Bakker JA, Lindhout M, Habets DD, van den Wijngaard A, Paulussen AD,
Bierau J. The Effect of ITPA Polymorphisms on the Enzyme Kinetic
Properties of Human Erythrocyte Inosine Triphosphatase Toward its Substrates
ITP and 6-Thio-ITP. Nucleosides Nucleotides Nucleic Acids. 2011
Nov;30(11):839-49. IF: 1.132
Bonneux S, Fransen E, Van
Eyken E, Van Laer L, Huyghe J, Van de Heyning P, Voets A, Gerards M, Stassen
AP, Hendrickx AT, Smeets HJ, Van Camp G. Inherited mitochondrial
variants are not a major cause of age-related hearing impairment in the European
population. Mitochondrion. 2011;Sep;11(5):729-34. IF: 4.145
CARIM + GROW(ontwikkelingsbiologie)
Borst B van den, Souren
NY, Gielen M, Loos RJ, Paulussen AD, Derom C,
Schols AM, Zeegers MP. Association between the IL6 -174G/C SNP and maximally
attained lung function. Thorax. 2011 Feb;66(2):179. IF: 7.041
RD, van Roozendaal K, Tserpelis D, García
EG, Blok MJ. Characterisation of unclassified variants in the BRCA1/2
genes with a putative effect on splicing. Breast Cancer Res Treat. 2011Oct;129(3):971-82. IF: 4.859
Christiaans I, Nannenberg
EA, Dooijes D, Jongbloed RJ, Michels M, Postema PG, Majoor-Krakauer D,
van den Wijngaard A, Mannens MM, van Tintelen JP, van Langen IM, Wilde AA.
Founder mutations in hypertrophic cardiomyopathy patients in the Netherlands.
Neth Heart J. 2010 May;18(5):248-54. IF: 1.392
Christiaans I, Birnie E,
Bonsel GJ, Mannens MM, Michels M, Majoor-Krakauer D, Dooijes D, van Tintelen JP,
van den Berg MP, Volders PG, Arens YH, van den Wijngaard A, Atsma DE,
Helderman-van den Enden AT, Houweling AC, de Boer K, van der Smagt JJ, Hauer RN,
Marcelis CL, Timmermans J, van Langen IM, Wilde AA. Manifest disease, risk
factors for sudden cardiac death, and cardiac events in a large nationwide
cohort of predictively tested hypertrophic cardiomyopathy mutation carriers:
determining the best cardiological screening strategy. Eur Heart J. 2011
May;32(9):1161-70. IF: 9.800
Cox MG, van der Zwaag PA,
van der Werf C, van der Smagt JJ, Noorman M, Bhuiyan ZA, Wiesfeld AC, Volders
PG, van Langen IM, Atsma DE, Dooijes D, van den Wijngaard A,
Houweling AC, Jongbloed JD, Jordaens L, Cramer MJ, Doevendans PA, de Bakker JM,
Wilde AA, van Tintelen JP, Hauer RN. Arrhythmogenic right ventricular
dysplasia/cardiomyopathy: pathogenic desmosome mutations in index-patients
predict outcome of family screening: Dutch arrhythmogenic right ventricular
dysplasia/cardiomyopathy genotype-phenotype follow-up study. Circulation. 2011
Jun 14;123(23):2690-700. IF: 14.432
Gerards M, van den Bosch BJ, Danhauser K, Serre V, van Weeghel M, Wanders RJ, Nicolaes GA, Sluiter W, Schoonderwoerd K, Scholte HR, Prokisch H, Rötig A, de Coo IF, Smeets HJ. Riboflavin-responsive oxidative phosphorylation complex I deficiency caused by defective ACAD9: new function for an old gene. Brain. 2011 Jan;134(Pt 1):210-9. IF: 9.490
Theeuwes M, van Dekken H,Sikkema M,Steyerberg EW,Lingsma HF,Siersema PD,Xia B,
Kusters JG,van der Woude CJ,Kuipers EJ. Biomarker-based prediction of inflammatory bowel
disease-related colorectal cancer: a case-control study.
Cell Oncol (Dordr). 2011 Apr;34(2):107-17. IF: 3.175
Hofman N, Jongbloed R,
Postema PG, Nannenberg E, Alders M, Wilde AA. Recurrent and Founder
Mutations in the Netherlands: the Long-QT Syndrome. Neth Heart J. 2011
Jan;19(1):10-16. IF: 1.392
Marion V, Sankaranarayanan S, de Theije C, van Dijk P, Lindsey P, Lamers MC, Harding HP, Ron D, Lamers WH, Köhler SE. Arginine deficiency causes runting in the suckling period by selectively activating the stress kinase GCN2. J Biol Chem. 2011 Mar 18;286(11):8866-74.
Paulussen AD, Stegmann AP, Blok MJ, Tserpelis D, Posma-Velter C,
Detisch Y, Smeets EE,
Wagemans A, Schrander JJ, van den Boogaard MJ, van der Smagt J, van
Haeringen A, Stolte-Dijkstra I, Kerstjens-Frederikse WS, Mancini GM, Wessels MW,
Hennekam RC, Vreeburg M, Geraedts J, de Ravel T, Fryns JP, Smeets HJ,
Devriendt K, Schrander-Stumpel CT. MLL2 mutation spectrum in 45
patients with Kabuki syndrome. Hum Mutat. 2011 Feb;32(2):E2018-25. IF: 6.887
Pieters S, Van Der
Vorst H, Burk WJ, Schoenmakers TM, Van Den Wildenberg E,
Lambrichs E, Field M, Engels RC, Wiers RW.
The effect of the OPRM1 and
DRD4 polymorphisms on the relation between attentional bias and alcohol use in adolescence and young adulthood. Dev Cogn Neurosci. 2011 Oct;1(4):591-9. PubMed PMID: 22436571.
Stassen, A.P.M., Ruivenkamp, C.A.L.,
Van Wezel, T., Fijneman, R.J.A., Hutson, A., Kakarlapudi, N., Hart, A.A.M. &
Demant, P. Most Lung and Colon Cancer Susceptibility Genes Are Pair-Wise Linked
in Mice, Humans and Rats. PloS ONE
2011 Feb;6(2), e14727. IF:
Ramus SJ, Kartsonaki C, Gayther SA, Pharoah PD, Sinilnikova OM, Beesley
J, Chen X, McGuffog L, Healey S, Couch FJ, Wang X, Fredericksen Z, Peterlongo P,
Manoukian S, Peissel B, Zaffaroni D, Roversi G, Barile M, Viel A, Allavena A,
Ottini L, Papi L, Gismondi V, Capra F, Radice P, Greene MH, Mai PL, Andrulis IL,
Glendon G, Ozcelik H; OCGN, Thomassen M, Gerdes AM, Kruse TA, Cruger D, Jensen
UB, Caligo MA, Olsson H, Kristoffersson U, Lindblom A, Arver B, Karlsson P,
Stenmark Askmalm M, Borg A, Neuhausen SL, Ding YC, Nathanson KL, Domchek SM,
Jakubowska A, Lubinski J, Huzarski T, Byrski T, Gronwald J, Górski B, Cybulski
C, Debniak T, Osorio A, Durán M, Tejada MI, Benítez J, Hamann U, Rookus MA,
Verhoef S, Tilanus-Linthorst MA, Vreeswijk MP, Bodmer D, Ausems MG, van Os TA,
Asperen CJ, Blok MJ, Meijers-Heijboer HE; HEBON; EMBRACE, Peock S, Cook
M, Oliver C, Frost D, Dunning AM, Evans DG, Eeles R, Pichert G, Cole T, Hodgson
S, Brewer C, Morrison PJ, Porteous M, Kennedy MJ, Rogers MT, Side LE, Donaldson
A, Gregory H, Godwin A, Stoppa-Lyonnet D, Moncoutier V, Castera L, Mazoyer S,
Barjhoux L, Bonadona V, Leroux D, Faivre L, Lidereau R, Nogues C, Bignon YJ,
Prieur F, Collonge-Rame MA, Venat-Bouvet L, Fert-Ferrer S; GEMO Study
Collaborators, Miron A, Buys SS, Hopper JL, Daly MB, John EM, Terry MB, Goldgar
D; BCFR, Hansen TV, Jřnson L, Ejlertsen B, Agnarsson BA, Offit K, Kirchhoff T,
Vijai J, Dutra-Clarke AV, Przybylo JA, Montagna M, Casella C, Imyanitov EN,
Janavicius R, Blanco I, Lázaro C, Moysich KB, Karlan BY, Gross J, Beattie MS,
Schmutzler R, Wappenschmidt B, Meindl A, Ruehl I, Fiebig B, Sutter C, Arnold N,
Deissler H, Varon-Mateeva R, Kast K, Niederacher D, Gadzicki D, Caldes T, de la
Hoya M, Nevanlinna H, Aittomäki K, Simard J, Soucy P; kConFab Investigators,
Spurdle AB, Holland H, Chenevix-Trench G, Easton DF, Antoniou AC; on behalf of
Consortium of Investigators of Modifiers of BRCA1/2. 2010. Genetic Variation at
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Schruers K. Esquivel G,
van Duinen M, Wichers M, Kenis G, Colasanti A, Knuts I, Goossens L, Jacobs N,
van Rozendaal J, Smeets H, van Os J, Griez E.
Genetic moderation of CO2-induced fear by 5-HTTLPR
genotype. J Psychopharmacol. 2011
Jan;25(1):37-42. IF: 3.801
Ŝíma M, Havelková
H, Quan L, Svobodová
J, Stassen A, Demant P, Lipoldova M. Genetic
Control of Resistance to Trypanosoma brucei
brucei Infection in Mice. PLoS Negl Trop Dis
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Stalpers X, Verrips A,
Braakhekke J, Lammens M, van den Wijngaard A, Mostert A. Scoliosis
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CARIM en GROW (ontwikkelingsbiologie)
Veeck, R.D. Brandao, J.C. van Elssen, K.K. van de Vijver, P.J. Lindsey,
M.J. Blok, K. Keymeulen, T. Ayoubi, H.J. Smeets, P.S. Hupperets.
A Randomised Trial Of Pre-Operative Celecoxib
Treatment Reveals Anti-Tumour Transcriptional Response In Primary Breast Cancer.
Annals of Oncology, 2011; 22: Supplement 2
Voets AM, van den Bosch BJ, Stassen AP, Hendrickx AT, Hellebrekers DM, Van
Laer L, Van Eyken E, Van Camp G, Pyle A, Baudouin SV, Chinnery PF, Smeets HJ.
Large scale mtDNA sequencing reveals sequence and functional conservation as
major determinants of homoplasmic mtDNA variant distribution. Mitochondrion.
CARIM + GROW(ontwikkelingsbiologie)
WH de, Houben F, Kamps M, Malhas A, Verheyen F, Cox J, Manders EM,
Verstraeten VL, van Steensel MA, Marcelis CL, van den Wijngaard A, Vaux
DJ, Ramaekers FC, Broers JL. Repetitive disruptions of the nuclear
envelope invoke temporary loss of cellular compartmentalization in
laminopathies. Hum Mol Genet. 2011 Nov 1;20(21):4175-86. IF: 8.058
GROW (oncologie) + CARIM
Wijngaard A van den, Volders P, Van Tintelen JP, Jongbloed JD, van den Berg
MP, Lekanne Deprez RH, Mannens MM, Hofmann N, Slegtenhorst M, Dooijes D, Michels
M, Arens Y, Jongbloed R, Smeets BJ. Recurrent and founder
mutations in the Netherlands: cardiac Troponin I (TNNI3) gene mutations as a
cause of severe forms of hypertrophic and restrictive cardiomyopathy. Neth Heart
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Eds. H.F. Baars, J.J. van der Smagt, P.A. Doevendans
Springer-Verlag London Ltd 2011
Introduction to Molecular Genetics: M.M.A.M. Mannens and H.J.M. Smeets
Florence H.J. van Tienen. Deciphering mitochondrial and molecular
pathology in Type 2 Diabetes Mellitus. 4 februari 2011
Promotor: prof.dr. H.J.M. Smeets
Ruben Martherus. Pulsing response of the cardiac transcriptome. 7
Promotores: prof.dr. H.J.M. Smeets en Prof.dr. J.P.M. Geraedts
Pre-implantation genetic diagnosis offers a fair chance of having
D. Hellebrekers1, R. Wolfe2, A. Hendrickx1, R. de Coo3, C. de Die1, J. Geraedts1, P. Chinnery4, H. Smeets1;1Maastricht University Medical Center, Maastricht, Netherlands, 2Schoolof Public Health and Preventive Medicine, Monash University, Melbourne,Australia, 3Department of Pediatric Neurology, Erasmus MC-Sophia Children’sHospital, Rotterdam, Netherlands, 4Institute of Human Genetics, NewcastleUniversity, Newcastle, United Kingdom. Oxidative phosphorylation disorders due to maternally inheritedhomoplasmic or heteroplasmic mtDNA mutations affect approximately1/5,000 individuals. The mutation level of transmitted heteroplasmic mtDNA point mutations varies significantly between embryos due to a segregational bottleneck. Prenatal (PND) or pre-implantation genetic diagnosis (PGD) of mtDNA disorders is complicated by the inability to accurately predict the threshold of clinical expression for most heteroplasmic mtDNA mutations. Here we tried to define a minimal mutant level below which the chance for an embryo of being affected is acceptably low, irrespective of the exact mtDNA point mutation. A systematic review was performed on muscle mutant levels of 159 different heteroplasmic mtDNA point mutations derived from 327 unrelated patients or pedigrees, excluding three overrepresented mtDNA mutations. We generated a distribution of mutation levels of all affected individuals (n=195) and their unaffected maternal relatives (n=19) from 137 pedigrees with a familial mtDNA mutation, and predicted the risk of being affected given a varying mutant level. This prediction required an assumption of overall prevalence of affected status in familial pedigrees. For familial mutations, little difference in mean muscle mutant level was observed between probands and affected maternal relatives and between affected individuals with a tRNA-versus protein-coding mutation. The overall prevalence of affected status in familial pedigrees was estimated as 0.477. A 95% or higher chance of being unaffected was associated with a muscle mutant level of 18% or less. PGD provides carriers of heteroplasmic mtDNA point mutations the opportunity to substantially increase the chance of having healthy offspring.
A HPE-like phenotype and heterotaxy associated with a 1,3 Mb deletion
encompassing the GLI2 gene at 2q14.2
S. H. G. Kevelam1, J. J. T. van Harssel1, B. van der Zwaag1, H. J. Smeets2, A. D. C. Paulussen2, K. D. Lichtenbelt1; 1Department of Medical Genetics, University Medical Center Utrecht, Utrecht, Netherlands, 2Department of Clinical Genetics, Maastricht University Medical Center, Maastricht, Netherlands. Loss-of-function mutations of GLI2 are associated with holoprosencephaly (HPE)-features, including abnormal pituitary gland formation and/or function, craniofacial abnormalities, branchial arch anomalies and polydactyly. We report a patient carrying a 1,3 Mb submicroscopic deletion in 2q14.2, encompassing the GLI2 gene. She presented with a HPE-like phenotype (bilateral cleft lip and palate and abnormal pituitary gland formation with panhypopituitarism),heterotaxy and normal psychomotor development. Submicroscopic deletions encompassing GLI2 have not been reported so far. Large microscopically visible interstitial deletions spanning 2q14.2 are rare and have been reported sporadically in patients with multiple congenital anomalies and mental retardation. The clinical and molecular findings in our proband and her family are presented. Additionally, we review the features of previously reported index patients with a GLI2 aberration (17 mutations and 7 large deletions spanning 2q14.2). With the report of this GLI2 deletion we confirm that haploinsufficiency of GLI2 is associated with HPE-like features. Comparable to what has been described in families with GLI2 mutations, we observed an incomplete penetrance of the deletion in our family, illustrating the multifactorial etiology of HPE and HPE-like features. The deletion contains five other genes, including the Epb4.1l5 gene. Based on observations in Epb4.1l5 mutant mice embryo’s, we argue that haploinsufficiency of Epb4.1l5 might be the cause of the heterotaxy in our proband. A pathogenic effect of the GLI2 deletion cannot be excluded. Finally, we conclude that, while other HPE-genes have been excluded, GLI2 is still a candidate gene for the Holoprosencephaly-polydactyly syndrome, also known as Pseudotrisomy 13.
Extended genotype - phenotype correlation study in Kabuki
C. Schrander-Stumpel1, M. Blok2, D. Tserpelis1, C. Posma-Velter1, C. Rusu3, K. Devriendt4, J. Schrander2, Y. Detisch1, B. Smeets1, A. Stegmann1, A. Paulussen1; 1Dpt Clinical Genetics, Maastricht,Netherlands, 2Dpt. Clinical Genetics, Maastricht, Netherlands, 3Dpt Clinical Genetics, Iasi, Romania, 4Dpt Clinical Genetics, Leuven, Belgium. Kabuki syndrome (KS, MIM: 147920) is a clinically recognizable syndrome of multiple congenital anomalies and mental retardation affecting approximately 1:30,000 live births. Key features are a characteristic face, growth retardation, developmental delay and additional features such as hypodontia and persistent foetal fingertip pads. Recently, a gene causing KS was identified through exome sequencing, reporting de novo mutations in the histone methyl transferase (HMT) gene MLL2 in 66% of 53 patients with Kabuki syndrome (Ng et al., Nat Genet.42(9):790-3, 2010). We confirmed the pathogenic significance of this gene in KS in our first published series, demonstrating 76% MLL2 mutations in 45 KS cases (Paulussen et al, Hum Mutat. 32(2), 2011). In another series of 31 patients, we identified 21 additional novel MLL2 mutations. Thus the combined mutation yield of Sanger-based MLL2 exon sequencing for both series is 55/76 = 72%, further supporting the major contribution of MLL2 mutations to the disease. We collected detailed clinical data of the 76 patients from both series (49 female, 27 male). Relevant clinical features were selected from the Dyscerne guidelines for KS (www. dyscerne.org) and interrogated using standardised questionnaires and checklists. Analysis focused on 7 clustered feature groups: facial, growth and development, cardiovascular, sensory (vision, speech, hearing), skeletal/orthopaedic malformations and accessory minor feautures. Statistical analysis using SPSS v12 was used to correlate (clustered) clinical features to MLL2 mutation carrier status We will present data from an extended pheno-genotype analysis, including the contribution of type-specific clustered mutations (i.e. nonsense, frameshift, missense and splice-site).
Riboflavin responsive OXPHOS complex I deficiency caused by
M. Gerards1,2, B. J. C. van den Bosch1,2, K. Danhauser3, V. Serre4, M. van Weeghel5, R. J. A. Wanders5, G. A. F. Nicolaes6, W. Sluiter7, K. Schoonderwoerd8, H. R. Scholte9, H. Prokisch3, A. Rötig4, I. F. M. de Coo10, H. J. M. Smeets1,2; 1Department of Genetics and Cell Biology, unit Clinical Genomics, Maastricht, Netherlands, 2School for Oncology and Developmental Biology, Maastricht University, Maastricht, Netherlands, 3Institute of Human Genetics, Technische Universität München, Germany and Institute of Human Genetics, Helmholtz Zentrum München, Munich, Germany, 4INSERM U781 and Department of Genetics, Hôpital Necker-Enfants Malades, Paris, France, 5Department of Clinical Chemistry, Laboratory Genetic Metabolic Diseases, Academic Medical Center, University of Amsterdam, Amsterdam, Netherlands, 6Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, Maastricht, Netherlands, 7Center for Lysosomal and Metabolic Diseases, Erasmus MC University Medical Center, Rotterdam, Netherlands, 8Department of Clinical Genetics, Erasmus MC University Medical Center, Rotterdam, Netherlands, 9Department of Neuroscience, Erasmus MC University Medical Center, Rotterdam, Netherlands, 10Department of Neurology, Erasmus MC University Medical Center, Rotterdam, Netherlands. Mitochondrial complex I deficiency is the most common OXPHOS defect. Mutations have been detected in mitochondrial and nuclear genes, but many patients remain unresolved and new genes are likely involved. In a consanguineous family, patients presented since early childhood with easy fatigability, exercise intolerance and lactic acidosis in blood. In muscle, subsarcolemmal mitochondrial proliferation and a severe complex I deficiency were observed. Exercise intolerance and complex I activity improved by supplementation with a high dosage of riboflavin. Homozygosity mapping revealed a candidate region on chromosome three containing six mitochondria-related genes. Four genes were screened for mutations and a homozygous substitution was identified in ACAD9 (c.1594C>T), changing the highly conserved arginine-532 into tryptophan. This mutation was absent in 188 ethnically matched controls. Protein modelling suggested a functional effect due to loss of a stabilizing hydrogen bond in an α-helix and a local flexibility change. To test whether the ACAD9 mutation caused the complex I deficiency, we transduced fibroblasts of patients with wild type and mutant ACAD9. Wild type ACAD9, but not mutant ACAD9, restored complex I activity. An unrelated patient with the same phenotype was compound heterozygous for c.380G>A and c.1405C>T, changing arginine-127 into glutamine and arginine-469 into tryptophan respectively. These amino acids were highly conserved and the substitutions were not present in controls, making them very likely pathogenic. Our data support a new function for ACAD9 in complex I function, making this gene an important new candidate for patients with complex I deficiency, which could be improved by riboflavin treatment.
Genes, viruses and inflammation in idiopathic (dilated)
I. P. C. Krapels, A. van den Wijngaard, R. J. E. Jongbloed, R. M. Dennert, C. G. M. J. Eurlings, A. T. J. M. Helderman-van den Enden, Y. H. J. M. Arens, S. R. H. Heymans; Maastricht University Medical Center, Maastricht, Netherlands.
Background: dilated cardiomyopathy (DCM) is characterized by impaired contractility and ventricular dilatation . Mutations in the currently known DCM-genes are only found in a small number of DCM patients and the clinical phenotype in families with a known DCM mutation varies considerably. Cardiotrophic viruses and/or inflammation also play an important role in DCM and could explain interindivuals clinical variability. We assessed the presence of genetic variants and cardiac viruses/ inflammation in a group of 30 patients with an idiopathic DCM. Methods: genetic analysis was performed on DNA through CardioCHIP analysis, a CHIP-based high throughput resequencing platform, simultaneously analysing 34 genes involved in inherited cardiomyopathies. The presence of cardiotrophic viruses and/or inflammation was assessed on cardiac biopsies. Results: 16 of the 30 patients (52%) had genetic variations at CardioCHIP analyses. Cardiac biopsy in these 30 patients revealed ParvoB19 virus infection (>500 copies per mcg isolated DNA) in 7 patients, inflammation (>14 CD45-cells/mm2) in 4 patients and active myocarditis in 1 patient. Of these 12 patients with viral infection and/ or inflammation (12/30=40%), 6 patients also had genetic alterations (6/12=50%) on cardioCHIP analysis. Family history was positive for sudden death in 3 of these 6 patients. We are currently performing DNA-analysis combined with cardiac biopsies in additional patients. Conclusion: although based on small numbers, these data demonstrate that viral or inflammatory components as a cause for DCM do not exclude a genetic component and vice versa. These data are important for genetic counseling of individuals with DCM.
Evaluation of RAD51C as a new breast cancer susceptibility
K. De Leeneer1, M. Van Bockstael1, N. Swietek1, J. Van den Ende2, K. Storm2, B. Blaumeiser2, K. Leunen3, C. J. van Asperen4, J. T. Wijnen4, E. Legius5, G. Michils5, G. Matthijs5, M. J. Blok6, E. B. Gomez-Garcia6, A. De Paepe1, B. Poppe1, K. Claes1; 1Center of Medical Genetics, Ghent University Hospital, Ghent, Belgium, 2Center of Medical Genetics, University Hospital of Antwerp, Antwerp, Belgium, 3Department of Gynaecological Oncology, Catholic University of Leuven, Leuven, Belgium, 4Dep. of clinical genetics, Leiden University Medical Center, Leiden, Netherlands, 5Department of Human Genetics, Catholic University of Leuven, Leuven, Belgium, 6Department of Clinical Genetics, Maastricht University Medical, Maastricht, Netherlands. Recently, germline mutations in RAD51C were found to be associated with an increased risk for breast and ovarian cancer. Meindl et al. (2010) detected six monoallelic pathogenic mutations in RAD51C by screening 1.100 unrelated German women with gynaecologic malignancies (breast and/or ovarian tumors). Strikingly, all six deleterious mutations were exclusively found within 480 BRCA1/2 negative breast and ovarian cancer families and not in breast cancer only families. With this study we aim to determine the prevalence of germline RAD51C mutations in Belgian/Dutch breast and ovarian cancer families, previously found to be negative for BRCA1&2 mutations. We performed mutational analysis in 350 index patients. Mutation detection was performed with High resolution melting curve analysis (HRMCA), followed by Sanger sequencing of the aberrant melting curves. Besides frequent single nucleotide polymorphisms and some novel, rare variants of which the clinical significance is currently under evaluation, we did not identify any deleterious mutation. To increase the number of probands to a similar number as in the initial report, we screened 100 additional families with HRMCA. Sequencing analysis of the fragments with aberrant melting curves is currently ongoing. We are the first group to investigate a cohort of breast and ovarian cancer families of comparable size as Meindl et al., and based on their findings, we had expected to detect at least three deleterious mutations. As other studies on (male) breast cancer families also did not identify deleterious germline RAD51C mutations, these may be less frequent than initially reported.
The BRCA1 variants c.692C>T and c.693G>A affect a putative
R. D. Brandăo1,2, K. van Roozendaal1, D. Tserpelis1, E. Gómez García1,2, M. J. Blok1; 1Maastricht University Hospital, Maastricht, Netherlands, 2GROW- School for Oncology and Developmental Biology, Maastricht, Netherlands. BRCA1 variants c.692C>T (p.T231M) and c.693G>A (p.=), of unknown clinical significance, were identified in two different patients from HBOC families. The variants lie in exon 11, within a region previously described as “critical region”, which spans between codons 200-300. Others suggested that this region was an important regulatory region and suggested the presence of two ESE motifs, one of these covering positions c.690 to c.695. We assessed the putative effect of these variants on RNA splicing. For splicing analysis, lymphocytes from the carriers and healthy controls were cultured and a fraction of these was treated with puromycin to prevent nonsense-mRNA mediated decay. The relative contribution of each allele to different transcripts was assessed based on allele specific cDNA analysis. In addition, an exon trapping vector including the putative ESE motif will be used to show its functionality in presence or absence of the variants. The alleles with either variant give rise to normal levels of the full-length transcript, but also induce higher expression of in-frame BRCA1Δ11 isoform, which was found to be weakly expressed in controls. Results of the ESE-dependent splicing assay will be reported. We now provide experimental evidence for the presence of a functional ESE motif in exon 11. Both variants in this motif affect splicing by increasing the expression of the BRCA1Δ11 transcript, besides expressing the full-length transcript. Currently, the clinical relevance of these variants remains elusive, as both the function and the critical expression level of the BRCA1Δ11 transcript are unknown.
Pre-implantation genetic diagnosis offers a fair chance of having unaffected. H.J.M. Smeets1,2, D.M.E.I. Hellebrekers1, R. Wolfe3, A.T.M. Hendrickx1, J.P.M. Geraedts1,2, P.F. Chinnery4, C.E. De Die1,2, I.F.M. De Coo5. 1) Dept Clinical Genetics, Maastricht UMC, Maastricht, Netherlands; 2) Research School GROW, Maastricht UMC, Maastricht, Netherlands; 3) School of Public Health and Preventive Medicine, Monash University, Melbourne, Australia; 4) Institute of Human Genetics, Newcastle University, Newcastle upon Tyne, UK; 5) Dept of Pediatric Neurology, Erasmus MC-Sophia Children's Hospital, Rotterdam, Netherlands. Mitochondrial disorders are often fatal multisystem disorders, for a significant part caused by heteroplasmic mtDNA point mutations. Prenatal diagnosis is in general not possible for these maternally inherited mutations due to extensive variation in mutation load among embryos and the inability to accurately predict the clinical expression. We hypothesized that preimplantation genetic diagnosis (PGD) should be a better alternative, if a minimal mutant level existed below which the chance for an embryo of being affected was acceptably low, irrespective of the mtDNA point mutation. We performed a systematic review of muscle mutant levels, evaluating 159 different heteroplasmic mtDNA point mutations derived from 327 unrelated patients or pedigrees, but excluding three overrepresented mtDNA mutations. Mutation levels were included for familial mtDNA point mutations only, covering all affected (n=195) and unaffected maternal relatives (n=19) from 137 pedigrees. Mean muscle mutant levels were comparable between probands and affected maternal relatives and between affected individuals with tRNA- versus protein-coding mutations. Using an estimated a-priori prevalence of being affected in pedigrees of 0.477, we calculated the risk of being affected at varying mutant levels. A 95% or higher chance of being unaffected was associated with a muscle mutant level of 18% or less. Most mtDNA mutation carriers will have oocytes below this threshold. Our data provide for the first time carriers of all heteroplasmic mtDNA point mutations a fair chance of having healthy offspring, by applying PGD.
Genetics of Kabuki Syndrome: MLL2 mutational spectrum in 100 KS. A.D.C. Paulussen1,3, M.J. Blok1,3, C.E. van Roozendaal1,3, D. Tserpelis1, C. Posma-Velter1, R. Vijzelaar4, J. Schrander2, Y. Detisch1,3, C.T.R.M. Schrander-Stumpel1,3, A.P.A. Stegmann1,3, H.J. Smeets1,3. 1) Dept Clinical Genomics, Maastricht Univ Med Ctr, Maastricht, Netherlands; 2) Department of Paediatrics, Maastricht UMC+,Maastricht, the Netherlands; 3) School for Oncology & Developmental Biology (GROW), Maastricht UMC+,Maastricht, the Netherlands; 4) MRC-Holland, Amsterdam, The Netherlands. Kabuki syndrome (KS, MIM: 147920) is a clinically recognizable syndrome of multiple congenital anomalies and mental retardation affecting approximately 1:30,000 live births. Key features are a characteristic face, growth retardation, developmental delay and additional features such as hypodontia and persistent foetal fingertip pads. Recently, a gene causing KS was identified through exome sequencing, reporting de novo mutations in the histone methyl transferase (HMT) gene MLL2 in 66% of 53 patients with Kabuki syndrome (Ng et al., Nat Genet.42(9):790-3, 2010). We confirmed the pathogenic significance of this gene in KS in our first published series, demonstrating 76% MLL2 mutations in 45 KS cases (Paulussen et al, Hum Mutat. 32(2), 2011). In a larger follow-up study we screened 100 patients for MLL2 mutations and included MLPAanalysis to study exonic deletions and duplications in the MLL2 mutation negative patients. Using Long-Range PCR we further examined the complete MLL2 gene, including introns and 5'/3' UTR in two typically clinical KS patients without a mutation in the coding region to search for exon deletions/duplications not covered by the MLPA kit and/ or deep-intronic de novo mutations. The effect of predicted splice-site mutations was investigated at the RNA level. Finally, we determined the mutational origin (paternal/maternal) of several mutations with allele specific PCR. Our data yields a more complete picture of the MLL2 mutational spectrum and further insight in the pathogenic mechanisms leading to KS.
Past and present investigations in a family with Christianson syndrome Kalscheuer4, W.O. Renier5, D. Tserpelis2, E.E. Smeets1,2, A.P. Stegmann1,6, R. Blok2,3, I.P. Krapels1,2, C.T.R.M. Schrander-Stumpel1,2, H.H. Ropers4, C.E.M. de Die-Smulders1,2. 1) Department of Clinical Genetics, Maastricht University Medical Center+, azM, Maastricht, the Netherlands; 2) Research Institute for Oncology and Developmental Biology, GROW, Maastricht University, Maastricht, The Netherlands; 3) Clinical Genomics, Department of Clinical Genetics, Maastricht University Medical Center+, azM, Maastricht, the Netherlands; 4) Max Planck Institute for Molecular Genetics, Department Human Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany; 5) Department of Neurology, Canisius-Wilhelmina Hospital, Nijmegen, The Netherlands; 6) Cytogenetic Laboratory, Department of Clinical Genetics, Maastricht University Medical Center+, azM, Maastricht, the Netherlands. Christianson syndrome (OMIM 300243) is a rare syndromic form of Xlinked intellectual disability disorder (XLID) which resembles clinically Angelman syndrome at young age. Affected men have profound cognitive dysfunction without or with nearly no speech. They show spasticity, ataxia and facial grimacing. They are relatively microcephalic and have cerebellar a-/hypoplasia. In adulthood, they develop a lean body and present with constant drooling. One-third of the carrier women have learning disabilities, dyslexia with or without additional clinical features. The underlying SLC9A6 (OMIM 300231) gene defect has been reported in families with Christianson syndrome. We present a new family with Christianson syndrome with additional brain pathology and molecular investigations. Clinically, classical features of Christianson syndrome were present in affected men. Open brain biopsy, performed in the past, in an affected man showed distinct pathological second and poorly differentiated third and fourth cerebral cortical layer using the Golgi-Cox staining. Cerebrospinal fluid investigations were normal, as well as other metabolic and infection parameters in blood. Carrier women had mild learning problems and were relatively microcephalic. Through next generation sequencing, the X-exome was systematically screened in this family. A SLC9A6 c.1481delG, p.Gly526fsX non-recurrent truncating mutation was identified which segregated with the clinical phenotype. Missense mutations in other causal XLID genes included GDI1 c.154-3C>T and SHROOM2 c.3608A>G (p.Asn1203Ser). Both mutations could, but FTSJ1 c.-87-10 (without splice effect after in silico analysis) variance could not be excluded through segregation analysis in this family. FTSJ1 expression analysis to investigate eventual modifying effect on the clinical phenotype in this family might be needed. We conclude, that through past and present investigations, the clinical phenotype of Christianson syndrome has further been delineated and that possible modifying effects of other XLID genes might interfere with the clinical phenotype or might explain inter and intra familial variability.. S.G.M. Frints1,2, C.E. van Roozendaal2,3, V.M.
Defective NDUFA9 as a novel cause of neonatally fatal complex I disease.
B.J.C. van den Bosch1,2, M. Gerards1,2, W. Sluiter3, A.P.A. Stegmann1, E.L.C. Jongen1, D.M.E.I. Hellebrekers1, E.H. Lambrichs2, H. Prokisch4,5, K. Danhauser4,5, K. Schoonderwoerd6, I.F.M. de Coo7, H.J.M. Smeets1,2. 1) Unit Clinical Genomics, Department of Clinical Genetics, Maastricht University, Maastricht, Netherlands; 2) School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands; 3) Centre for Lysosomal and Metabolic Diseases, Erasmus MC, Rotterdam, The Netherlands; 4) Institute of Human Genetics, Technische Universität München, Munich, Germany; 5) Institute of Human Genetics, Helmholtz Zentrum München, German Research Centre for Environmental Health, Neuherberg, Germany; 6) Department of Clinical Genetics, Erasmus MC, Rotterdam, The Netherlands; 7) Department of Neurology, Erasmus MC, Rotterdam, The Netherlands. Mitochondrial disorders are associated with abnormalities of the oxidative phosphorylation (OXPHOS) system and cause significant morbidity and mortality in the population. The extensive clinical and genetic heterogeneity of these disorders due to a broad variety of mutations in either the mitochondrial DNA (mtDNA) or nuclear DNA (nDNA) impedes a straightforward genetic diagnosis. Several approaches exist to identify the underlying gene defect, such as (large scale) screening of candidate genes, linkage analysis, homozygosity mapping and/or whole exome/genome sequencing. Here we present a single Kurdisch patient from consanguineous parents with neonatally fatal Leigh syndrome and complex I deficiency in which the gene defect was identified by homozygosity mapping and subsequent positional candidate gene analysis of complex I genes. A pathogenic mutation was identified in the complex I subunit encoding NDUFA9 gene, changing a highly conserved arginine at position 321 by proline, which is the first mutation reported for NDUFA9. Complex I activity was restored in fibroblasts of the patient by lentiviral transduction with wild type but not mutant NDUFA9, confirming that the mutation causes the complex I deficiency and related disease. Our data shows that homozygosity mapping and candidate gene analysis is still an efficient way to detect mutations in (single) consanguineous patients with OXPHOS deficiency, especially when the enzyme deficiency in fibroblasts allows appropriate candidate gene selection and functional complementation
Genes and viruses in idiopathic (dilated) cardiomyopathy; a pilot study.
I.PC. Krapels1, Y.H.J.M. Arens1, A.T.J.M. Helderman-van den Enden1, C. Eurlings2, R. Jongbloed1, A. van den Wijngaard1, S.R.B. Heymans2. 1) Clinical Genetics, Maastricht University Medical Center, Maastricht, Netherlands; 2) Cardiology, Maastricht University Medical Center, Maastricht, the Netherlands. Background: dilated cardiomyopathy (DCM) is characterized by impaired contractility and dilatation of the ventricles. Mutations in the currently known DCM-genes only explain a small number of DCM patients. The clinical phenotype in families with a known DCM mutation varies considerably. Cardiotropic viruses are involved in DCM and could explain the differences in clinical phenotype among individuals. We assessed the interaction between viral en genetic components in a group of idiopathic DCM patients. Methods: genetic analysis was performed through CardioCHIP analysis, a CHIP-based high throughput resequencing platform, simultaneously analysing 34 genes involved in inherited cardiomyopathies. The presence of cardiotropic viruses was assessed on the cardiac biopsies. Results: cardiac biopsy data was available for 177 patient; 59 (33%) of them had viral presence on cardiac biopsy (Parvo virus, >250 copies per mcg isolated DNA). One patient had an active myocarditis. Cardiochip data was available for 55 patients; 34 of them (62%) had genetic variations (either polymorphism, unclassified variant of mutation). The combined data of the Cardiochip analysis and cardiac biopsy was available for 30 patients. Of these 30 patients, 22 patients had no virus presence on biopsy. 12 of these 22 had genetic variants (55%), compared to 50% (4/8) in the group of patients with virus presence. Conclusion: Although based on small numbers, these data demonstrate that the presence of a viral infection as a cause for dilated cardiomyopathy does not exclude a genetic component. Further studies will address whether environmental factors such as a viral infections explain the difference in phenotype between patients and families with a known DCM mutation.
Evaluation of RAD51C as a new breast cancer susceptibility gene in the. K. Claes1, M. Van Bockstal1, S. De Brouwer1, N. Swietek1, K. Storm2, J. Van den Ende2, S. Willocx2, E.B. Gomez-Garcia6, B. Blaumeiser2, K. Leunen3, C.J. Van Asperen4, J. Wijnen4, E. Legius5, G. Michils5, G. Matthijs5, M.J. Blok6, A. De Paepe1, B. Poppe1, K. De Leeneer1. 1) Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium; 2) Center for Medical Genetics, University Hospital of Antwerp, Antwerp, Belgium; 3) Department of Gynaecological Oncology, Catholic University of Leuven, Leuven, Belgium; 4) Department of Clinical Genetics, Leiden University Medical Center, Leiden, Netherlands; 5) Department of Human Genetics, Catholic University of Leuven, Leuven, Belgium; 6) Department of Clinical Genetics, Maastricht University Medical, Maastricht, Netherlands. Recently, germline mutations in RAD51C were found to be associated with an increased risk for breast and ovarian cancer. Meindl et al. (2010) detected six monoallelic pathogenic mutations in RAD51C by screening 1.100 unrelated German women with gynaecologic malignancies (breast and/or ovarian tumors). Strikingly, all six deleterious mutations were exclusively found within 480 BRCA1/2 negative breast and ovarian cancer families and not in breast cancer only families. With this study we aim to determine the prevalence of germline RAD51C mutations in Belgian/Dutch breast and ovarian cancer families, previously found to be negative for BRCA1&2 mutations. We performed mutational analysis in 482 index patients. Mutation detection was performed with High resolution melting (HRM), followed by Sanger sequencing of the aberrant melting curves. No unequivocal deleterious RAD51C mutation was identified in our study population. In total our mutation analysis revealed 11 unique sequence variations of which 3 are novel. The most interesting is a novel 3'UTR variant: c.*131 A>G, identified in an ovarian cancer patient, diagnosed at the age of 60 and a positive family history of breast and colon cancer. In silico miRNA binding seed predictions (PITA Scan) revealed the creation of a miR-126 binding site. Functional assays are currently ongoing to find out if this variant alters AD51C expression mediated by miRNAs. Two other novel variants (RAD51C c.-36 A>G (5'UTR) and c.572-17 G>T (intronic)) were detected in our study population, none of them are predicted in silico to affect splicing. Since the initial publication, we and others were unable to confirm a major role for RAD51C germline mutations in breast and ovarian cancer families. Since the investigated cohort is of similar size and geographically close to the patients investigated in the initial report, we conclude that the prevalence of RAD51C mutations may be lower than initially expected. However, further investigations in populations of various ethnic origins are required before RAD51C can be excluded as a major breast and ovarian cancer susceptibility gene.
Veeck, R.D. Brandao, J.C. van Elssen, K.K. van de Vijver, P.J. Lindsey,
M.J. Blok, K. Keymeulen, T. Ayoubi, H.J. Smeets, P.S. Hupperets.
A Randomised Trial Of Pre-Operative Celecoxib
Treatment Reveals Anti-Tumour Transcriptional Response In Primary Breast Cancer.
Annals of Oncology, 2011; 22: Supplement 2