Christi J. van Asperen

TP53 germline mutation testing in 180 families suspected of Li–Fraumeni syndrome: mutation detection rate and relative frequency of cancers in different familial phenotypes

Background Li–Fraumeni syndrome (LFS) is a rare autosomal dominant cancer predisposition syndrome. Most families fulfilling the classical diagnostic criteria harbour TP53 germline mutations. However, TP53 germline mutations may also occur in less obvious phenotypes. As a result, different criteria are in use to decide which patients qualify for TP53 mutation analysis, including the LFS, Li–Fraumeni-like (LFL) and Chompret criteria. We investigated which criteria for TP53 mutation analysis resulted in the highest mutation detection rate and sensitivity in Dutch families. We describe the tumour spectrum in TP53-positive families and calculated tumour type specific relative risks. Method A total of 180 Dutch families referred for TP53 mutation analysis were evaluated. Tumour phenotypes were verified by pathology reports or clinical records. Results A TP53 germline mutation was identified in 24 families. When the Chompret criteria were used 22/24 mutations were detected (sensitivity 92%, mutation detection rate 21%). In LFS and LFL families 18/24 mutations were found (sensitivity 75%). The two mutations detected outside the ‘Chompret group’ were found in a child with rhabdomyosarcoma and a young woman with breast cancer. In the mutation carriers, in addition to the classical LFS tumour types, colon and pancreatic cancer were also found significantly more often than in the general population. Conclusion We suggest TP53 mutation testing for all families fulfilling the Chompret criteria. In addition, TP53 mutation testing can be considered in the event of childhood sarcoma and breast cancer before 30 years. In addition to the risk for established LFS tumour types, TP53-positive individuals may also have an elevated risk for pancreatic and colon cancer.

Exposure to diagnostic radiation and risk of breast cancer among carriers of BRCA1/2 mutations: retrospective cohort study (GENE-RAD-RISK)

Objective To estimate the risk of breast cancer associated with diagnostic radiation in carriers of BRCA1/2 mutations. Design Retrospective cohort study (GENE-RAD-RISK). Setting Three nationwide studies (GENEPSO, EMBRACE, HEBON) in France, United Kingdom, and the Netherlands, Participants 1993 female carriers of BRCA1/2 mutations recruited in 2006-09. Main outcome measure Risk of breast cancer estimated with a weighted Cox proportional hazards model with a time dependent individually estimated cumulative breast dose, based on nominal estimates of organ dose and frequency of self reported diagnostic procedures. To correct for potential survival bias, the analysis excluded carriers who were diagnosed more than five years before completion of the study questionnaire. Results In carriers of BRCA1/2 mutations any exposure to diagnostic radiation before the age of 30 was associated with an increased risk of breast cancer (hazard ratio 1.90, 95% confidence interval 1.20 to 3.00), with a dose-response pattern. The risks by quarter of estimated cumulative dose <0.0020 Gy, ≥0.0020-0.0065 Gy, ≥0.0066-0.0173 Gy, and ≥0.0174 Gy were 1.63 (0.96 to 2.77), 1.78 (0.88 to 3.58), 1.75 (0.72 to 4.25), and 3.84 (1.67 to 8.79), respectively. Analyses on the different types of diagnostic procedures showed a pattern of increasing risk with increasing number of radiographs before age 20 and before age 30 compared with no exposure. A history of mammography before age 30 was also associated with an increased risk of breast cancer (hazard ratio 1.43, 0.85 to 2.40). Sensitivity analysis showed that this finding was not caused by confounding by indication of family history. Conclusion In this large European study among carriers of BRCA1/2 mutations, exposure to diagnostic radiation before age 30 was associated with an increased risk of breast cancer at dose levels considerably lower than those at which increases have been found in other cohorts exposed to radiation. The results of this study support the use of non-ionising radiation imaging techniques (such as magnetic resonance imaging) as the main tool for surveillance in young women with BRCA1/2 mutations.

A genome wide linkage search for breast cancer susceptibility genes.

Mutations in known breast cancer susceptibility genes account for a minority of the familial aggregation of the disease. To search for further breast cancer susceptibility genes, we performed a combined analysis of four genome‐wide linkage screens, which included a total of 149 multiple case breast cancer families. All families included at least three cases of breast cancer diagnosed below age 60 years, at least one of whom had been tested and found not to carry a BRCA1 or BRCA2 mutation. Evidence for linkage was assessed using parametric linkage analysis, assuming both a dominant and a recessive mode of inheritance, and using nonparametric methods. The highest LOD score obtained in any analysis of the combined data was 1.80 under the dominant model, in a region on chromosome 4 close to marker D4S392. Three further LOD scores over 1 were identified in the parametric analyses and two in the nonparametric analyses. A maximum LOD score of 2.40 was found on chromosome arm 2p in families with four or more cases of breast cancer diagnosed below age 50 years. The number of linkage peaks did not differ from the number expected by chance. These results suggest regions that may harbor novel breast cancer susceptibility genes. They also indicate that no single gene is likely to account for a large fraction of the familial aggregation of breast cancer that is not due to mutations in BRCA1 or BRCA2.

Deletions spanning the neurofibromatosis type 1 gene: Implications for genotype-phenotype correlations in neurofibromatosis type 1?

Neurofibromatosis type 1 (NF1) is an autosomal‐dominant disorder characterized by abnormalities of tissues predominantly derived from the neural crest. Symptoms are highly variable and severity cannot be predicted, even within families. DNA of 84 unrelated patients with NF1, unselected for clinical features or severity, were screened with intragenic polymorphic repeat markers and by Southern analysis with cDNA probes. Deletions of the entire gene were detected in five patients from four unrelated families. Their phenotype resembled that of five previously reported patients with deletions, including intellectual impairment and dysmorphic features, but without an excessive number of dermal neurofibromas. This report supports the hypothesis that large deletions spanning the entire NF1 gene may lead to a specific phenotype. Hum Mutat 9:458–464, 1997

Intronic variants in BRCA1 and BRCA2 that affect RNA splicing can be reliably selected by splice-site prediction programs†

A large number of sequence variants identified in BRCA1 and BRCA2 cannot be distinguished as either disease‐causing mutations or neutral variants. These so‐called unclassified variants (UVs) include variants that are located in the intronic sequences of BRCA1 and BRCA2. The purpose of this study was to assess the use of splice‐site prediction programs (SSPPs) to select intronic variants in BRCA1 and BRCA2 that are likely to affect RNA splicing. We performed in vitro molecular characterization of RNA of six intronic variants in BRCA1 and BRCA2. In four cases (BRCA1, c.81–6T>A and c.4986+5G>T; BRCA2, c.7617+2T>G and c.8754+5G>A) a deleterious effect on RNA splicing was seen, whereas the c.135‐15_‐12del variant in BRCA1 showed no effect on RNA splicing. In the case of the BRCA2 c.68–7T>A variant, RNA analysis was not sufficient to establish the clinical significance. Six SSPPs were used to predict whether an effect on RNA splicing was expected for these six variants as well as for 23 intronic variants in BRCA1 for which the effect on RNA splicing has been published. Out of a total of 174 predictions, 161 (93%) were informative (i.e., the wild‐type splice‐site was recognized). No false‐negative predictions were observed; an effect on RNA splicing was always predicted by these programs. In four cases (2.5%) a false‐positive prediction was observed. For DNA diagnostic laboratories, these programs are therefore very useful to select intronic variants that are likely to affect RNA splicing for further analysis. Hum Mutat 0,1–8, 2008.

Clinical correlates of low-risk variants in FGFR2, TNRC9, MAP3K1, LSP1 and 8q24 in a Dutch cohort of incident breast cancer cases

IntroductionSeven SNPs in five genomic loci were recently found to confer a mildly increased risk of breast cancer.MethodsWe have investigated the correlations between disease characteristics and the patient genotypes of these SNPs in an unselected prospective cohort of 1,267 consecutive patients with primary breast cancer.ResultsHeterozygote carriers and minor allele homozygote carriers for SNP rs889312 in the MAP3K1 gene were less likely to be lymph node positive at breast cancer diagnosis (P = 0.044) relative to major allele homozygote carriers. Heterozygote carriers and minor allele homozygote carriers for SNP rs3803662 near the TNCR9 gene were more likely to be diagnosed before the age of 60 years (P = 0.025) relative to major allele homozygote carriers. We also noted a correlation between the number of minor alleles of rs2981582 in FGFR2 and the average number of first-degree and second-degree relatives with breast cancer and/or ovarian cancer (P = 0.05). All other disease characteristics, including tumour size and grade, and oestrogen or progesterone receptor status, were not significantly associated with any of these variants.ConclusionSome recently discovered genomic variants associated with a mildly increased risk of breast cancer are also associated with breast cancer characteristics or family history of breast cancer and ovarian cancer. These findings provide interesting new clues for further research on these low-risk susceptibility alleles.

Differences and similarities in breast cancer risk assessment models in clinical practice: which model to choose?

Abstract To show differences and similarities between risk estimation models for breast cancer in healthy women from BRCA1/2-negative or untested families. After a systematic literature search seven models were selected: Gail-2, Claus Model, Claus Tables, BOADICEA, Jonker Model, Claus-Extended Formula, and Tyrer–Cuzick. Life-time risks (LTRs) for developing breast cancer were estimated for two healthy counsellees, aged 40, with a variety in family histories and personal risk factors. Comparisons were made with guideline thresholds for individual screening. Without a clinically significant family history LTRs varied from 6.7% (Gail-2 Model) to 12.8% (Tyrer–Cuzick Model). Adding more information on personal risk factors increased the LTRs and yearly mammography will be advised in most situations. Older models (i.e. Gail-2 and Claus) are likely to underestimate the LTR for developing breast cancer as their baseline risk for women is too low. When models include personal risk factors, surveillance thresholds have to be reformulated. For current clinical practice, the Tyrer–Cuzick Model and the BOADICEA Model seem good choices.

Whole Exome Sequencing Suggests Much of Non-BRCA1/BRCA2 Familial Breast Cancer Is Due to Moderate and Low Penetrance Susceptibility Alleles

The identification of the two most prevalent susceptibility genes in breast cancer, BRCA1 and BRCA2, was the beginning of a sustained effort to uncover new genes explaining the missing heritability in this disease. Today, additional high, moderate and low penetrance genes have been identified in breast cancer, such as P53, PTEN, STK11, PALB2 or ATM, globally accounting for around 35 percent of the familial cases. In the present study we used massively parallel sequencing to analyze 7 BRCA1/BRCA2 negative families, each having at least 6 affected women with breast cancer (between 6 and 10) diagnosed under the age of 60 across generations. After extensive filtering, Sanger sequencing validation and co-segregation studies, variants were prioritized through either control-population studies, including up to 750 healthy individuals, or case-control assays comprising approximately 5300 samples. As a result, a known moderate susceptibility indel variant (CHEK2 1100delC) and a catalogue of 11 rare variants presenting signs of association with breast cancer were identified. All the affected genes are involved in important cellular mechanisms like DNA repair, cell proliferation and survival or cell cycle regulation. This study highlights the need to investigate the role of rare variants in familial cancer development by means of novel high throughput analysis strategies optimized for genetically heterogeneous scenarios. Even considering the intrinsic limitations of exome resequencing studies, our findings support the hypothesis that the majority of non-BRCA1/BRCA2 breast cancer families might be explained by the action of moderate and/or low penetrance susceptibility alleles.

Clinical Characteristics Affect the Impact of an Uninformative DNA Test Result: The Course of Worry and Distress Experienced by Women Who Apply for Genetic Testing for Breast Cancer

PURPOSE DNA mutation testing for breast cancer usually yields an uninformative result, which is a negative result in the absence of a known BRCA mutation within the family. However, few data are available on the psychological impact of this result. Moreover, the clinical heterogeneity within this group has not yet been considered. This study provides prospective data about the course of cancer-specific worry and distress for different groups of test applicants. PATIENTS AND METHODS All DNA test applicants (n = 238) completed three questionnaires: before and 1 and 7 months after disclosure of a DNA mutation test. With repeated-measures analysis of variance, differences were assessed between BRCA1/2-positive women (n = 42), BRCA1/2-true-negative women (n = 43), and women with an uninformative test result (n = 153). RESULTS On the group level, women with an uninformative result seemed to be reassured after disclosure (P < .001), but to a lesser extent than those women who received a true-negative result. However, not all women with an uninformative result reacted similarly: higher levels of worry and distress could be explained by relatively straightforward clinical variables, namely a personal history of cancer (P < or = .001) and a higher pedigree-based risk (P < or = .005). Furthermore, these clinical variables determined whether these women were either comparable to women who received a true-negative result or to BRCA mutation carriers. CONCLUSION Women with an uninformative result form a heterogeneous group of test applicants. The subpopulation of those with both a personal history of cancer and a relatively high pedigree-based risk expressed the highest levels of worry 7 months after DNA testing.

Genetic testing for familial/hereditary breast cancer-comparison of guidelines and recommendations from the UK, France, the Netherlands and Germany.

In this review, the national guidelines and recommendations for genetic testing for familial/hereditary breast cancer from the UK, France, the Netherlands and Germany were evaluated as to the inclusion criteria for genetic testing. In all four countries, access to genetic testing relies basically on the family history of breast and ovarian cancer. Similarities are obvious for most selection criteria. All four guidelines recommend embedding genetic testing within a framework of genetic counselling, and all agree to perform genetic testing first in an affected person. However, there are differences regarding the thresholds based on certain familial constellations, detailed description of selection criteria, the degree of relatedness between affected individuals and the counsellee, the age of diagnosis, the individual history of early onset breast cancer, bilateral breast cancer, the tumour morphology or the access to intensified surveillance. These differences and open questions not covered by the guidelines, e.g. on how to deal with phenocopies, unclassified variants, genetic variants in newly identified breast cancer susceptibility genes or with family constellations not fitting the criteria, are discussed. New evidence is usually slowly integrated into the guidelines. An exchange process towards the harmonization of the guidelines will ensure high quality health care across Europe.