with non-specific neuroimaging results that can help prioritize further investigation but don’t suggest a specific molecular diagnosis.MethodsTo genetically explore a cohort of 18 Egyptian families with undiagnosed cerebellar atrophy identified on MRI, we sequenced probands and some non-affected family members via high-coverage whole exome sequencing (WES; >97 % of the exome covered at least by 30x). Patients were mostly from consanguineous families, either sporadic or multiplex. We analyzed WES data and filtered variants according to dominant and recessive inheritance models.ResultsWe successfully identified disease-causing mutations in half of the families screened (9/18). These mutations are located in seven different genes, PLA2G6 being the gene most frequently mutated (n = 3). We also identified a recurrent de novo mutation in the KIF1A gene and a molybdenum cofactor deficiency caused by the loss of the start codon in the MOCS2A open-reading frame in a mildly affected subject.ConclusionsThis study illustrates the necessity of screening for dominant mutations in WES data from consanguineous families. Our identification of a patient with a mild and improving phenotype carrying a previously characterized severe loss of function mutation also broadens the clinical spectrum associated with molybdenum cofactor deficiency

Characterization of greater middle eastern genetic variation for enhanced disease gene discovery

The Greater Middle East (GME) has been a central hub of human migration and population admixture. The tradition of consanguinity, variably practiced in the Persian Gulf region, North Africa, and Central Asia1-3, has resulted in an elevated burden of recessive disease4. Here we generated a whole-exome GME variome from 1,111 unrelated subjects. We detected substantial diversity and admixture in continental and subregional populations, corresponding to several ancient founder populations with little evidence of bottlenecks. Measured consanguinity rates were an order of magnitude above those in other sampled populations, and the GME population exhibited an increased burden of runs of homozygosity (ROHs) but showed no evidence for reduced burden of deleterious variation due to classically theorized ‘genetic purging’. Applying this database to unsolved recessive conditions in the GME population reduced the number of potential disease-causing variants by four- to sevenfold. These results show variegated genetic architecture in GME populations and support future human genetic discoveries in Mendelian and population genetics

Wnt/Wg pathway activation in medulloblastoma and disease risk stratification

Despite improvements in the diagnosis and therapy of medulloblastoma (MB), the most common malignant paediatric brain tumour, survival rates are still only 40-60% for high-risk patients, and surviving patients offer suffer life-long therapy-associated late effects. Improving the current clinical disease risk classification may provide opportunities to tailor therapy to disease-risk more accurately, aiming to improve survival whilst minimizing late effects. The overall aim of this study was to identify and validate clinicopathological and molecular markers of prognostic significance in a large cohort of cases derived from the PNET3 clinical trial, which could be used in improved disease stratification schemes. Initial investigations focussed on defects of chromosome 17, the most common chromosomal aberrations identified in MB, which have been associated with adverse outcome in some previous studies. During this initial study, methods for testing for regions of genetic loss in DNA samples extracted from routinely collected and stored pathology material were developed and validated. Using these methods, 17p loss was assessed in 284 cases overall, but did not show an association with poor prognosis. 17p loss was rare in patients 3 years (21/59, p=0.009), supporting the notion that infants with MB may represent a biologically distinct disease subgroup. During the course of this study, evidence of a microsatellite instability phenotype was also found in one case (1/29; 3.8%), suggesting a role for this phenomenon in a small subset of medulloblastomas. Previous studies have identified MBs with activation of the Wnt/Wg signalling pathway to be associated with a favourable outcome. Three independent molecular markers of Wnt/Wg activation in medulloblastoma (β-catenin nuclear localization, CTNNB1 mutation and chromosome 6 loss) have recently been identified, and were therefore assessed in the PNET3 cohort to examine their inter-relationships and relative prognostic utilities. β-catenin nuclear localization was detected in 34/207 (16.4%) cases, CTNNB1 mutation in 20/197 (10.1%), and chromosome 6 LOH in 20/190 (10.5%); all three markers were highly associated with each other (p=0.001). Univariate survival analysis showed significant associations between clinicopathological (gender, pathology, M stage and extent of tumour resection) and molecular variables (β-catenin nuclear localization and CTNNB1 mutation) with survival (all 'p' values <0.03) in the PNET3 cohort. Multivariate survival analysis and Cox proportional regression hazard analysis identified M stage, large cell / anaplastic pathology, gender and β-catenin nuclear staining to be significantly associated with prognosis (p<0.04), with hazard ratios of 1.924, 3.338, 0.57 and 0.215, respectively. Finally, combined models for disease risk prediction were constructed using the prognostically significant clinicopathological and molecular variables identified, including models to precisely classify the overall disease hazard for individual cases based on their clinical, pathological and molecular profile. In summary, Wnt/Wg pathway activation was associated with favourable outcome and can be most accurately identified using β-catenin nuclear staining. Combined models which incorporate Wnt/Wg status alongside other prognostically significant clinical and pathological indices validated in this study provide schemes for improved disease risk prediction in medulloblastoma, which should now be tested prospectively in large clinical trials.EThOS - Electronic Theses Online ServiceEgyptian GovernmentGBUnited Kingdo

Correlation between clinical features and MECP2 gene mutations in patients with Rett syndrome

Background: Rett syndrome is a progressive neurodevelopment disorder which mainly affects females and is a common cause of mental retardation. Loss of purposeful hand movements, regression of acquired cognitive and motor skills and autistic features are the main criteria associated with this disorder. Sixty to ninety percent of the cases show MECP2 gene mutations, which reside on the X chromosome. MECP2 regulates gene expression in a repressive manner. The aim of this study is to estimate the incidence of MECP2 mutations in 32 female Egyptian patients clinically diagnosed with Rett syndrome, and to correlate their clinical features with MECP2 mutation status. Patients: 32 female Egyptian patients with a mean age of 36.9 months diagnosed clinically to suffer from Rett syndrome are the cohort of this study. Methods: Thorough clinical examination, MRI, EEG and testing for MECP2 gene mutations. Results: Twenty of the 32 (62.5%) patients showed MECP2 mutations an incidence which falls within that reported in the literature. Patients with MECP2 gene mutations presented with more severe clinical abnormalities. Conclusions: Mutation screening for MECP2 is a fast and reliable method to diagnose patients clinically suspected to suffer from Rett syndrome or female patients with atypical Rett syndrome features, mental retardation, developmental delay and other neurological abnormalities who do not fit any specific diagnosis. Also, patients with MECP2 mutation presented with a more severe phenotype

Rapid diagnosis of medulloblastoma molecular subgroups

Purpose: Microarray studies indicate medulloblastoma comprises distinct molecular disease subgroups, which offer potential for improved clinical management. Experimental Design: Minimal mRNA expression signatures diagnostic for the Wnt/Wingless (WNT) and Sonic Hedgehog (SHH) subgroups were developed, validated, and used to assign subgroup affiliation in 173 tumors from four independent cohorts, alongside a systematic investigation of subgroup clinical and molecular characteristics. Results: WNT tumors [12% (21/173)] were diagnosed >5 years of age (peak, 10 years), displayed classic histology, CTNNB1 mutation (19/20), and associated chromosome 6 loss, and have previously been associated with favorable prognosis. SHH cases [24% (42/173)] predominated in infants (<3 years) and showed an age-dependent relationship to desmoplastic/nodular pathology; all infant desmoplastic/nodular cases (previously associated with a good outcome) were SHH-positive, but these relationships broke down in noninfants. PTCH1 mutations were common [34% (11/32)], but PTCH1 exon1c hypermethylation, chromosome 9q and REN (KCTD11) genetic loss were not SHH associated, and SMO or SUFU mutation, PTCH1 exon1a or SUFU hypermethylation did not play a role, indicating novel activating mechanisms in the majority of SHH cases. SHH tumors were associated with an absence of COL1A2 methylation. WNT/SHH-independent medulloblastomas [64% (110/173)] showed all histologies, peaked at 3 and 6 years, and were exclusively associated with chromosome 17p loss. Conclusions: Medulloblastoma subgroups are characterized by distinct genomic, epigenomic and clinicopathologic features, and clinical outcomes. Validated array-independent gene expression assays for the rapid assessment of subgroup affiliation in small biopsies provide a basis for their routine clinical application, in strategies including molecular disease-risk stratification and delivery of targeted therapeutics

MYC family amplification and clinical risk-factors interact to predict an extremely poor prognosis in childhood medulloblastoma

The MYC oncogenes are the most commonly amplified loci in medulloblastoma, and have previously been proposed as biomarkers of adverse disease prognosis by us and others. Here, we report focussed and comprehensive investigations of MYCC, MYCN and MYCL in an extensive medulloblastoma cohort (n = 292), aimed to define more precisely their biological significance and optimal clinical application to direct improved disease risk-stratification and individualisation of therapy. MYCC and MYCN expression elevations were multifactorial, associated with high-risk (gene amplification, large-cell/anaplastic pathology (LCA)) and favourable-risk (WNT/SHH molecular subgroups) disease features. Highly variable cellular gene amplification patterns underlay overall MYC copy number elevations observed in tumour biopsies; we used these alternative measures together to define quantitative methodologies and thresholds for amplification detection in routinely collected tumour material. MYCC and MYCN amplification, but not gain, each had independent prognostic significance in non-infants (≥3.0–16.0 years), but MYCC conferred a greater hazard to survival than MYCN when considered across this treatment group. MYCN’s weaker group-wide survival relationship may be explained by its pleiotropic behaviour between clinical disease-risk groups; MYCN predicted poor prognosis in clinical high-risk (metastatic (M+) or LCA), but not standard-risk, patients. Extending these findings, survival decreased in proportion to the total number of independently significant high-risk features present (LCA, M+ or MYCC/MYCN amplification). This cumulative-risk model defines a patient group characterised by ≥2 independent risk-factors and an extremely poor prognosis (<15% survival), which can be identified straightforwardly using the reported MYC amplification detection methodologies alongside clinical assessments, enabling targeting for novel/intensified therapies in future clinical studies

Clinical and genetic characterization of ten Egyptian patients with Wolf–Hirschhorn syndrome and review of literature

Abstract Background Wolf–Hirschhorn syndrome (WHS) (OMIM 194190) is a multiple congenital anomalies/intellectual disability syndrome. It is caused by partial loss of genetic material from the distal portion of the short arm of chromosome. Methods We studied the phenotype–genotype correlation. Results We present the clinical manifestations and cytogenetic results of 10 unrelated Egyptian patients with 4p deletions. Karyotyping, FISH and MLPA was performed for screening for microdeletion syndromes. Array CGH was done for two patients. All patients exhibited the cardinal clinical manifestation of WHS. FISH proved deletion of the specific WHS locus in all patients. MLPA detected microdeletion of the specific locus in two patients with normal karyotypes, while array CGH, performed for two patients, has delineated the extent of the deleted segments and the involved genes. LETM1, the main candidate gene for the seizure phenotype, was found deleted in the two patients tested by array CGH; nevertheless, one of them did not manifest seizures. The study emphasized the previous. Conclusion WHS is a contiguous gene syndrome resulting from hemizygosity of the terminal 2 Mb of 4p16.3 region. The Branchial fistula, detected in one of our patients is a new finding that, to our knowledge, was not reported