Multiple-case Breast Cancer Families Research Articles

Detecting BRCA2 Protein Truncation in Tissue Biopsies to Identify Breast Cancers That Arise in BRCA2 Gene Mutation Carriers

Mutations in the BRCA2 gene are dominantly inherited but cause cancers when the wild-type allele has loss of heterozygosity (LOH) within the cancer. Because most disease-associated BRCA2 mutations are protein-truncating mutations, a test for truncated BRCA2 proteins should identify most BRCA2 hereditary cancers. We have developed a tissue truncation test to identify truncated BRCA2 proteins in breast cancer tissue biopsies in vivo that does not use amplification or genetic manipulations. N-terminal and C-terminal antibodies are used to visualize protein truncation by demonstrating that the beginning of the protein is present but the end (ie, terminus) is absent. A quantitative C-terminal immunostaining score or a C-terminal to N-terminal truncation ratio correctly classified 20 of 21 breast cancers arising in BRCA2 mutation carriers and 57 of 58 cancers arising outside the context of a multiple-case breast cancer family. This represents a sensitivity of 95% and a specificity of 98%. Because of the presence of C-terminal BRCA2 protein and atypical clinical features of the misclassified cancer in a BRCA2 mutation carrier, we performed polymerase chain reaction and sequence analyses on this cancer. The results showed continued presence of the BRCA2 wild-type allele in the cancer, which indicated that intact BRCA2 protein was present in this cancer. This immunohistochemistry-based test (which takes only 4 hours) appears to identify BRCA2 hereditary cancer with high accuracy. The test also appears to diagnose the biochemical loss of BRCA2 protein in cancers (ie, BRCA2-mutant genotype), which will usually but not always agree with the presence of a germline BRCA2 mutation found by susceptibility testing by DNA sequencing of blood samples.

Predictors of the use of complementary and alternative medicine (CAM) by women at high risk for breast cancer

Background Few data exist regarding the use of complementary and alternative medicine (CAM) by unaffected women at high risk of breast cancer. Methods Self-reported CAM use by women from multiple-case breast cancer families was obtained by questionnaire. Factors associated with CAM use were assessed using multiple logistic regression. Results Of 892 women, 55% ( n = 489) used CAM, 6% ( n = 53) specifically to prevent cancer. CAM use was independently associated with tertiary education level (OR 2.56, 95% CI 1.83–3.58, p < 0.001), greater physical activity (OR 1.05 per hour of physical activity/week, 95% CI 1.00–1.10, p = 0.049), greater anxiety (OR 1.92, 95% CI 1.16–3.16, p = 0.01), not currently smoking (OR 0.64, 95% CI 0.42–0.97, p = 0.037) and lower perceived BC risk (OR 0.82 per 20 percentage points, 95% CI 0.72–0.94, p = 0.005). Conclusions The majority of high-risk women use CAM, but mostly for reasons other than cancer prevention. Most predictors of CAM use are consistent with the limited literature for women at high risk for cancer.

The RAD51D E233G variant and breast cancer risk: population-based and clinic-based family studies of Australian women

RAD51D is a homolog of the RAD51 protein, which is known to be an important component of the DNA repair pathway. A rare missense variant in the RAD51D gene, E233G (c.A>G), has been reported to be more prevalent in breast cancer cases from specific multiple-case breast cancer families, with an odds ratio of 2.6 (95% confidence interval (CI): 1.12-6.03). We assessed whether this variant was associated with breast cancer risk using two studies: a population-based case-control-family study based on 1,110 cases and 629 controls, and a clinic-based study based on 390 cases from multiple-case breast cancer families. We conducted case-control analyses and modified segregation analyses of carrier families. The carrier frequencies (95% CI) of the RAD51D variant were 4.1% (2.4-6.6) for clinic-based cases, 3.9% (2.8-5.2) for population-based cases, and 3.7% (2.3-5.4) for population-based controls, and were not significantly higher in case groups than controls (P=0.7 and P=0.8, respectively). After genotyping the relatives of cases who carried the variant, modified segregation analyses of these families were conducted, and the estimated hazard ratio for breast cancer corresponding to the E233G variant was 1.30 (95% CI: 0.66-2.58; P=0.4) for familial breast cancer families and 1.28 (95% CI: 0.47-3.43; P=0.6) for families unselected for family history. Therefore, despite being well powered to detect moderate risks, no evidence for an association between the E233G variant and breast cancer risk was observed in any setting. Larger studies would be required to determine if this variant is associated with a smaller risk of breast cancer.

Is MSH2 a breast cancer susceptibility gene?

Mutations in the DNA mismatch repair gene MSH2 lead to increased replication error and microsatellite instability and account for a substantial proportion of hereditary non-polyposis colorectal cancer (Lynch syndrome). A recent international collaborative genome-wide linkage scan (GWS) for breast cancer susceptibility loci found some evidence for there being a breast cancer susceptibility gene in a genomic region on chromosome 2p close to MSH2. We sought to investigate the possibility that mutations in MSH2 might explain the multiple cases of breast cancer in some families that were included in the international GWS. DNA samples from the affected probands of 59 multiple-case breast cancer families, many of whom gave LOD scores >0.5 in the MSH2 region, were screened for large genomic alterations in MSH2 via the Multiplex Ligation-dependent Probe Amplification (MLPA) assay and for coding region mutations via exonic sequencing. Several of the families also contained cases of colorectal cancer in addition to breast cancer and had been included in the GWS that had identified a positive LOD score on chromosome 2p. Using MLPA, c.1236C > T was identified in one proband but this variant was not predicted to create an alternate acceptor/donor site within exon 7 MSH2 using in silico analyses. A c.1734T > C was identified in a second proband via exonic sequencing but testing of the variant in other family members did not support segregation of this variant with disease. Extensive screening of 59 multiple-case breast cancer families did not identify any coding region mutations or larger genomic alterations in MSH2 that might implicate MSH2 as a breast cancer susceptibility gene.

Progesterone receptor polymorphisms and risk of breast cancer: results from two Australian breast cancer studies

Association studies aimed at identifying breast cancer susceptibility variants in the progesterone receptor (PGR) gene have been previously reported in the literature with conflicting results, ranging from a protective effect conferred by the PROGINS allele in German Caucasian women, to an increased risk for the leucine variant of the Val660Leu (rs1042838) polymorphism in East Anglian cases. We report the results of genotype and haplotype analyses of five PGR polymorphisms, +44C/T (rs518162), +331G/A (rs10895068), V660L (rs1042838), H770H (rs1042839) and Q886Q (rs500760), conducted on 1,847 Australian breast cancer cases (including 276 cases from a cohort of multiple-case breast cancer families) and 833 controls. Genotype and haplotype analyses of the five polymorphisms showed no evidence of an association with breast cancer (p>0.3). We also conducted a meta-analysis of the V660 L (rs1042838) polymorphism on our and six other published studies including 10,205 cases and 11,320 controls. Compared to the VV homozygotes, VL heterozygotes and LL homozygotes were associated with a non-significant increased risk for breast cancer [Odds ratio (OR)VL, 1.06; 95% confidence intervals (95% CI), 0.97-1.15, ORLL, 1.05; 95% CI, 0.75-1.49]. Analysis of a per-leucine allele risk under a codominant model, however, suggested a significant leucine-allele dose effect, ORper-L, 1.07; 95% CI, 1.02-1.13, p=0.01. We conclude that the leucine allele of the V660L SNP may be associated with a small increase in breast cancer risk, while the other four PGR SNPs, +44C/T (rs518162), +331G/A (rs10895068), H770H (rs1042839) and Q886Q (rs500760), do not substantially increase breast cancer risk.

High proportion of BRCA1/2 founder mutations in Hispanic breast/ovarian cancer families from Colombia

In South America, a high proportion of the population is of Hispanic origin with an important representation in Colombia. Since nothing is known about the contribution of BRCA1 and BRCA2 germline mutations to hereditary breast/ovarian cancer in the Hispanic population from Colombia, we conducted the first study of 53 breast/ovarian cancer families from this country. Comprehensive BRCA mutation screening was performed using a range of techniques, including DHPLC, SSCP, and PTT, followed by DNA sequencing analysis. Thirteen deleterious germline mutations (24.5%) were identified in 53 families, comprising eight in BRCA1 and five in BRCA2. The two recurrent BRCA1 mutations, 3450 delCAAG and A1708E, accounted for 100% of all BRCA1 mutations identified in this cohort and the recurrent 3034 delACAA BRCA2 mutation for 40% of all BRCA2 mutations. Haplotype analyses suggested that each of these mutations has arisen from a common ancestor. The prevalence of BRCA1 or BRCA2 mutations was 50% in multiple case breast cancer families, and was 33% for the breast-ovarian cancer families. Our findings show that BRCA mutations account for a substantial proportion of hereditary breast/ovarian cancer in Colombia. The spectrum of mutations differed completely to that previously reported in Hispanic families of predominantly Mexican origin from Southern California [1] suggesting that specific genetic risk assessment strategies for the different Hispanic populations in South America and in the United States need to be developed.

Histopathological features of breast cancer in carriers of ATM gene variants

Germline variants in the ataxia telangiectasia mutated (ATM) gene have been implicated in increased breast cancer risk. The aim of this study was to determine whether the histopathology of breast cancers occurring in ATM variant carriers is distinctive or resembles the described BRCA1 mutation-associated phenotype. The histopathological features of breast cancers occurring in ATM variant carriers from multiple-case breast cancer families were compared with matched controls. The test group included 21 cases of in situ and/or invasive cancer from carriers of either the IVS10-6T-->G, 2424V-->G or 1420L-->F ATM variants in the absence of BRCA1 or BRCA2 mutations. An additional four invasive cancers from carriers of a pathogenic BRCA1 mutation in the context of a familial ATM variant were also examined. The histopathology of breast cancers in ATM variant-only carriers was not significantly different from controls and known features of BRCA1 mutation-associated cancer were rarely seen. In contrast, these features were prominent in the small group of cases with a pathogenic BRCA1 mutation. Breast cancer occurring in carriers of ATM variants is not associated with distinctive histopathological features and does not resemble the tumour phenotype commonly observed in BRCA1 mutation carriers.

Characterization of the breast cancer associated ATM 7271T&gt;G (V2424G) mutation by gene expression profiling

Mutations in ATM are responsible for the autosomal recessive disorder ataxia telangiectasia. Heterozygous mutations in ATM have been associated with an elevated risk of breast cancer. We previously reported one breast cancer family in which ATM 7271T>G (V2424G) segregated with disease, and apparently acted in a dominant negative manner. We now report the screening of 782 multiple-case breast cancer families that identified two additional index cases with ATM 7271T>G. Phylogenetic sequence analysis showed that V2424 is a highly conserved residue, and that the 2424G variant is likely to interfere with function. To elucidate the consequences of this mutation, we expression profiled wild-type, heterozygous, and homozygous lymphoblastoid cell lines (LCLs) from Scottish and Australian families using an oligonucleotide microarray. Cluster analysis revealed 77 genes that were differentially expressed in homozygous and heterozygous V2424G cells (compared to wild-type) and 11 genes differentially expressed in the homozygous cells. We also evaluated the profiles of LCLs after exposure to ionizing radiation (IR) and identified 77 genes that were differentially expressed in wild-type cells, but not in homozygous or heterozygous V2424G cells. We validated the expression differences by RT-PCR in additional heterozygous V2424G LCLs from another breast cancer family. We found no consistent cytotoxicity or abrogation of ATM kinase activity after IR in seven heterozygous V2424G LCLs, compared to wild-type LCLs, but did find an increase in the number of chromosomal aberrations. These data suggest that the V2424G missense mutation acts largely as a dominant negative in terms of the associated expression profiles.

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.

Genetic and Histopathologic Evaluation ofBRCA1andBRCA2DNA Sequence Variants of Unknown Clinical Significance

Classification of rare missense variants as neutral or disease causing is a challenge and has important implications for genetic counseling. A multifactorial likelihood model for classification of unclassified variants in BRCA1 and BRCA2 has previously been developed, which uses data on co-occurrence of the unclassified variant with pathogenic mutations in the same gene, cosegregation of the unclassified variant with affected status, and Grantham analysis of the fit between the missense substitution and the evolutionary range of variation observed at its position in the protein. We have further developed this model to take into account relevant features of BRCA1- and BRCA2-associated tumors, such as the characteristic histopathology and immunochemical profiles associated with pathogenic mutations in BRCA1, and the fact that approximately 80% of tumors from BRCA1 and BRCA2 carriers undergo inactivation of the wild-type allele by loss of heterozygosity. We examined 10 BRCA1 and 15 BRCA2 unclassified variants identified in Australian, multiple-case breast cancer families. By a combination of genetic, in silico, and histopathologic analyses, we were able to classify one BRCA1 variant as pathogenic and six BRCA1 and seven BRCA2 variants as neutral. Five of these neutral variants were also found in at least 1 of 180 healthy controls, suggesting that screening a large number of appropriate controls might be a useful adjunct to other methods for evaluation of unclassified variants.

Analysis of cancer risk and BRCA1 and BRCA2mutation prevalence in the kConFab familial breast cancer resource

IntroductionThe Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer (kConFab) is a multidisciplinary, collaborative framework for the investigation of familial breast cancer. Based in Australia, the primary aim of kConFab is to facilitate high-quality research by amassing a large and comprehensive resource of epidemiological and clinical data with biospecimens from individuals at high risk of breast and/or ovarian cancer, and from their close relatives.MethodsEpidemiological, family history and lifestyle data, as well as biospecimens, are collected from multiple-case breast cancer families ascertained through family cancer clinics in Australia and New Zealand. We used the Tyrer-Cuzick algorithms to assess the prospective risk of breast cancer in women in the kConFab cohort who were unaffected with breast cancer at the time of enrolment in the study.ResultsOf kConFab's first 822 families, 518 families had multiple cases of female breast cancer alone, 239 had cases of female breast and ovarian cancer, 37 had cases of female and male breast cancer, and 14 had both ovarian cancer as well as male and female breast cancer. Data are currently held for 11,422 people and germline DNAs for 7,389. Among the 812 families with at least one germline sample collected, the mean number of germline DNA samples collected per family is nine. Of the 747 families that have undergone some form of mutation screening, 229 (31%) carry a pathogenic or splice-site mutation in BRCA1 or BRCA2. Germline DNAs and data are stored from 773 proven carriers of BRCA1 or BRCA1 mutations. kConFab's fresh tissue bank includes 253 specimens of breast or ovarian tissue – both normal and malignant – including 126 from carriers of BRCA1 or BRCA2 mutations.ConclusionThese kConFab resources are available to researchers anywhere in the world, who may apply to kConFab for biospecimens and data for use in ethically approved, peer-reviewed projects. A high calculated risk from the Tyrer-Cuzick algorithms correlated closely with the subsequent occurrence of breast cancer in BRCA1 and BRCA2 mutation positive families, but this was less evident in families in which no pathogenic BRCA1 or BRCA2 mutation has been detected.

Mutation analysis of FANCD2, BRIP1/BACH1, LMO4 and SFN in familial breast cancer

IntroductionMutations in known predisposition genes account for only about a third of all multiple-case breast cancer families. We hypothesized that germline mutations in FANCD2, BRIP1/BACH1, LMO4 and SFN may account for some of the unexplained multiple-case breast cancer families.MethodsThe families used in this study were ascertained through the Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer (kConFab). Denaturing high performance liquid chromatography (DHPLC) analysis of the coding regions of these four genes was conducted in the youngest affected cases of 30 to 267 non-BRCA1/2 breast cancer families. In addition, a further 399 index cases were also screened for mutations in two functionally significant regions of the FANCD2 gene and 253 index cases were screened for two previously reported mutations in BACH1 (p. P47A and p. M299I).ResultsDHPLC analysis of FANCD2 identified six silent exonic variants, and a large number of intronic variants, which tagged two common haplotypes. One protein truncating variant was found in BRIP1/BACH1, as well as four missense variants, a silent change and a variant in the 3' untranslated region. No missense or splice site mutations were found in LMO4 or SFN. Analysis of the missense, silent and frameshift variants of FANCD2 and BACH1 in relatives of the index cases, and in a panel of controls, found no evidence suggestive of pathogenicity.ConclusionThere is no evidence that highly penetrant exonic or splice site mutations in FANCD2, BRIP1/BACH1, LMO4 or SFN contribute to familial breast cancer. Large scale association studies will be necessary to determine whether any of the polymorphisms or haplotypes identified in these genes contributes to breast cancer risk.

Low frequency of CHEK2 1100delC allele in Australian multiple-case breast cancer families: functional analysis in heterozygous individuals

A protein-truncating variant of CHEK2, 1100delC, is associated with a moderate increase in breast cancer risk. We have determined the prevalence of this allele in index cases from 300 Australian multiple-case breast cancer families, 95% of which had been found to be negative for mutations in BRCA1 and BRCA2. Only two (0.6%) index cases heterozygous for the CHEK2 mutation were identified. All available relatives in these two families were genotyped, but there was no evidence of co-segregation between the CHEK2 variant and breast cancer. Lymphoblastoid cell lines established from a heterozygous carrier contained approximately 20% of the CHEK2 1100delC mRNA relative to wild-type CHEK2 transcript. However, no truncated CHK2 protein was detectable. Analyses of expression and phosphorylation of wild-type CHK2 suggest that the variant is likely to act by haploinsufficiency. Analysis of CDC25A degradation, a downstream target of CHK2, suggests that some compensation occurs to allow normal degradation of CDC25A. Such compensation of the 1100delC defect in CHEK2 might explain the rather low breast cancer risk associated with the CHEK2 variant, compared to that associated with truncating mutations in BRCA1 or BRCA2.

Molecular characterization and cancer risk associated withBRCA1 andBRCA2 splice site variants identified in multiple-case breast cancer families

Genetic screening of women from multiple-case breast cancer families and other research-based endeavors have identified an extensive collection of germline variations of BRCA1 and BRCA2 that can be classified as deleterious and have clinical relevance. For some variants, such as those in the conserved intronic splice site regions which are highly likely to alter splicing, it is not possible to classify them based on the identified DNA sequence variation alone. We studied 11 multiple-case breast cancer families carrying seven distinct splice site region genetic alterations in BRCA1 or BRCA2 (BRCA1, c.IVS6-2delA, c.IVS9-2A>C, c.IVS4-1G>T, c.IVS20+1G>A and BRCA2, c.IVS17-1G>C, c.IVS20+1G>A, c.IVS7-1G>A) and applied SpliceSiteFinder to predict possible changes in efficiency of splice donor and acceptor sites, characterized the transcripts, and estimated the average age-specific cumulative risk (penetrance) using a modified segregation analysis. SpliceSiteFinder predicted and we identified transcipts that illustrated that all variants caused exon skipping, and all but two led to frameshifts. The risks of breast cancer to age 70 yrs, averaged over all variants, over BRCA1 variants alone, and over BRCA2 variants alone, were 73% (95% confidence interval 47-93), 64% (95%CI 28-96) and 79% (95%CI 48-98) respectively (all P<0.0001). Therefore five of these seven consensus splice site variants of BRCA1 and BRCA2 produce a transcript similar to that of other previously described deleterious exonic variants and are associated with similar high lifetime risks.

ATM and Genome Maintenance: Defining Its Role in Breast Cancer Susceptibility

The ATM gene is mutated in ataxia-telangiectasia (A-T), a genetic instability syndrome characterized by increased cancer risk, as well as other features. Recent studies have shown that the ATM protein kinase plays a critical role in maintaining genome integrity by activating a biochemical chain reaction that in turn leads to cell cycle checkpoint activation and repair of DNA damage. ATM targets include well-known tumor suppressor genes such as p53 and BRCA1, both of which play an important role in predisposition to breast cancer. Studies of A-T families have consistently reported an increased risk of breast cancer in women with one mutated ATM gene, but so far an increased frequency of ATM mutations has not been found in women with breast cancer. Some specific missense and protein truncating variants of ATM have been reported to confer increased breast cancer risk, but the magnitude of this risk remains uncertain. A more comprehensive analysis of ATM is needed in large case-control studies, and in multiple-case breast cancer families.

Frequency of the ATM IVS10-6T-->G variant in Australian multiple-case breast cancer families.

BackgroundGermline mutations in the genes BRCA1 and BRCA2 account for only a proportion of hereditary breast cancer, suggesting that additional genes contribute to hereditary breast cancer. Recently a heterozygous variant in the ataxia–telangiectasia mutated (ATM) gene, IVS10-6T→G, was reported by an Australian multiple-case breast cancer family cohort study (the Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer) to confer a substantial breast cancer risk. Although this variant can result in a truncated ATM product, its clinical significance as a high-penetrance breast cancer allele or its role as a low-penetrance risk-modifier is controversial.MethodsWe determined the frequency of ATM IVS10-6T→G variants in a cohort of individuals affected by breast and/or ovarian cancer who underwent BRCA1 and BRCA2 genetic testing at four major Australian familial cancer clinics.ResultsSeven of 495 patients (1.4%) were heterozygous for the IVS10-6T→G variant; the carrier rate in unselected Australian women with no family history of breast cancer is reported to be 6 of 725 (0.83%) (P = 0.4). Two of the seven probands also harboured a pathogenic BRCA1 mutation and one patient had a BRCA1 unclassified variant of uncertain significance.ConclusionThese findings indicate that the ATM IVS10-6T→G variant does not seem to occur at a significantly higher frequency in affected individuals from high-risk families than in the general population. A role for this variant as a low-penetrance allele or as a modifying gene in association with other genes (such as BRCA1) remains possible. Routine testing for ATM IVS10-6T→G is not warranted in mutation screening of affected individuals from high-risk families.

Average age-specific cumulative risk of breast cancer according to type and site of germline mutations in BRCA1 and BRCA2 estimated from multiple-case breast cancer families attending Australian family cancer clinics.

If the risk of disease is not the same for all germline mutations in a given gene, or if there are other familial modifiers of risk in carriers, then family-history-based estimates of average risk for detected mutations in that gene will depend on how carriers are sampled. Risk may also depend on the site or type of mutation. We studied 51 families with strong histories of breast cancer who attended Australian family cancer clinics and in which a germline mutation in BRCA1 or BRCA2 had been identified (28 and 23 families, respectively). Breast cancer risk in carriers was estimated under maximum likelihood theory, using information from all family members including those not tested, with adjustment for ascertainment by conditioning on genotype of the proband and family phenotype. The average cumulative risk of breast cancer for mutations in either BRCA1 or BRCA2 was 27% (95% confidence interval 16-43%) to age 50 and 64% (44-83%) to age 70. When grouped, the incidence in carriers was on average 17 (10-30) times that in non-carriers, independent of gene or mutation type (hazard ratios: 11 (4-29) for BRCA1, 23 (12-43) for BRCA2 (P for difference = 0.23); 13 (6-29) for protein-truncating mutations, 30 (9-104) for missense mutations and 30 (10-90) for splice-site mutations). For missense mutations, this was equivalent to a cumulative risk to age 70 of 83% (40-100%) and was due in part, but not totally, to the missense mutations 300 T>G in BRCA1 and 4486 G>T in BRCA2, which were individually found to be associated with high risk (P<0.001). Mutations in the central region of BRCA1 may be associated with a lower risk. The issue of the pathogenicity of specific variants may be addressed analytically providing there are one or more suitably informative families with that mutation.

Dominant negative ATM mutations in breast cancer families.

The ATM gene encoding a putative protein kinase is mutated in ataxia-telangiectasia (A-T), an autosomal recessive disorder with a predisposition for cancer. Studies of A-T families suggest that female heterozygotes have an increased risk of breast cancer compared with noncarriers. However, neither linkage analyses nor mutation studies have provided supporting evidence for a role of ATM in breast cancer predisposition. Nevertheless, two recurrent ATM mutations, T7271G and IVS10-6T-->G, reportedly increase the risk of breast cancer. We examined these two ATM mutations in a population-based, case-control series of breast cancer families and multiple-case breast cancer families. Five hundred twenty-five or 262 case patients with breast cancer and 381 or 68 control subjects, respectively, were genotyped for the T7271G and IVS10-6T-->G ATM mutations, as were index patients from 76 non-BRCA1/2 multiple-case breast cancer families. Linkage and penetrance were analyzed. ATM protein expression and kinase activity were analyzed in lymphoblastoid cell lines from mutation carriers. All statistical tests were two-sided. In case and control subjects unselected for family history of breast cancer, one case patient had the T7271G mutation, and none had the IVS10-6T-->G mutation. In three multiple-case families, one of these two mutations segregated with breast cancer. The estimated average penetrance of the mutations was 60% (95% confidence interval [CI] = 32% to 90%) to age 70 years, equivalent to a 15.7-fold (95% CI = 6.4-fold to 38.0-fold) increased relative risk compared with that of the general population. Expression and activity analyses of ATM in heterozygous cell lines indicated that both mutations are dominant negative. At least two ATM mutations are associated with a sufficiently high risk of breast cancer to be found in multiple-case breast cancer families. Full mutation analysis of the ATM gene in such families could help clarify the role of ATM in breast cancer susceptibility.

Evaluation of linkage of breast cancer to the putative BRCA3 locus on chromosome 13q21 in 128 multiple case families from the Breast Cancer Linkage Consortium.

The known susceptibility genes for breast cancer, including BRCA1 and BRCA2, only account for a minority of the familial aggregation of the disease. A recent study of 77 multiple case breast cancer families from Scandinavia found evidence of linkage between the disease and polymorphic markers on chromosome 13q21. We have evaluated the contribution of this candidate "BRCA3" locus to breast cancer susceptibility in 128 high-risk breast cancer families of Western European ancestry with no identified BRCA1 or BRCA2 mutations. No evidence of linkage was found. The estimated proportion (alpha) of families linked to a susceptibility locus at D13S1308, the location estimated by Kainu et al. [(2000) Proc. Natl. Acad. Sci. USA 97, 9603-9608], was 0 (upper 95% confidence limit 0.13). Adjustment for possible bias due to selection of families on the basis of linkage evidence at BRCA2 did not materially alter this result (alpha = 0, upper 95% confidence limit 0.18). The proportion of linked families reported by Kainu et al. (0.65) is excluded with a high degree of confidence in our dataset [heterogeneity logarithm of odds (HLOD) at alpha = 0.65 was -11.0]. We conclude that, if a susceptibility gene does exist at this locus, it can only account for a small proportion of non-BRCA1/2 families with multiple cases of early-onset breast cancer.

Validation of family history of breast cancer and identification of the BRCA1 and other syndromes using a population-based cancer registry

A major risk factor for breast cancer is family history of the disease in first-degree relatives. This study evaluates the validity of family history information on breast cancer in mothers and sisters of breast cancer probands from the cancer registry (CR) compared to personal interviews (PI) of 359 consecutive population-based cases of breast cancer. Breast cancer is seen in mothers of 14% of probands by CR compared to 12% by PI. Further, 13% of probands have a sister with breast cancer using CR compared to 12% by PI. Using PI as the standard, the sensitivity of the CR family history data in mothers is 92% and the specificity is 99%, while in sisters they are 88% and 99%, respectively. These estimates were calculated on cases where family history information is available in the CR. Sensitivity and specificity are recalculated, recording an "error" whenever family history information is not available, and they are 75% and 68%, respectively, for mothers and 72% and 70%, respectively, for sisters. Estimates of proband-mother and proband-sisters familial breast cancer from CR and PI are sufficiently similar to warrant the use of CR family history data in studies of genetic epidemiology. The family phenotype consistent with the BRCA1 syndrome was found in four (1.1%) probands, all below age 50 years, while for BRCA2 there were five (1.4%) probands, three below age 50 years and two 50 years or older. Site-specific familial breast cancer was found in 23 (6.4%) probands. Population-based multiple-case breast cancer families can rapidly be identified through CR. These families can make substantial contributions to the study of genetic and environmental etiology of the disease as well as benefit from preventive and therapeutic efforts. As new knowledge and tools in molecular genetics become available, there is an urgent need for large population-based registries of families at high risk for cancer.