Hereditary breast cancer; Genetic penetrance and current status with BRCA

The most important cause of developing hereditary breast cancer is germline mutations occurring in breast cancer (BCs) susceptibility genes, for example, BRCA1, BRCA2, TP53, CHEK2, PTEN, ATM, and PPM1D. Many BC susceptibility genes can be grouped into two classes, high- and low-penetrance genes, each of which interact with multiple genes and environmental factors. However, the penetrance of genes can also be represented by a spectrum, which ranges between high and low. Two of the most common susceptibility genes are BRCA1 and BRCA2, which perform vital cellular functions for repair of homologous DNA. Loss of heterozygosity accompanied by hereditary mutations in BRCA1 or BRCA2 increases chromosomal instability and the likelihood of cancer, as well as playing a key role in stimulating malignant transformation. With regard to pathological features, familial breast cancers caused by BRCA1 mutations usually differ from those caused by BRCA2 mutations and nonfamilial BCs. It is essential to acquire an understanding of these pathological features along with the genetic history of the patient to offer an individualized treatment. Germline mutations in BRCA1 and BRCA2 genes are the main genetic and inherited factors for breast and ovarian cancer. In fact, these mutations are very important in developing early onset and increasing the risk of familial breast and ovarian cancer and responsible for 90% of hereditary BC cases. Therefore, according to the conducted studies, screening of BRCA1 and BRCA2 genes is recommended as an important marker for early detection of all patients with breast or ovarian cancer risk with family history of the disease. In this review, we summarize the role of hereditary genes, mainly BRCA1 and BRCA2, in BC.     

Genetik des Prostatakarzinoms

Prostate cancer is the most frequent malignancy in males and its etiology is strongly influenced by genetic factors. Nevertheless, no mutated genes which could be used for diagnosis have been identified in a major proportion of familial cases. Three genes with germline mutations have been identified after linkage analysis (ELAC2, RNASEL, MSR1), but these mutations are very rare and their penetrance is not well defined. The association of most genes with genetic variants is weak, and only BRCA2/familial breast cancer is of clinical relevance. As a consequence of the extreme genetic heterogeneity, diagnostic tools are not available and genetic counseling has to rely on risk estimates from pedigree data in which a single affected first degree relative indicates a relevant risk.     

Rare copy number variants observed in hereditary breast cancer cases disrupt genes in estrogen signaling and TP53 tumor suppression network

Breast cancer is the most common cancer in women in developed countries, and the contribution of genetic susceptibility to breast cancer development has been well-recognized. However, a great proportion of these hereditary predisposing factors still remain unidentified. To examine the contribution of rare copy number variants (CNVs) in breast cancer predisposition, high-resolution genome-wide scans were performed on genomic DNA of 103 BRCA1, BRCA2, and PALB2 mutation negative familial breast cancer cases and 128 geographically matched healthy female controls; for replication an independent cohort of 75 similarly mutation negative young breast cancer patients was used. All observed rare variants were confirmed by independent methods. The studied breast cancer cases showed a consistent increase in the frequency of rare CNVs when compared to controls. Furthermore, the biological networks of the disrupted genes differed between the two groups. In familial cases the observed mutations disrupted genes, which were significantly overrepresented in cellular functions related to maintenance of genomic integrity, including DNA double-strand break repair (P = 0.0211). Biological network analysis in the two independent breast cancer cohorts showed that the disrupted genes were closely related to estrogen signaling and TP53 centered tumor suppressor network. These results suggest that rare CNVs represent an alternative source of genetic variation influencing hereditary risk for breast cancer.     

Exome Sequencing of Germline DNA from Non-BRCA1/2 Familial Breast Cancer Cases Selected on the Basis of aCGH Tumor Profiling

The bulk of familial breast cancer risk (∼70%) cannot be explained by mutations in the known predisposition genes, primarily BRCA1 and BRCA2. Underlying genetic heterogeneity in these cases is the probable explanation for the failure of all attempts to identify further high-risk alleles. While exome sequencing of non-BRCA1/2 breast cancer cases is a promising strategy to detect new high-risk genes, rational approaches to the rigorous pre-selection of cases are needed to reduce heterogeneity. We selected six families in which the tumours of multiple cases showed a specific genomic profile on array comparative genomic hybridization (aCGH). Linkage analysis in these families revealed a region on chromosome 4 with a LOD score of 2.49 under homogeneity. We then analysed the germline DNA of two patients from each family using exome sequencing. Initially focusing on the linkage region, no potentially pathogenic variants could be identified in more than one family. Variants outside the linkage region were then analysed, and we detected multiple possibly pathogenic variants in genes that encode DNA integrity maintenance proteins. However, further analysis led to the rejection of all variants due to poor co-segregation or a relatively high allele frequency in a control population. We concluded that using CGH results to focus on a sub-set of families for sequencing analysis did not enable us to identify a common genetic change responsible for the aggregation of breast cancer in these families. Our data also support the emerging view that non-BRCA1/2 hereditary breast cancer families have a very heterogeneous genetic basis.     

Variants in CHEK2 other than 1100delC do not make a major contribution to breast cancer susceptibility

We recently reported that a sequence variant in the cell-cycle-checkpoint kinase CHEK2 (CHEK2 1100delC) is a low-penetrance breast cancer-susceptibility allele in noncarriers of BRCA1 or BRCA2 mutations. To investigate whether other CHEK2 variants confer susceptibility to breast cancer, we screened the full CHEK2 coding sequence in BRCA1/2-negative breast cancer cases from 89 pedigrees with three or more cases of breast cancer. We identified one novel germline variant, R117G, in two separate families. To evaluate the possible association of R117G and two germline variants reported elsewhere, R145W and I157T with breast cancer, we screened 737 BRCA1/2-negative familial breast cancer cases from 605 families, 459 BRCA1/2-positive cases from 335 families, and 723 controls from the United Kingdom, the Netherlands, and North America. All three variants were rare in all groups, and none occurred at significantly elevated frequency in familial breast cancer cases compared with controls. These results indicate that 1100delC may be the only CHEK2 allele that makes an appreciable contribution to breast cancer susceptibility.     

Targeted Sequencing of the Mitochondrial Genome of Women at High Risk of Breast Cancer without Detectable Mutations in BRCA1/2

Breast Cancer is a complex multifactorial disease for which high-penetrance mutations have been identified. Approaches used to date have identified genomic features explaining about 50% of breast cancer heritability. A number of low- to medium penetrance alleles (per-allele odds ratio < 1.5 and 4.0, respectively) have been identified, suggesting that the remaining heritability is likely to be explained by the cumulative effect of such alleles and/or by rare high-penetrance alleles. Relatively few studies have specifically explored the mitochondrial genome for variants potentially implicated in breast cancer risk. For these reasons, we propose an exploration of the variability of the mitochondrial genome in individuals diagnosed with breast cancer, having a positive breast cancer family history but testing negative for BRCA1/2 pathogenic mutations. We sequenced the mitochondrial genome of 436 index breast cancer cases from the GENESIS study. As expected, no pathogenic genomic pattern common to the 436 women included in our study was observed. The mitochondrial genes MT-ATP6 and MT-CYB were observed to carry the highest number of variants in the study. The proteins encoded by these genes are involved in the structure of the mitochondrial respiration chain, and variants in these genes may impact reactive oxygen species production contributing to carcinogenesis. More functional and epidemiological studies are needed to further investigate to what extent variants identified may influence familial breast cancer risk.     

Analysis of PALB2 Gene in BRCA1/BRCA2 Negative Spanish Hereditary Breast/Ovarian Cancer Families with Pancreatic Cancer Cases

Background

The PALB2 gene, also known as FANCN, forms a bond and co-localizes with BRCA2 in DNA repair. Germline mutations in PALB2 have been identified in approximately 1% of familial breast cancer and 3–4% of familial pancreatic cancer. The goal of this study was to determine the prevalence of PALB2 mutations in a population of BRCA1/BRCA2 negative breast cancer patients selected from either a personal or family history of pancreatic cancer.

Methods

132 non-BRCA1/BRCA2 breast/ovarian cancer families with at least one pancreatic cancer case were included in the study. PALB2 mutational analysis was performed by direct sequencing of all coding exons and intron/exon boundaries, as well as multiplex ligation-dependent probe amplification.

Results

Two PALB2 truncating mutations, the c.1653T>A (p.Tyr551Stop) previously reported, and c.3362del (p.Gly1121ValfsX3) which is a novel frameshift mutation, were identified. Moreover, several PALB2 variants were detected; some of them were predicted as pathological by bioinformatic analysis. Considering truncating mutations, the prevalence rate of our population of BRCA1/2-negative breast cancer patients with pancreatic cancer is 1.5%.

Conclusions

The prevalence rate of PALB2 mutations in non-BRCA1/BRCA2 breast/ovarian cancer families, selected from either a personal or family pancreatic cancer history, is similar to that previously described for unselected breast/ovarian cancer families. Future research directed towards identifying other gene(s) involved in the development of breast/pancreatic cancer families is required.     

Hereditary breast cancer: new genetic developments,new therapeutic avenues

Six genes confer a high risk for developing breast cancer (BRCA1/2, TP53, PTEN, STK11, CDH1). Both BRCA1 and BRCA2 have DNA repair functions, and BRCA1/2 deficient tumors are now being targeted by poly(ADP-ribose) polymerase inhibitors. Other genes conferring an increased risk for breast cancer include ATM, CHEK2, PALB2, BRIP1 and genome-wide association studies have identified lower penetrance alleles including FGFR2, a minor allele of which is associated with breast cancer. We review recent findings related to the function of some of these genes, and discuss how they can be targeted by various drugs. Gaining deeper insights in breast cancer susceptibility will improve our ability to identify those families at increased risk and permit the development of new and more specific therapeutic approaches.     

RAD51 and Breast Cancer Susceptibility: No Evidence for Rare Variant Association in the Breast Cancer Family Registry Study

Background

Although inherited breast cancer has been associated with germline mutations in genes that are functionally involved in the DNA homologous recombination repair (HRR) pathway, including BRCA1, BRCA2, TP53, ATM, BRIP1, CHEK2 and PALB2, about 70% of breast cancer heritability remains unexplained. Because of their critical functions in maintaining genome integrity and already well-established associations with breast cancer susceptibility, it is likely that additional genes involved in the HRR pathway harbor sequence variants associated with increased risk of breast cancer. RAD51 plays a central biological function in DNA repair and despite the fact that rare, likely dysfunctional variants in three of its five paralogs, RAD51C, RAD51D, and XRCC2, have been associated with breast and/or ovarian cancer risk, no population-based case-control mutation screening data are available for the RAD51 gene. We thus postulated that RAD51 could harbor rare germline mutations that confer increased risk of breast cancer.

Methodology/Principal Findings

We screened the coding exons and proximal splice junction regions of the gene for germline sequence variation in 1,330 early-onset breast cancer cases and 1,123 controls from the Breast Cancer Family Registry, using the same population-based sampling and analytical strategy that we developed for assessment of rare sequence variants in ATM and CHEK2. In total, 12 distinct very rare or private variants were characterized in RAD51, with 10 cases (0.75%) and 9 controls (0.80%) carrying such a variant. Variants were either likely neutral missense substitutions (3), silent substitutions (4) or non-coding substitutions (5) that were predicted to have little effect on efficiency of the splicing machinery.

Conclusion

Altogether, our data suggest that RAD51 tolerates so little dysfunctional sequence variation that rare variants in the gene contribute little, if anything, to breast cancer susceptibility.     

The Validation and Clinical Implementation of BRCAplus: A Comprehensive High-Risk Breast Cancer Diagnostic Assay

Breast cancer is the most commonly diagnosed cancer in women, with 10% of disease attributed to hereditary factors. Although BRCA1 and BRCA2 account for a high percentage of hereditary cases, there are more than 25 susceptibility genes that differentially impact the risk for breast cancer. Traditionally, germline testing for breast cancer was performed by Sanger dideoxy terminator sequencing in a reflexive manner, beginning with BRCA1 and BRCA2. The introduction of next-generation sequencing (NGS) has enabled the simultaneous testing of all genes implicated in breast cancer resulting in diagnostic labs offering large, comprehensive gene panels. However, some physicians prefer to only test for those genes in which established surveillance and treatment protocol exists. The NGS based BRCAplus test utilizes a custom tiled PCR based target enrichment design and bioinformatics pipeline coupled with array comparative genomic hybridization (aCGH) to identify mutations in the six high-risk genes: BRCA1, BRCA2, PTEN, TP53, CDH1, and STK11. Validation of the assay with 250 previously characterized samples resulted in 100% detection of 3,025 known variants and analytical specificity of 99.99%. Analysis of the clinical performance of the first 3,000 BRCAplus samples referred for testing revealed an average coverage greater than 9,000X per target base pair resulting in excellent specificity and the sensitivity to detect low level mosaicism and allele-drop out. The unique design of the assay enabled the detection of pathogenic mutations missed by previous testing. With the abundance of NGS diagnostic tests being released, it is essential that clinicians understand the advantages and limitations of different test designs.     

Genetic variants associated with breast cancer risk for Ashkenazi Jewish women with strong family histories but no identifiable BRCA1/2 mutation

The ability to establish genetic risk models is critical for early identification and optimal treatment of breast cancer. For such a model to gain clinical utility, more variants must be identified beyond those discovered in previous genome-wide association studies (GWAS). This is especially true for women at high risk because of family history, but without BRCA1/2 mutations. This study incorporates three datasets in a GWAS analysis of women with Ashkenazi Jewish (AJ) homogeneous ancestry. Two independent discovery cohorts comprised 239 and 238 AJ women with invasive breast cancer or preinvasive ductal carcinoma in situ and strong family histories of breast cancer, but lacking the three BRCA1/2 founder mutations, along with 294 and 230 AJ controls, respectively. An independent, third cohort of 203 AJ cases with familial breast cancer history and 263 healthy controls of AJ women was used for validation. A total of 19 SNPs were identified as associated with familial breast cancer risk in AJ women. Among these SNPs, 13 were identified from a panel of 109 discovery SNPs, including an FGFR2 haplotype. In addition, six previously identified breast cancer GWAS SNPs were confirmed in this population. Seven of the 19 markers were significant in a multivariate predictive model of familial breast cancer in AJ women, three novel SNPs [rs17663555(5q13.2), rs566164(6q21), and rs11075884(16q22.2)], the FGFR2 haplotype, and three previously published SNPs [rs13387042(2q35), rs2046210(ESR1), and rs3112612(TOX3)], yielding moderate predictive power with an area under the curve (AUC) of the ROC (receiver-operator characteristic curve) of 0.74. Population-specific genetic variants in addition to variants shared with populations of European ancestry may improve breast cancer risk prediction among AJ women from high-risk families without founder BRCA1/2 mutations.     

BRCA1 and BRCA2 missense variants of high and low clinical significance influence lymphoblastoid cell line post-irradiation gene expression

The functional consequences of missense variants in disease genes are difficult to predict. We assessed if gene expression profiles could distinguish between BRCA1 or BRCA2 pathogenic truncating and missense mutation carriers and familial breast cancer cases whose disease was not attributable to BRCA1 or BRCA2 mutations (BRCAX cases). 72 cell lines from affected women in high-risk breast ovarian families were assayed after exposure to ionising irradiation, including 23 BRCA1 carriers, 22 BRCA2 carriers, and 27 BRCAX individuals. A subset of 10 BRCAX individuals carried rare BRCA1/2 sequence variants considered to be of low clinical significance (LCS). BRCA1 and BRCA2 mutation carriers had similar expression profiles, with some subclustering of missense mutation carriers. The majority of BRCAX individuals formed a distinct cluster, but BRCAX individuals with LCS variants had expression profiles similar to BRCA1/2 mutation carriers. Gaussian Process Classifier predicted BRCA1, BRCA2 and BRCAX status, with a maximum of 62% accuracy, and prediction accuracy decreased with inclusion of BRCAX samples carrying an LCS variant, and inclusion of pathogenic missense carriers. Similarly, prediction of mutation status with gene lists derived using Support Vector Machines was good for BRCAX samples without an LCS variant (82–94%), poor for BRCAX with an LCS (40–50%), and improved for pathogenic BRCA1/2 mutation carriers when the gene list used for prediction was appropriate to mutation effect being tested (71–100%). This study indicates that mutation effect, and presence of rare variants possibly associated with a low risk of cancer, must be considered in the development of array-based assays of variant pathogenicity.     

Analysis of 17β-hydroxysteroid dehydrogenase types 5, 7, and 12 genetic sequence variants in breast cancer cases from French Canadian Families with high risk of breast and ovarian cancer

A family history and estrogen exposure are well-known risk factors for breast cancer. Members of the 17β-hydroxysteroid dehydrogenase family are responsible for important steps in the metabolism of androgens and estrogens in peripheral tissues, including the mammary gland. The crucial biological function of 17β-HSDs renders these genes good candidates for being involved in breast cancer etiology. This study screened for mutations in HSD17B7 and HSD17B12 genes, which encode enzymes involved in estradiol biosynthesis and in AKR1C3, which codes for 17β-HSD type 5 enzyme involved in androgen and progesterone metabolism, to assess whether high penetrance allelic variants in these genes could be involved in breast cancer susceptibility. Mutation screening of 50 breast cancer cases from non-BRCA1/2 high-risk French Canadian families failed to identify germline likely high-risk mutations in HSD17B7, HSD17B12 and AKR1C3 genes. However, 107 sequence variants were identified, including seven missense variants. Assessment of the impact of missense variants on enzymatic activity of the corresponding enzymes revealed no difference in catalytic properties between variants of 17β-HSD types 7 and 12 and wild-type enzymes, while variants p.Glu77Gly and p.Lys183Arg in 17β-HSD type 5 showed a slightly decreased activity. Finally, a haplotype-based approach was used to determine tagging SNPs providing valuable information for studies investigating associations of common variants in these genes with breast cancer risk.     

Germline Variation in Cancer-Susceptibility Genes in a Healthy,Ancestrally Diverse Cohort: Implications for Individual Genome Sequencing

Technological advances coupled with decreasing costs are bringing whole genome and whole exome sequencing closer to routine clinical use. One of the hurdles to clinical implementation is the high number of variants of unknown significance. For cancer-susceptibility genes, the difficulty in interpreting the clinical relevance of the genomic variants is compounded by the fact that most of what is known about these variants comes from the study of highly selected populations, such as cancer patients or individuals with a family history of cancer. The genetic variation in known cancer-susceptibility genes in the general population has not been well characterized to date. To address this gap, we profiled the nonsynonymous genomic variation in 158 genes causally implicated in carcinogenesis using high-quality whole genome sequences from an ancestrally diverse cohort of 681 healthy individuals. We found that all individuals carry multiple variants that may impact cancer susceptibility, with an average of 68 variants per individual. Of the 2,688 allelic variants identified within the cohort, most are very rare, with 75% found in only 1 or 2 individuals in our population. Allele frequencies vary between ancestral groups, and there are 21 variants for which the minor allele in one population is the major allele in another. Detailed analysis of a selected subset of 5 clinically important cancer genes, BRCA1, BRCA2, KRAS, TP53, and PTEN, highlights differences between germline variants and reported somatic mutations. The dataset can serve a resource of genetic variation in cancer-susceptibility genes in 6 ancestry groups, an important foundation for the interpretation of cancer risk from personal genome sequences.     

Genome-wide Linkage Scan Reveals Three Putative Breast-Cancer-Susceptibility Loci

Despite all the research efforts made during the last few decades, most of the cases of families with breast cancer remain unexplained. Mutations in BRCA1 and BRCA2, and in other breast-cancer-susceptibility genes, account for about 25% of familial breast cancer. Linkage studies have failed to identify other breast-cancer-susceptibility genes. The selection criteria of the families, differences in the population background, or clinical and genetic heterogeneity, among other factors, might determine the power to detect the linkage signal. We have performed a SNP-based linkage scan with a total of 6000 SNP markers across the genome in 41 breast-cancer Spanish families, with an average of four breast-cancer cases per family not associated with BRCA1 or BRCA2 germline mutations. In addition, we have included three BRCA-positive families to test the power in linkage detection from a low-complexity family in which a high-penetrance mutation segregates. We have identified three regions of interest, located on 3q25, 6q24, and 21q22. The two former regions showed a suggestive linkage signal (HLOD scores 3.01 and 2.26, respectively), and the latter region showed a significant linkage signal (HLOD score 3.55). Moreover, we found that a subset of 13 families with bilateral breast cancer presented a HLOD of 3.13 on the 3q25 region. Our results suggest that several variables must be taken into account before performing a linkage study in familial breast cancer because of the high heterogeneity within non-BRCA1/2 families. Phenotypic and geographic homogeneity could be the most important factors.     

Eleven Candidate Susceptibility Genes for Common Familial Colorectal Cancer

Hereditary factors are presumed to play a role in one third of colorectal cancer (CRC) cases. However, in the majority of familial CRC cases the genetic basis of predisposition remains unexplained. This is particularly true for families with few affected individuals. To identify susceptibility genes for this common phenotype, we examined familial cases derived from a consecutive series of 1514 Finnish CRC patients. Ninety-six familial CRC patients with no previous diagnosis of a hereditary CRC syndrome were included in the analysis. Eighty-six patients had one affected first-degree relative, and ten patients had two or more. Exome sequencing was utilized to search for genes harboring putative loss-of-function variants, because such alterations are likely candidates for disease-causing mutations. Eleven genes with rare truncating variants in two or three familial CRC cases were identified: UACA, SFXN4, TWSG1, PSPH, NUDT7, ZNF490, PRSS37, CCDC18, PRADC1, MRPL3, and AKR1C4. Loss of heterozygosity was examined in all respective cancer samples, and was detected in seven occasions involving four of the candidate genes. In all seven occasions the wild-type allele was lost (P = 0.0078) providing additional evidence that these eleven genes are likely to include true culprits. The study provides a set of candidate predisposition genes which may explain a subset of common familial CRC. Additional genetic validation in other populations is required to provide firm evidence for causality, as well as to characterize the natural history of the respective phenotypes.     

Prevalence of BRCA1 Mutations in Familial and Sporadic Greek Ovarian Cancer Cases

Germline mutations in the BRCA1 and BRCA2 genes contribute to approximately 18% of hereditary ovarian cancers conferring an estimated lifetime risk from 15% to 50%. A variable incidence of mutations has been reported for these genes in ovarian cancer cases from different populations. In Greece, six mutations in BRCA1 account for 63% of all mutations detected in both BRCA1 and BRCA2 genes. This study aimed to determine the prevalence of BRCA1 mutations in a Greek cohort of 106 familial ovarian cancer patients that had strong family history or metachronous breast cancer and 592 sporadic ovarian cancer cases. All 698 patients were screened for the six recurrent Greek mutations (including founder mutations c.5266dupC, p.G1738R and the three large deletions of exon 20, exons 23–24 and exon 24). In familial cases, the BRCA1 gene was consequently screened for exons 5, 11, 12, 20, 21, 22, 23, 24. A deleterious BRCA1 mutation was found in 43/106 (40.6%) of familial cancer cases and in 27/592 (4.6%) of sporadic cases. The variant of unknown clinical significance p.V1833M was identified in 9/698 patients (1.3%). The majority of BRCA1 carriers (71.2%) presented a high-grade serous phenotype. Identifying a mutation in the BRCA1 gene among breast and/or ovarian cancer families is important, as it enables carriers to take preventive measures. All ovarian cancer patients with a serous phenotype should be considered for genetic testing. Further studies are warranted to determine the prevalence of mutations in the rest of the BRCA1 gene, in the BRCA2 gene, and other novel predisposing genes for breast and ovarian cancer.