| 11. |
- Moens, Lotte N. J., et al.
(author)
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HaloPlex Targeted Resequencing for Mutation Detection in Clinical Formalin-Fixed, Paraffin-Embedded Tumor Samples
- 2015
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In: Journal of Molecular Diagnostics. - : Elsevier BV. - 1525-1578 .- 1943-7811. ; 17:6, s. 729-739
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Journal article (peer-reviewed)abstract
- In recent years, the advent of massively parallel next-generation sequencing technologies has enabled substantial advances in the study of human diseases. Combined with targeted DNA enrichment methods, high sequence coverage can be obtained for different genes simultaneously at a reduced cost per sample, creating unique opportunities for clinical cancer diagnostics. However, the formalin-fixed, paraffin-embedded (FFPE) process of tissue samples, routinely used in pathology departments, results in DNA fragmentation and nucleotide modifications that introduce a number of technical challenges for downstream biomotecular analyses. We evaluated the HaloPlex target enrichment system for somatic mutation detection in 80 tissue fractions derived from 20 clinical cancer cases with paired tumor and normal tissue available in both FFPE and fresh-frozen format. Several modifications to the standard method were introduced, including a reduced target fragment Length and two strand capturing. We found that FFPE material can be used for HaloPlex-based target enrichment and next-generation sequencing, even when starting from small amounts of DNA. By specifically capturing both strands for each target fragment, we were able to reduce the number of false-positive errors caused by FFPE-induced artifacts and Lower the detection limit for somatic mutations. We believe that the HaloPlex method presented here will be broadly applicable as a tool for somatic mutation detection in clinical cancer settings.
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| 12. |
- Moens, Lotte N, et al.
(author)
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PCM1 and schizophrenia : a replication study in the Northern Swedish population
- 2010
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In: American Journal of Medical Genetics. - : Wiley. - 0148-7299 .- 1096-8628. ; 153B:6, s. 1240-1243
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Journal article (peer-reviewed)abstract
- Previous studies implicated centrosomal dysfunction as a source of various neuropsychiatric disorders, including schizophrenia (SZ). Two recent reports [Gurling et al., 2006; Datta et al., 2008. Mol Psychiatry] described an association between polymorphisms in the PCM1 gene and SZ in a UK/Scottish population. In this study, we aimed to replicate these findings in a Northern Swedish association sample of 486 research subjects with SZ and 512 unrelated control individuals. We genotyped 12 previously described SNP markers and carried out haplotype analyses using the same multi-marker haplotypes previously reported. Though we could not replicate the association with SNPs rs445422 and rs208747, we did observe a significant protective association with intronic SNP rs13276297. Furthermore, we performed a meta-analysis comprising 1,794 SZ patients and 1,553 controls, which confirmed the previously reported association with rs445422 and rs208747. These data provide further evidence that PCM1-though certainly not a major risk factor in the Northern Swedish population-cannot be ruled out as a contributor to SZ risk and/or protection, and deserves further replication in larger populations to elucidate its role in disease etiology.
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| 13. |
- Moens, Lotte N, et al.
(author)
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Sequencing of DISC1 Pathway Genes Reveals Increased Burden of Rare Missense Variants in Schizophrenia Patients from a Northern Swedish Population
- 2011
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In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:8, s. e23450-
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Journal article (peer-reviewed)abstract
- In recent years, DISC1 has emerged as one of the most credible and best supported candidate genes for schizophrenia and related neuropsychiatric disorders. Furthermore, increasing evidence - both genetic and functional - indicates that many of its protein interaction partners are also involved in the development of these diseases. In this study, we applied a pooled sample 454 sequencing strategy, to explore the contribution of genetic variation in DISC1 and 10 of its interaction partners (ATF5, Grb2, FEZ1, LIS-1, PDE4B, NDE1, NDEL1, TRAF3IP1, YWHAE, and ZNF365) to schizophrenia susceptibility in an isolated northern Swedish population. Mutation burden analysis of the identified variants in a population of 486 SZ patients and 514 control individuals, revealed that non-synonymous rare variants with a MAF<0.01 were significantly more present in patients compared to controls (8.64% versus 4.7%, P = 0.018), providing further evidence for the involvement of DISC1 and some of its interaction partners in psychiatric disorders. This increased burden of rare missense variants was even more striking in a subgroup of early onset patients (12.9% versus 4.7%, P = 0.0004), highlighting the importance of studying subgroups of patients and identifying endophenotypes. Upon investigation of the potential functional effects associated with the identified missense variants, we found that similar to 90% of these variants reside in intrinsically disordered protein regions. The observed increase in mutation burden in patients provides further support for the role of the DISC1 pathway in schizophrenia. Furthermore, this study presents the first evidence supporting the involvement of mutations within intrinsically disordered protein regions in the pathogenesis of psychiatric disorders. As many important biological functions depend directly on the disordered state, alteration of this disorder in key pathways may represent an intriguing new disease mechanism for schizophrenia and related neuropsychiatric diseases. Further research into this unexplored domain will be required to elucidate the role of the identified variants in schizophrenia etiology.
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| 14. |
- Sassi, Atfa, et al.
(author)
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Hypomorphic homozygous mutations in phosphoglucomutase 3 (PGM3) impair immunity and increase serum IgE levels
- 2014
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In: Journal of Allergy and Clinical Immunology. - : Elsevier BV. - 0091-6749 .- 1097-6825. ; 133:5, s. 1410-U681
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Journal article (peer-reviewed)abstract
- Background: Recurrent bacterial and fungal infections, eczema, and increased serum IgE levels characterize patients with the hyper-IgE syndrome (HIES). Known genetic causes for HIES are mutations in signal transducer and activator of transcription 3 (STAT3) and dedicator of cytokinesis 8 (DOCK8), which are involved in signal transduction pathways. However, glycosylation defects have not been described in patients with HIES. One crucial enzyme in the glycosylation pathway is phosphoglucomutase 3 (PGM3), which catalyzes a key step in the synthesis of uridine diphosphate N-acetylglucosamine, which is required for the biosynthesis of N-glycans. Objective: We sought to elucidate the genetic cause in patients with HIES who do not carry mutations in STAT3 or DOCK8. Methods: After establishing a linkage interval by means of SNPchip genotyping and homozygosity mapping in 2 families with HIES from Tunisia, mutational analysis was performed with selector-based, high-throughput sequencing. Protein expression was analyzed by means of Western blotting, and glycosylation was profiled by using mass spectrometry. Results: Mutational analysis of candidate genes in an 11.9-Mb linkage region on chromosome 6 shared by 2 multiplex families identified 2 homozygous mutations in PGM3 that segregated with disease status and followed recessive inheritance. The mutations predict amino acid changes in PGM3 (p. Glu340del and p. Leu83Ser). A third homozygous mutation (p. Asp502Tyr) and the p. Leu83Ser variant were identified in 2 other affected families, respectively. These hypomorphic mutations have an effect on the biosynthetic reactions involving uridine diphosphate N-acetylglucosamine. Glycomic analysis revealed an aberrant glycosylation pattern in leukocytes demonstrated by a reduced level of tri-antennary and tetra-antennary N-glycans. T-cell proliferation and differentiation were impaired in patients. Most patients had developmental delay, and many had psychomotor retardation. Conclusion: Impairment of PGM3 function leads to a novel primary (inborn) error of development and immunity because biallelic hypomorphic mutations are associated with impaired glycosylation and a hyper-IgE-like phenotype.
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