Showing papers by "Soma Das published in 2011"

The core FOXG1 syndrome phenotype consists of postnatal microcephaly, severe mental retardation, absent language, dyskinesia, and corpus callosum hypogenesis

Abstract: Background Submicroscopic deletions in 14q12 spanning FOXG1 or intragenic mutations have been reported in patients with a developmental disorder described as a congenital variant of Rett syndrome. This study aimed to further characterise and delineate the phenotype of FOXG1 mutation positive patients. Method The study mapped the breakpoints of a 2;14 translocation by fluorescence in situ hybridisation and analysed three chromosome rearrangements in 14q12 by cytogenetic analysis and/or array comparative genomic hybridisation. The FOXG1 gene was sequenced in 210 patients, including 129 patients with unexplained developmental disorders and 81 MECP2 mutation negative individuals. Results One known mutation, seen in two patients, and nine novel mutations of FOXG1 including two deletions, two chromosome rearrangements disrupting or displacing putative cis -regulatory elements from FOXG1 , and seven sequence changes, are reported. Analysis of 11 patients in this study, and a further 15 patients reported in the literature, demonstrates a complex constellation of features including mild postnatal growth deficiency, severe postnatal microcephaly, severe mental retardation with absent language development, deficient social reciprocity resembling autism, combined stereotypies and frank dyskinesias, epilepsy, poor sleep patterns, irritability in infancy, unexplained episodes of crying, recurrent aspiration, and gastro-oesophageal reflux. Brain imaging studies reveal simplified gyral pattern and reduced white matter volume in the frontal lobes, corpus callosum hypogenesis, and variable mild frontal pachgyria. Conclusions These findings have significantly expanded the number of FOXG1 mutations and identified two affecting possible cis -regulatory elements. While the phenotype of the patients overlaps both classic and congenital Rett syndrome, extensive clinical evaluation demonstrates a distinctive and clinically recognisable phenotype which the authors suggest designating as the FOXG1 syndrome.

Phenotypic spectrum associated with CASK loss-of-function mutations

Abstract: Background Heterozygous mutations in the CASK gene in Xp11.4 have been shown to be associated with a distinct brain malformation phenotype in females, including disproportionate pontine and cerebellar hypoplasia. Methods The study characterised the CASK alteration in 20 new female patients by molecular karyotyping, fluorescence in situ hybridisation, sequencing, reverse transcriptase (RT) and/or quantitative real-time PCR. Clinical and brain imaging data of a total of 25 patients were reviewed. Results 11 submicroscopic copy number alterations, including nine deletions of ∼11 kb to 4.5 Mb and two duplications, all covering (part of) CASK , four splice, four nonsense, and one 1 bp deletion are reported. These heterozygous CASK mutations most likely lead to a null allele. Brain imaging consistently showed diffuse brainstem and cerebellar hypoplasia with a dilated fourth ventricle, but of remarkably varying degrees. Analysis of 20 patients in this study, and five previously reported patients, revealed a core clinical phenotype comprising severe developmental delay/intellectual disability, severe postnatal microcephaly, often associated with growth retardation, (axial) hypotonia with or without hypertonia of extremities, optic nerve hypoplasia, and/or other eye abnormalities. A recognisable facial phenotype emerged, including prominent and broad nasal bridge and tip, small or short nose, long philtrum, small chin, and/or large ears. Conclusions These findings define the phenotypic spectrum associated with CASK loss-of-function mutations. The combination of developmental and brain imaging features together with mild facial dysmorphism is highly suggestive of this disorder and should prompt subsequent testing of the CASK gene.

Exome sequencing reveals a novel mutation for autosomal recessive non-syndromic mental retardation in the TECR gene on chromosome 19p13

Abstract: Exome sequencing is a powerful tool for discovery of the Mendelian disease genes. Previously, we reported a novel locus for autosomal recessive non-syndromic mental retardation (NSMR) in a consanguineous family [Nolan, D.K., Chen, P., Das, S., Ober, C. and Waggoner, D. (2008) Fine mapping of a locus for nonsyndromic mental retardation on chromosome 19p13. Am. J. Med. Genet. A, 146A, 1414-1422]. Using linkage and homozygosity mapping, we previously localized the gene to chromosome 19p13. The parents of this sibship were recently included in an exome sequencing project. Using a series of filters, we narrowed the putative causal mutation to a single variant site that segregated with NSMR: the mutation was homozygous in five affected siblings but in none of eight unaffected siblings. This mutation causes a substitution of a leucine for a highly conserved proline at amino acid 182 in TECR (trans-2,3-enoyl-CoA reductase), a synaptic glycoprotein. Our results reveal the value of massively parallel sequencing for identification of novel disease genes that could not be found using traditional approaches and identifies only the seventh causal mutation for autosomal recessive NSMR.

Exome Sequencing and the Genetics of Intellectual Disability

Abstract: Exome sequencing has greatly impacted the speed at which new disease genes are identified. In the last year alone, six studies have used exome sequencing to identify new genes involved in intellectual disability, a genetically heterogeneous condition affecting 1-3% of the population. These studies encompass the full gamut of modes of inheritance and phenotypic presentation, including syndromic and non-syndromic conditions, sporadic and familial cases, and dominant and recessive inheritance patterns. Because different disease presentations require different approaches to gene discovery, studies of intellectual disability provide a nearly comprehensive showcase of strategies for exome-driven gene discovery. Despite these successes, the etiology of ~60% of cases of intellectual disability remains unknown. The application of exome sequencing to the clinical diagnosis of intellectual disability in the near future will ultimately reduce the number of idiopathic cases and provide a rich source of sequence variation for the identification of new intellectual disability genes.

Novel Functional Germline Variants in the VEGF Receptor 2 Gene and Their Effect on Gene Expression and Microvessel Density in Lung Cancer

Abstract: Purpose: VEGFR-2 plays a crucial role in mediating angiogenic endothelial cell responses via the VEGF pathway and angiogenesis inhibitors targeting VEGFR-2 are in clinical use. As angiogenesis is a host-driven process, functional heritable variation in KDR , the gene encoding VEGFR-2, may affect VEGFR-2 function, and ultimately, the extent of tumor angiogenesis. Experimental Design: We resequenced KDR using 24 DNAs each from healthy Caucasian, African American and Asian groups. Non-synonymous genetic variants were assessed for function using phosphorylation assays. Luciferase reporter gene assays were used to examine effects of variants on gene expression. KDR mRNA and protein expression, and microvessel density (MVD) were measured in non-small cell lung cancer (NSCLC) tumor samples and matching patient DNA samples were genotyped to test for associations with variants of interest. Results: KDR resequencing led to the discovery of 120 genetic variants, of which 25 had not been previously reported. Q472H had increased VEGFR-2 protein phosphorylation and associated with increased MVD in NSCLC tumor samples. -2854C and -2455A increased luciferase expression and associated with higher KDR mRNA levels in NSCLC samples. -271A reduced luciferase expression and associated with lower VEGFR-2 levels in NSCLC samples. -906C and 23408G, associated with higher KDR mRNA levels in NSCLC samples. Conclusions: This study has defined KDR genetic variation in three populations and identified common variants that impact on tumoral KDR expression and vascularization. These findings may have important implications for understanding the molecular basis of genetic associations between KDR variation and clinical phenotypes related to VEGFR-2 function.

Platinum sensitivity-related germline polymorphism discovered via a cell-based approach and analysis of its association with outcome in ovarian cancer patients

Abstract: Purpose: Cell-based approaches were used to identify genetic markers predictive of patients9 risk for poor response prior to chemotherapy. Experimental Design: We conducted genome-wide association studies (GWAS) to identify single-nucleotide polymorphisms (SNP) associated with cellular sensitivity to carboplatin through their effects on mRNA expression using International HapMap lymphoblastoid cell lines (LCL) and replicated them in additional LCLs. SNPs passing both stages of the cell-based study were tested for association with progression-free survival (PFS) in patients. Phase 1 validation was based on 377 ovarian cancer patients receiving at least four cycles of carboplatin and paclitaxel from the Australian Ovarian Cancer Study (AOCS). Positive associations were then assessed in phase 2 validation analysis of 1,326 patients from the Ovarian Cancer Association Consortium and The Cancer Genome Atlas. Results: In the initial GWAS, 342 SNPs were associated with carboplatin-induced cytotoxicity, of which 18 unique SNPs were retained after assessing their association with gene expression. One SNP (rs1649942) was replicated in an independent LCL set (Bonferroni adjusted P P per-allele = 2 × 10 −2 ), with a stronger effect in the subset of women with optimally debulked tumors ( P per-allele = 4 × 10 −3 ). rs1649942 was also associated with poorer overall survival in women with optimally debulked tumors ( P per-allele = 9 × 10 −3 ). However, this SNP was not significant in phase 2 validation analysis with patients from numerous cohorts. Conclusion: This study shows the potential of cell-based, genome-wide approaches to identify germline predictors of treatment outcome and highlights the need for extensive validation in patients to assess their clinical effect. Clin Cancer Res; 17(16); 5490–500. ©2011 AACR .

Abstract 5136: Discovery of novel functional germline variations in the vascular endothelial growth factor receptor 2 gene (KDR) and their effect on gene expression and microvessel density in lung cancer

Abstract: Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Background: VEGFR-2 plays a crucial role in mediating angiogenic endothelial cell responses via the VEGF pathway and VEGFR-2 inhibitors are in clinical use. Angiogenesis is a host-driven process and heritable variation in KDR, the gene encoding VEGFR-2, may impact on angiogenesis. We have previously characterized KDR germline genetic variation by resequencing (Ye et al., AACR, 2007;737). However, very limited data are available on the molecular function of KDR germline variants and their resulting effect on tumoral expression and vascularization. The purpose of this study was to identify functional variants affecting KDR expression and microvessel density (MVD) in lung cancer. Methods: The molecular function of the non-synonymous variants R106W, V297I, Q472H and C482R was assessed by testing their effect on VEGFR-2 phosphorylation (measured via immunoblotting) after VEGFA165 exposure in HEK293 cells. Four 5’ upstream region SNPs were examined by luciferase reporter assay in endothelial SVEC4-10 cells. To validate these results and examine the effects of other KDR SNPs, KDR mRNA and VEGFR-2 protein expression were measured by quantitative PCR and immunohistochemistry in non-small cell lung cancer (NSCLC) samples from Caucasian patients and MVD was assessed by immunohistochemistry (anti-CD31). Patients were genotyped for KDR SNPs and their association with mRNA, protein expression, or MVD was evaluated by linear regression. Results: Among the non-synonymous SNPs, the only significant observation was that Q472H had a 46% increase in VEGFR-2 phosphorylation relative to the wild-type variant after VEGFA165 stimulation (p=0.035); in agreement with this finding, Q472H was associated with 22% greater MVD in 168 NSCLC patients (p=0.05). Of the putatively regulatory SNPs, -2008G and -1942G had no effect on luciferase expression. -2854C and -2455A showed 10-20% higher luciferase expression than the wild-type variant (p<0.05) and they were associated with 23% greater tumor KDR mRNA expression in 66 NSCLC patients (p=0.014), but had no effect on tumor VEFGR-2 expression; -271A reduced luciferase expression by approximately 50% (Ye et al., AACR 2007;737) and correlated with 10% lower tumor VEGFR-2 expression in 162 NSCLC patients (p=0.01); -906C and 23408G were associated with 20-25% higher tumor KDR mRNA levels (p=0.009 and 0.025, respectively), although these variants had no effect on tumor VEGFR-2 expression. Conclusions: This study has identified novel functional variants which correlate with in vitro gene expression or VEGFR-2 phosphorylation and KDR expression or MVD in NSCLC. This information may be critical for understanding the role of heritable KDR variations as contributing factors to prognosis and lung cancer outcome. Acknowledgment: This research was supported by the American Cancer Society. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5136. doi:10.1158/1538-7445.AM2011-5136