ABSTRACT
SLX4/FANCP is a recently discovered novel disease gene for Fanconi anemia (FA), a rare recessive disorder characterized by chromosomal instability and increased cancer susceptibility. Three ...of the 15 FA genes are breast cancer susceptibility genes in heterozygous mutation carriers—BRCA2, PALB2, and BRIP1. To investigate if defects in SLX4 also predispose to breast cancer, the gene was sequenced in a cohort of 729 BRCA1/BRCA2‐negative familial breast cancer cases. We identified a single splice site mutation (c.2013+2T>A), which causes a frameshift by skipping of exon 8. We also identified 39 missense variants, four of which were selected for functional testing in a Mitomycin C‐induced growth inhibition assay, and appeared indistinguishable from wild type. Although this is the first study that describes a truncating SLX4 mutation in breast cancer patients, our data indicate that germline mutations in SLX4 are very rare and are unlikely to make a significant contribution to familial breast cancer.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Objective
We previously identified a pathogenic germline DICER1 variant in a child with transposition of the great arteries who was a member of a family with DICER1 syndrome. In view of a report ...linking Dicer1 knockout in murine cardiomyocytes to cardiac outflow defects, we investigated the involvement of DICER1 in transposition of the great arteries.
Design
We used Fluidigm access array followed by next generation sequencing to screen for variants in the coding exons, their exon/intron boundaries and the 3′ untranslated region of DICER1 in patient DNA.
Cases
Germline DNA was collected from 129 patients with either sporadic or familial forms of transposition of the great arteries from two sites in Australia and Italy.
Results
Most cases (85%) did not have any germline DICER1 variants. In the remaining 15% of cases, we identified 16 previously reported variants (5 synonymous, 6 intronic, and 5 missense) and 2 novel variants (1 intronic and 1 missense). None of the identified variants were predicted to be pathogenic.
Conclusions
Here, we report that neither likely pathogenic nor pathogenic variants in DICER1 appear to play a major role in transposition of the great arteries.
Germ-line RB-1 mutations predispose to pineoblastoma (PinB), but other predisposing genetic factors are not well established. We recently identified a germline D1CER1 mutation in a child with a PinB. ...This was accompanied by loss of heterozygosity (LOH) of the wildtype allele within the tumour. We set out to establish the prevalence of D1CER1 mutations in an opportunistically ascertained series of PinBs. Twenty-one PinB cases were studied: eighteen cases had not undergone previous testing for D1CER1 mutations; three patients were known carriers of germ-line D1CER1 mutations. The eighteen PinBs were sequenced by Sanger and/or Fluidigm-based next-generation sequencing to identify D1CER1 mutations in blood gDNA and/or tumour gDNA. Testing for somatic D1CER1 mutations was also conducted on one case with a known germ-line D1CER1 mutation. From the eighteen PinBs, we identified four deleterious D1CER1 mutations, three of which were germ line in origin, and one for which a germ line versus somatic origin could not be determined; in all four, the second allele was also inactivated leading to complete loss of DICER1 protein. No somatic D1CER1 RNase IIIb mutations were identified. One PinB arising in a germline D1CER1 mutation carrier was found to have LOH. This Division of Children's Leukaemia and Cancer Research, Telethon Kids Institute, The University of Western Australia, Perth, Australia study suggests that germ-line D1CER1 mutations make a clinically significant contribution to PinB, establishing D1CER1 as an important susceptibility gene for PinB and demonstrates PinB to be a manifestation of a germ-line D1CER1 mutation. The means by which the second allele is inactivated may differ from other D1CER1 -related tumours. Keywords D1CER1 * miRNA processing * Paediatric brain tumours * Pineal gland * Childhood cancer * Mutation * Pineoblastoma * OMIM #601200
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Background Around half of familial breast cancer cases are caused by germ-line mutations in genes which are critically involved in the maintenance of genome stability. Mutations in related genes ...functioning in DNA repair may account for currently unattributed cases. Two such genes, RAP80 and Abraxas, have recently been identified to be in a complex with BRCA1, and are required for the localization of BRCA1 to DNA damage foci. Methods RAP80 and Abraxas variants were screened for in a cohort of 95 high risk, non-BRCA1/2 breast cancer cases of varying ethnicity: those of Ashkenazi Jewish (n = 35), mixed Canadian (n = 34) and Swiss descent (n = 26). Results We have identified four missense variants, four silent SNPs, three SNPs in the UTRs and seven intronic variants in RAP80. Two of the previously reported RAP80 variants were further investigated. In Abraxas, we have identified two missense, nine intronic and two variants in the 3' UTR. Conclusions Overall, it seems unlikely that moderate to highly penetrant alleles of either RAP80 or Abraxas, confer a significantly high relative risk of breast cancer.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
PALB2 has emerged as a breast cancer susceptibility gene. Mutations in PALB2 have been identified in almost all breast cancer populations studied to date, but the rarity of these mutations and lack ...of information regarding their penetrance makes genetic counseling for these families challenging. We studied BRCA1/2 -negative breast and/or ovarian cancer families to a) assess the contribution of PALB2 mutations in this series and b) identify clinical, pathological and family history characteristics that might make PALB2 screening more efficient.
The coding region of the PALB2 gene was analyzed in 175 probands with family histories of breast and/or ovarian cancer ascertained from a single Canadian institution in Eastern Ontario.
We identified 2 probands with PALB2 mutations that are known or strongly considered to be pathogenic and 3 probands with missense mutations that are possibly pathogenic. One of the identified truncating mutations c.3113G > A (p.Gly1000_Trp1038del - major product), has been previously described while the other four mutations c.3507_3508delTC (p.H1170Ffs*19), c.1846G > C (p.D616H), c.3418 T > G (p.W1140G), c.3287A > G (p.N1096S) have not been previously reported. Loss of heterozygosity was detected in two breast tumors from one c.3507_3508delTC mutation carrier but not in other available tumors from that family or in tumors from carriers of other mutations.
PALB2 mutation screening identifies a small, but significant number of mutations in BRCA1/2 -negative breast and/or ovarian cancer families. We show that mutations are more likely to be found in families with three or more breast cancers as well as other BRCA2-related cancers. In our cohort, both clearly pathogenic mutations were identified in premenopausal breast cancer cases (2/77, 2.6%). Testing should be preferentially offered to affected women from such families.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Byline: David J. Novak (1,2,3), Nelly Sabbaghian (1,3), Philippe Maillet (4), Pierre O. Chappuis (5), William D. Foulkes (1,2,3,6), Marc Tischkowitz (1,2,3) Keywords: Hereditary breast cancer; RAP80; ...Abraxas Background Around half of familial breast cancer cases are caused by germ-line mutations in genes which are critically involved in the maintenance of genome stability. Mutations in related genes functioning in DNA repair may account for currently unattributed cases. Two such genes, RAP80 and Abraxas, have recently been identified to be in a complex with BRCA1, and are required for the localization of BRCA1 to DNA damage foci. Methods RAP80 and Abraxas variants were screened for in a cohort of 95 high risk, non-BRCA1/2 breast cancer cases of varying ethnicity: those of Ashkenazi Jewish (n = 35), mixed Canadian (n = 34) and Swiss descent (n = 26). Results We have identified four missense variants, four silent SNPs, three SNPs in the UTRs and seven intronic variants in RAP80. Two of the previously reported RAP80 variants were further investigated. In Abraxas, we have identified two missense, nine intronic and two variants in the 3 UTR. Conclusions Overall, it seems unlikely that moderate to highly penetrant alleles of either RAP80 or Abraxas, confer a significantly high relative risk of breast cancer. Author Affiliation: (1) Departments of Oncology and Human Genetics, Program in Cancer Genetics, McGill University, Montreal, QC, Canada, H2W 1S6 (2) Departments of Medicine and Human Genetics, McGill University, Montreal, QC, Canada, H2W 1S6 (3) Segal Cancer Center, McGill University Sir M. B. Davis-Jewish General Hospital, 3755 Cote Ste-Catherine, Montreal, QC, Canada, H3T 1E2 (4) Laboratory of Molecular Oncology, Division of Laboratory Medicine, Department of Genetic Medicine and Laboratory, University Hospitals of Geneva, 1211, Geneva 14, Switzerland (5) Unit of Oncogenetics and Cancer Prevention, Division of Oncology, Department of Internal Medicine Division of Genetic Medicine, Department of Genetic Medicine and Laboratory, University Hospitals of Geneva, 1211, Geneva 14, Switzerland (6) The Research Institute, McGill University Health Centre, Montreal, QC, Canada, H3G 1A4 Article History: Registration Date: 11/07/2008 Received Date: 28/01/2008 Accepted Date: 11/07/2008 Online Date: 10/08/2008
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Massively parallel sequencing (MPS) has revolutionised biomedical research and offers enormous capacity for clinical application. We previously reported Hi-Plex, a streamlined highly-multiplexed ...PCR-MPS approach, allowing a given library to be sequenced with both the Ion Torrent and TruSeq chemistries. Comparable sequencing efficiency was achieved using material derived from lymphoblastoid cell lines and formalin-fixed paraffin-embedded tumour.
Here, we report high-throughput application of Hi-Plex by performing blinded mutation screening of the coding regions of the breast cancer susceptibility gene PALB2 on a set of 95 blood-derived DNA samples that had previously been screened using Sanger sequencing and high-resolution melting curve analysis (n = 90), or genotyped by Taqman probe-based assays (n = 5). Hi-Plex libraries were prepared simultaneously using relatively inexpensive, readily available reagents in a simple half-day protocol followed by MPS on a single MiSeq run.
We observed that 99.93% of amplicons were represented at ≥10X coverage. All 56 previously identified variant calls were detected and no false positive calls were assigned. Four additional variant calls were made and confirmed upon re-analysis of previous data or subsequent Sanger sequencing.
These results support Hi-Plex as a powerful approach for rapid, cost-effective and accurate high-throughput mutation screening. They further demonstrate that Hi-Plex methods are suitable for and can meet the demands of high-throughput genetic testing in research and clinical settings.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The RNase III enzyme DICER1 plays a central role in maturation of microRNAs. Identification of neoplasia-associated germ-line and somatic mutations in DICER1 indicates that mis-expression of miRNAs ...in cancer may result from defects in their processing. As part of a recent study of DICER1 RNase III domains in 96 testicular germ cell tumors, a single RNase IIIb domain mutation was identified in a seminoma. To further explore the importance of DICER1 mutations in the etiology of testicular germ cell tumors (TGCT), we studied germ-line DNA samples from 43 probands diagnosed with familial TGCT.
We carried out High Resolution Melting Curve Analysis of DICER1 exons 2-12, 14-19, 21 and 24-27. All questionable melt curves were subjected to confirmatory Sanger sequencing.Sanger sequencing was used for exons 13, 20, 22 and 23. Intron-exon boundaries were included in all analyses. We identified 12 previously reported single nucleotide polymorphisms and two novel single nucleotide variants. No likely deleterious variants were identified; notably no mutations that were predicted to truncate the protein were identified.
Taken together with previous studies, the findings reported here suggest a very limited role for either germ-line or somatic DICER1 mutations in the etiology of TGCT.
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