Targeted sequencing of BRCA1 and BRCA2 across a large unselected breast cancer cohort suggests that one-third of mutations are somatic.

BACKGROUND: A mutation found in the BRCA1 or BRCA2 gene of a breast tumor could be either germline or somatically acquired. The prevalence of somatic BRCA1/2 mutations and the ratio between somatic and germline BRCA1/2 mutations in unselected breast cancer patients are currently unclear. PATIENTS AND METHODS: Paired normal and tumor DNA was analyzed for BRCA1/2 mutations by massively parallel sequencing in an unselected cohort of 273 breast cancer patients from south Sweden. RESULTS: Deleterious germline mutations in BRCA1 (n = 10) or BRCA2 (n = 10) were detected in 20 patients (7%). Deleterious somatic mutations in BRCA1 (n = 4) or BRCA2 (n = 5) were detected in 9 patients (3%). Accordingly, about 1 in 9 breast carcinomas (11%) in our cohort harbor a BRCA1/2 mutation. For each gene, the tumor phenotypes were very similar regardless of the mutation being germline or somatically acquired, whereas the tumor phenotypes differed significantly between wild-type and mutated cases. For age at diagnosis, the patients with somatic BRCA1/2 mutations resembled the wild-type patients (median age at diagnosis, germline BRCA1: 41.5 years; germline BRCA2: 49.5 years; somatic BRCA1/2: 65 years; wild-type BRCA1/2: 62.5 years). CONCLUSIONS: In a population without strong germline founder mutations, the likelihood of a BRCA1/2 mutation found in a breast carcinoma being somatic was ∼1/3 and germline 2/3. This may have implications for treatment and genetic counseling.

Familial aggregation of stroke amongst young patients in Lund Stroke Register.

BACKGROUND AND PURPOSE: The known monogenic forms of stroke are rare. The aim of this study was to analyze pedigrees of young stroke patients regarding possible monogenic cerebrovascular disease and to evaluate the possibility of genetic stroke in these families. This may contribute to a better understanding of disease mechanism in stroke. METHODS: Lund Stroke Register includes consecutive patients with first-ever stroke from a defined geographical area in southern Sweden. Early-onset (≤55 years) stroke patients were systematically screened with regard to family history (FHx), and families with stroke aggregation were compiled. Participants provided information in a questionnaire on occurrence of stroke or transient ischaemic attack (TIA) in their families. Information on cardiovascular risk factors (VRFs) and clinical stroke subtype was collected. FHx for stroke was considered positive when the patient reported either ≥1 first-degree relative with stroke/TIA, or no first-degree relative but ≥3 second- or third-degree relatives with stroke/TIA in a distribution compatible with monogenic inheritance. RESULTS: Of 4103 stroke patients registered, 426 (10%) had first-ever stroke at ≤55 years and 338 (79%) of these answered the questionnaire. Of them, 159 (47%) reported a positive FHx. Twenty-eight (18%) of the probands with positive FHx had no known VRFs. Thirty-two families with ≥4 members with stroke were identified. In all these larger families the affected individuals with stroke were present in more than one generation. CONCLUSION: Aggregation of stroke in families of early-onset stroke patients is not uncommon. Genetic factors with impact on stroke risk, including monogenic causes, need to be evaluated in future stroke studies.

Health needs assessment for medical genetic services for congenital disorders in middle- and low-income nations.

Medical genetic services for the care and prevention of congenital disorders have received little attention in most middle- and low-income countries to date. In 2010, the World Health Organisation prioritized services for the care and prevention of birth defects in these nations, emphasising their importance in assisting such countries to reach their Millennium Development Goals. Health Needs Assessment is an inclusive, rational, epidemiological-assisted approach for providing information to plan, introduce and beneficially change health care services to improve the health of populations. It is intrinsic to much of the development of health care systems in industrialised nations. Its use by middle- and low-income countries to introduce and develop medical genetic services commensurate with their needs and circumstances would be beneficial. An approach to applying Health Needs Assessment in these circumstances is described.

Are 25 SNPs from the CARDIoGRAM study associated with ischaemic stroke?

BACKGROUND AND PURPOSE: The Coronary Artery Disease Genome-Wide Replication and Meta-Analysis Study (CARDIoGRAM) reported 25 single-nucleotide polymorphisms (SNPs) on 15 chromosomes to be associated with coronary artery disease (CAD) risk. Because common vascular risk factors are shared between CAD and ischaemic stroke (IS), these SNPs may also be related to IS overall or one or more of its pathogenetic subtypes. METHODS: We performed a candidate gene study comprising 3986 patients with IS and 2459 control subjects. The 25 CAD-associated SNPs reported by CARDIoGRAM were examined by allelic association analysis including logistic regression. Weighted and unweighted genetic risk scores (GRSs) were also compiled and likewise analysed against IS. We furthermore considered the IS main subtypes large-vessel disease (LVD), small-vessel disease and cardioembolic stroke [according to Trial of Org 10172 in Acute Stroke Treatment (TOAST)] separately. RESULTS: SNP rs4977574 on chromosome 9p21.3 was associated with overall IS [odds ratio (OR) = 1.12; 95% confidence interval (CI): 1.04-1.20; P = 0.002] as well as LVD (OR = 1.36; 95% CI: 1.13-1.64; P = 0.001). No other SNP was significantly associated with IS or any of its main subtypes. Analogously, the GRSs did not show any noticeable effect. CONCLUSIONS: Besides the previously reported association with SNPs on chromosome 9p21, this study did not detect any significant association between IS and CAD-susceptible genetic variants. Also, GRSs compiled from these variants did not predict IS or any pathogenetic IS subtype, despite a total sample size of 6445 participants.

Association of PHB 1630 C>T and MTHFR 677 C>T polymorphisms with breast and ovarian cancer risk in BRCA1/2 mutation carriers: results from a multicenter study.

BACKGROUND: The variable penetrance of breast cancer in BRCA1/2 mutation carriers suggests that other genetic or environmental factors modify breast cancer risk. Two genes of special interest are prohibitin (PHB) and methylene-tetrahydrofolate reductase (MTHFR), both of which are important either directly or indirectly in maintaining genomic integrity. METHODS: To evaluate the potential role of genetic variants within PHB and MTHFR in breast and ovarian cancer risk, 4102 BRCA1 and 2093 BRCA2 mutation carriers, and 6211 BRCA1 and 2902 BRCA2 carriers from the Consortium of Investigators of Modifiers of BRCA1 and BRCA2 (CIMBA) were genotyped for the PHB 1630 C>T (rs6917) polymorphism and the MTHFR 677 C>T (rs1801133) polymorphism, respectively. RESULTS: There was no evidence of association between the PHB 1630 C>T and MTHFR 677 C>T polymorphisms with either disease for BRCA1 or BRCA2 mutation carriers when breast and ovarian cancer associations were evaluated separately. Analysis that evaluated associations for breast and ovarian cancer simultaneously showed some evidence that BRCA1 mutation carriers who had the rare homozygote genotype (TT) of the PHB 1630 C>T polymorphism were at increased risk of both breast and ovarian cancer (HR 1.50, 95%CI 1.10-2.04 and HR 2.16, 95%CI 1.24-3.76, respectively). However, there was no evidence of association under a multiplicative model for the effect of each minor allele. CONCLUSION: The PHB 1630TT genotype may modify breast and ovarian cancer risks in BRCA1 mutation carriers. This association need to be evaluated in larger series of BRCA1 mutation carriers.

Genetic screening in Europe.

Genetic screening has been defined as any kind of test performed systematically for the early detection or exclusion of a genetic disease, genetic predisposition or resistance to a disease, or to determine whether a person carries a gene variant that may produce disease in his or her offspring. In comparison to 'genetic testing', the term 'genetic screening' should be reserved for the explicit and systematic application of a diagnostic genetic test across a whole population of asymptomatic people (population screening) or a subset of a population such as pregnant women (prenatal/antenatal screening) or newborn infants (neonatal screening). This survey intends to present the current (2006-2008) status of genetic screening and the organization of genetic screening programmes in selected European countries as a background for future attempts to harmonize standards and procedures of genetic screening, an explicit aim of the European Network of Excellence, EuroGentest (www.eurogentest.org). Our report builds on the first comprehensive assessment of genetic screening programmes in Germany by the European Society of Human Genetics, starting with a workshop of experts in 1999, the production of background documentation in 2000, and a final report in 2003.

External quality assessment for mutation detection in the BRCA1 and BRCA2 genes: EMQN's experience of 3 years.

BACKGROUND: The European Molecular Genetics Quality Network (EMQN) was formed in order to improve external quality assessment for molecular genetic testing in Europe. From 1999 to 2002 it received funding from the European Union under the Standards, Measurement and Testing programme (contract no. SMT4-CT98-7515). Since then, its maintenance has been supported through subscription of the participants, and it has been coordinated by the National Genetic Reference Laboratory at Manchester, UK (Rob Elles and Simon Patton; www.emqn.org). MATERIALS AND METHODS: Among other external quality assessment (EQA) schemes, EMQN has provided an EQA scheme for mutation detection in the breast cancer genes, BRCA1 and BRCA2, designed to cover the two important aspects of genetic testing: (i) genotyping and (ii) interpretation and reporting of results. The fourth full scheme was completed in 2003, with data evaluation pending for the 47 participants. RESULTS: Analysis of genotyping data has pinpointed two main types of errors: (i) missing a mutation (in nine of the 17 false results a normal sequence was reported); and (ii) description of the observed sequence change by an incorrect nomenclature. Compared with the more technical process of genotyping, the writing of reports displayed a much wider variation between laboratories. CONCLUSIONS: From the reported data it is clear that external quality control should become an integral part of quality assessment in the laboratory, thus contributing to maintaining confidence in the reliability of genetic testing among patients and health professionals.

One in 10 ovarian cancer patients carry germ line BRCA1 or BRCA2 mutations: results of a prospective study in Southern Sweden.

At least 10% of all ovarian cancers are estimated to have a hereditary background. Hereditary breast-ovarian cancer (HBOC) due to mutations in the BRCA genes is a major cause of hereditary ovarian cancer, although its frequency and relationship to age and family history in unselected series of ovarian cancers is not completely known. We report here the results of a full mutational screening analysis for germ line BRCA1 and BRCA2 mutations in 161 patients with invasive epithelial ovarian carcinomas. Age at diagnosis ranged from 22 to 82 years (mean 59 years). Deleterious (frame-shift, nonsense and missense) mutations were detected in 13/161 (8%) of the patients and affected BRCA1 in 12 cases and BRCA2 in one case. Four additional missense variants (one in BRCA1 and three in BRCA2) with a possible association with an increased risk ovarian cancer were revealed, resulting in a total frequency of BRCA gene alterations of 17/161 (11%). The 13 patients with deleterious mutations had a mean age of 57 years (range 41-76 years) and only three of these patients were below 50 years of age. A family history of at least one breast cancer and/or ovarian cancer was reported in all but 1 of the patients with BRCA mutations compared with only 24% of patients without mutations. Our findings in this prospective study confirm approximately 1 in 10 patients with ovarian cancer carry a germ line BRCA gene mutation associated with HBOC, and also indicate that a large number of these patients are over 50 years of age at diagnosis.

HRPT2, encoding parafibromin, is mutated in hyperparathyroidism-jaw tumor syndrome.

We report here the identification of a gene associated with the hyperparathyroidism-jaw tumor (HPT-JT) syndrome. A single locus associated with HPT-JT (HRPT2) was previously mapped to chromosomal region 1q25-q32. We refined this region to a critical interval of 12 cM by genotyping in 26 affected kindreds. Using a positional candidate approach, we identified thirteen different heterozygous, germline, inactivating mutations in a single gene in fourteen families with HPT-JT. The proposed role of HRPT2 as a tumor suppressor was supported by mutation screening in 48 parathyroid adenomas with cystic features, which identified three somatic inactivating mutations, all located in exon 1. None of these mutations were detected in normal controls, and all were predicted to cause deficient or impaired protein function. HRPT2 is a ubiquitously expressed, evolutionarily conserved gene encoding a predicted protein of 531 amino acids, for which we propose the name parafibromin. Our findings suggest that HRPT2 is a tumor-suppressor gene, the inactivation of which is directly involved in predisposition to HPT-JT and in development of some sporadic parathyroid tumors.

Family history of breast and ovarian cancers and BRCA1 and BRCA2 mutations in a population-based series of early-onset breast cancer.

BACKGROUND: BRCA1 and BRCA2 are the two major susceptibility genes involved in hereditary breast cancer. This study was undertaken to provide reliable population-based estimates of genetic influence and to characterize the nature and prevalence of BRCA1 and BRCA2 germline mutations in early-onset breast cancer. METHODS: In a series comprising all women diagnosed with breast cancer under the age of 41 years in southern Sweden during 1990 through 1995 (n = 262), family history of cancer was evaluated in 95% (n = 250) of the case subjects and germline mutations in BRCA1 and BRCA2 were analyzed in 89% (n = 234). All statistical tests were two-sided. RESULTS: A total of 97 case subjects had at least one first- or second-degree relative with breast or ovarian cancer; 34 (14%; 95% confidence interval [CI] = 9.6% to 18%) cases had at least two first- or second-degree relatives, 22 (8.8%; 95%CI = 5.3% to 12%) had one first-degree relative, and 41 (16%; 95% CI = 12% to 21%) had one second-degree relative with either cancer. If two females affected with breast or ovarian cancer who were related through an unaffected male were also defined as first-degree relatives, then a higher number of case subjects, 120 (48%; 95% CI = 42% to 54%), had at least one first-degree or second-degree relative with breast or ovarian cancer. Sixteen (6.8%; 95% CI = 4.0% to 11%) BRCA1 mutation carriers and five (2.1%; 95% CI = 0.70% to 4.9%) BRCA2 mutation carriers were identified. Among case subjects with one first- or more than one first- or second-degree relative with breast or ovarian cancer, BRCA mutations were more frequent (P<.001) than among the case subjects without this degree of family history. BRCA mutations were also statistically significantly more common among women with bilateral breast cancer than among women with unilateral breast cancer (P =.002). BRCA mutations were more common among younger case subjects than among older ones (P =.0027). CONCLUSIONS: Almost half (48%) of women in southern Sweden with early-onset breast cancer have some family history of breast or ovarian cancer, and 9.0% of early-onset breast cancer cases are associated with a germline mutation in BRCA1 or BRCA2. Mutation carriers were more prevalent among young women, women with at least one first- or second-degree relative with breast or ovarian cancer, and women with bilateral breast cancer.

Functional analysis of BRCA1 C-terminal missense mutations identified in breast and ovarian cancer families.

Germline mutations in the breast and ovarian cancer susceptibility gene BRCA1 are responsible for the majority of cases involving hereditary breast and ovarian cancer. Whereas all truncating mutations are considered as functionally deleterious, most of the missense variants identified to date cannot be readily distinguished as either disease-associated mutations or benign polymorphisms. The C-terminal domain of BRCA1 displays an intrinsic transactivation activity, and mutations linked to disease predisposition have been shown to confer loss of such activity in yeast and mammalian cells. In an attempt to clarify the functional importance of the BRCA1 C-terminus as a transcription activator in cancer predisposition, we have characterized the effect of C-terminal germline variants identified in Scandinavian breast and ovarian cancer families. Missense variants A1669S, C1697R, R1699W, R1699Q, A1708E, S1715R and G1738E and a truncating mutation, W1837X, were characterized using yeast- and mammalian-based transcription assays. In addition, four additional missense variants (V1665M, D1692N, S1715N and D1733G) and one in-frame deletion (V1688del) were included in the study. Our findings demonstrate that transactivation activity may reflect a tumor-suppressing function of BRCA1 and further support the role of BRCA1 missense mutations in disease predisposition. We also report a discrepancy between results from yeast- and mammalian-based assays, indicating that it may not be possible to unambiguously characterize variants with the yeast assay alone. We show that transcription-based assays can aid in the characterization of deleterious mutations in the C-terminal part of BRCA1 and may form the basis of a functional assay.

Identification of human candidate genes for male infertility by digital differential display.

Evidence for the importance of genetic factors in male fertility is accumulating. In the literature and the Mendelian Cytogenetics Network database, 265 cases of infertile males with balanced reciprocal translocations have been described. The candidacy for infertility of 14 testis-expressed transcripts (TETs) were examined by comparing their chromosomal mapping position to the position of balanced reciprocal translocation breakpoints found in the 265 infertile males. The 14 TETs were selected by using digital differential display (electronic subtraction) to search for apparently testis-specific transcripts in the TIGR database. The testis specificity of the 14 TETs was further examined by reverse transcription-polymerase chain reaction (RT-PCR) on adult and fetal tissues showing that four TETs (TET1 to TET4) were testis-expressed only, six TETs (TET5 to TET10) appeared to be differentially expressed and the remaining four TETs (TET11 to TET14) were ubiquitously expressed. Interestingly, the two tesis expressed-only transcripts, TET1 and TET2, mapped to chromosomal regions where seven and six translocation breakpoints have been reported in infertile males respectively. Furthermore, one ubiquitously, but predominantly testis-expressed, transcript, TET11, mapped to 1p32-33, where 13 translocation breakpoints have been found in infertile males. Interestingly, the mouse mutation, skeletal fusions with sterility, sks, maps to the syntenic region in the mouse genome. Another transcript, TET7, was the human homologue of rat Tpx-1, which functions in the specific interaction of spermatogenic cells with Sertoli cells. TPX-1 maps to 6p21 where three cases of chromosomal breakpoints in infertile males have been reported. Finally, TET8 was a novel transcript which in the fetal stage is testis-specific, but in the adult is expressed in multiple tissues, including testis. We named this novel transcript fetal and adult testis-expressed transcript (FATE).

Disease associated balanced chromosome rearrangements: a resource for large scale genotype-phenotype delineation in man.

Disease associated balanced chromosomal rearrangements (DBCRs), which truncate, delete, or otherwise inactivate specific genes, have been instrumental for positional cloning of many disease genes. A network of cytogenetic laboratories, Mendelian Cytogenetics Network (MCN), has been established to facilitate the identification and mapping of DBCRs. To get an estimate of the potential of this approach, we surveyed all cytogenetic archives in Denmark and southern Sweden, with a population of approximately 6.6 million. The nine laboratories have performed 71 739 postnatal cytogenetic tests. Excluding Robertsonian translocations and chromosome 9 inversions, we identified 216 DBCRs ( approximately 0.3%), including a minimum estimate of 114 de novo reciprocal translocations (0.16%) and eight de novo inversions (0.01%). Altogether, this is six times more frequent than in the general population, suggesting a causal relationship with the traits involved in most of these cases. Of the identified cases, only 25 (12%) have been published, including 12 cases with known syndromes and 13 cases with unspecified mental retardation/congenital malformations. The remaining DBCRs were associated with a plethora of traits including mental retardation, dysmorphic features, major congenital malformations, autism, and male and female infertility. Several of the unpublished DBCRs defined candidate breakpoints for nail-patella, Prader-Willi, and Schmidt syndromes, ataxia, and ulna aplasia. The implication of the survey is apparent when compared with MCN; altogether, the 292 participating laboratories have performed >2.5 million postnatal analyses, with an estimated approximately 7500 DBCRs stored in their archives, of which more than half might be causative mutations. In addition, an estimated 450-500 novel cases should be detected each year. Our data illustrate that DBCRs and MCN are resources for large scale establishment of phenotype-genotype relationships in man.

Risk perception, screening practice and interest in genetic testing among unaffected men in families with hereditary prostate cancer.

Approximately 5-10% of prostate cancer cases are caused by dominantly inherited susceptibility to the disease. Although advances have been made in research concerning the genetic mechanisms of hereditary prostate cancer, little is known about the psychological consequences for men at high risk of developing the disease. The aims of the present study were to examine risk perception, interest in genetic investigations, cancer-specific worry, and screening practice among unaffected men, aged 40-72 years old, with a pedigree consistent with hereditary prostate cancer and an estimated lifetime risk of prostate cancer of 35-45%. A questionnaire was sent by mail to 120 subjects, of whom 110 responded. Most of the men (n = 90, 82%) worried about having an inherited susceptibility to prostate cancer, and 34 (31%) claimed that worry about prostate cancer affected their daily life (3 (3%) fairly much, 31 (28%) slightly). As many as 40% of the study subjects perceived their lifetime risk of prostate cancer as 67% or more. Perceived high risk was associated with symptoms of depression and with cancer worry affecting daily living. Two-thirds of the men aged 50 years old or more were regularly screened for prostate cancer. Subjects with high levels of cancer-specific stress, as measured by the avoidance subscale of the Impact of Event Scale, were less likely to opt for screening. Almost all of the men (94%) were interested in presymptomatic genetic testing (84 (76%) "definitely yes" and 20 (18%) "probably yes"). We conclude that hereditary susceptibility to prostate cancer has significant psychological consequences although it rarely causes psychiatric morbidity. The present study underlines the importance of giving thorough, repeated information to men at high risk of prostate cancer.

Novel germline APC mutations in Swedish patients with familial adenomatous polyposis and Gardner syndrome.

BACKGROUND: Familial adenomatous polyposis (FAP) is a familial cancer syndrome in which affected individuals develop multiple adenomatous polyps and are thereby at greatly increased risk of developing colorectal cancer. Gardner syndrome is a variant of FAP, in which the patients also develop extraintestinal tumors, in particular osteomas and desmoid tumors. An attenuated form of the disease (AFAP) is associated with fewer polyps, but still a high risk for colorectal cancer. Germline mutations in the adenomatosis polyposis coli (APC) gene cause FAP and Gardner syndrome and have recently been associated also with the development of AFAP. METHODS: We have analysed the entire APC gene for germline mutations in 7 patients with FAP and in 6 patients with suspected AFAP. Mutation screening was performed by direct sequencing of exons 1-14 and using the protein truncation test for analysis of exon 15. RESULTS: Novel disease-causing germline mutations, all of which resulted in truncation of the APC protein, were identified in 6 of the 7 patients with FAP or Gardner syndrome. No APC mutation was detected in any of the 6 patients with suspected AFAP. CONCLUSIONS: This study reports novel FAP- and Gardner syndrome-causing mutations in the APC gene. The lack of APC mutations in patients with multiple polyps at young age indicates that other genetic defects may cause this phenotype.