BRCA1 mutations in African Americans

The breast cancer predisposing gene, BRCA1, was analyzed for germline mutations in 45 African American families at high-risk for hereditary breast cancer. Patients were considered high-risk if they had a family history of the disease, early onset breast cancer, bilateral breast cancer, or breast and ovarian cancer. The entire BRCA1 coding and flanking intron regions have been examined by single stranded conformation polymorphism analysis followed by sequencing of variant bands. Eleven different BRCA1 germline mutations/variations were identified in 7 patients from the 45 high-risk families. Two pathogenic, protein-truncating mutations were detected in exon 11. A ten base pair tandem duplication, 943ins10, was present in a woman with breast and ovarian cancer whose first-degree relatives had prostate cancer. A four base pair deletion, 3450del4, was detected in a breast cancer patient with five cases of breast cancer in the family; two of the proband's sisters with breast cancer also carried the same mutation. Four amino acid substitutions (Lys1183Arg, Leu1564Pro, Gln1785His, and Glu1794Asp) and four nucleotide substitutions in intron 22 (IVS22+78 C/A, IVS22+67 T/C, IVS22+8 T/A and IVS22+7 T/C) were observed in patients and not in control subjects. One early onset breast cancer patient carried five distinct BRCA1 variations, two amino acid substitutions and three substitutions in intron 22. An amino acid substitution in exon 11, Ser1140Gly, was identified in 3 different unrelated patients and in 6 of 92 control samples. The latter probably represents a benign polymorphism. Electronic Publication     

Contribution of BRCA1 germline mutation in patients with sporadic breast cancer

Hereditary artifacts in BRCA1 gene have a significant contributory role in familial cases of breast cancer. However, its germline mutational penetrance in sporadic breast cancer cases with respect to Pakistani population has not yet been very well defined. This study was designed to assess the contributory role of germline mutations of this gene in sporadic cases of breast cancer. 150 cases of unilateral breast cancer patients, with no prior family history of breast cancer and no other disorders or diseases in general with age range 35–75 yrs, were included in this study.Mutational analysis for hot spots on Exon 2, 3 and 13 of BRCA1 was done by using Single Strand Conformational Polymorphism (SSCP). Sequence analysis revealed five variants (missense) and one novel splice site mutation at exon 13. No germline mutation was observed on the remaining exons with respect sporadic breast cancer cases in Pakistani population. A vast majority of breast cancer cases are sporadic; the present study may be helpful for designing a better genetic screening tool for germline BRCA mutations in sporadic breast cancer patients of Pakistani population. Further studies involving a screening of entire coding region of BRCA1 is required to explore the merits of genetic diagnosis and counseling in breast cancer patients.     

The genetic basis of Cowden’s syndrome: three novel mutations in PTEN/MMAC1/TEP1

Cowden’s syndrome (CS) is an autosomal dominant disorder associated with an increased risk of developing benign and malignant tumors in a variety of tissues, including the skin, thyroid, breast and brain. Women with CS are felt to have an increased risk of developing breast cancer, and virtually all women with CS develop bilateral fibrocystic disease of the breast. Recently, a series of germline mutations have been identified from CS families in a gene known as PTEN/MMAC1/TEP1. In this study, we used heteroduplex analysis and direct sequencing analysis and identified three novel germline mutations in the PTEN/MMAC1/TEP1 coding sequence from unrelated individuals with CS. We report a de novo transition (T→C) at nucleotide 335 in exon 5. This missense mutation resulted in a leucine to proline (CTA to CCA) change at codon 112. We also describe a novel splice site mutation (801+2T→G) in intron 7 that caused exon skipping in PTEN/MMAC1/TEP1 mRNA. The third mutation we report is a missense mutation, consisting of a transition (T→C) at nucleotide 202 in exon 3, resulting in a tyrosine to histidine (TAC to CAC) change at codon 68. Finally, we also detected a rare polymorphism in exon 7 of the PTEN/MMAC1/TEP1 coding sequence. These data confirm the observation that mutations of the PTEN/MMAC1/ TEP1 coding sequence are responsible for at least some cases of CS, and further define the spectrum of mutations in this autosomal dominant disorder. Received: 21 October 1997 / Accepted: 15 December 1997     

Gain- and Loss-of-Function Mutations in the Breast Cancer Gene GATA3 Result in Differential Drug Sensitivity

Patterns of somatic mutations in cancer genes provide information about their functional role in tumourigenesis, and thus indicate their potential for therapeutic exploitation. Yet, the classical distinction between oncogene and tumour suppressor may not always apply. For instance, TP53 has been simultaneously associated with tumour suppressing and promoting activities. Here, we uncover a similar phenomenon for GATA3, a frequently mutated, yet poorly understood, breast cancer gene. We identify two functional classes of frameshift mutations that are associated with distinct expression profiles in tumours, differential disease-free patient survival and gain- and loss-of-function activities in a cell line model. Furthermore, we find an estrogen receptor-independent synthetic lethal interaction between a GATA3 frameshift mutant with an extended C-terminus and the histone methyltransferases G9A and GLP, indicating perturbed epigenetic regulation. Our findings reveal important insights into mutant GATA3 function and breast cancer, provide the first potential therapeutic strategy and suggest that dual tumour suppressive and oncogenic activities are more widespread than previously appreciated.     

Significance of PIK3CA Mutations in Patients with Early Breast Cancer Treated with Adjuvant Chemotherapy: A Hellenic Cooperative Oncology Group (HeCOG) Study

Background

The PI3K-AKT pathway is frequently activated in breast cancer. PIK3CA mutations are most frequently found in the helical (exon 9) and kinase (exon 20) domains of this protein. The aim of the present study was to examine the role of different types of PIK3CA mutations in combination with molecular biomarkers related to PI3K-AKT signaling in patients with early breast cancer.

Methods

Tumor tissue samples from 1008 early breast cancer patients treated with adjuvant chemotherapy in two similar randomized trials of HeCOG were examined. Tumors were subtyped with immunohistochemistry (IHC) and FISH for ER, PgR, Ki67, HER2 and androgen receptor (AR). PIK3CA mutations were analyzed by Sanger sequencing (exon 20) and qPCR (exon 9) (Sanger/qPCR mutations). In 610 cases, next generation sequencing (NGS) PIK3CA mutation data were also available. PIK3CA mutations and PTEN protein expression (IHC) were analyzed in luminal tumors (ER and/or PgR positive), molecular apocrine carcinomas (MAC; ER/PgR negative / AR positive) and hormone receptor (ER/PgR/AR) negative tumors.

Results

PIK3CA mutations were detected in 235/1008 tumors (23%) with Sanger/qPCR and in 149/610 tumors (24%) with NGS. Concordance between the two methods was good with a Kappa coefficient of 0.76 (95% CI 0.69–0.82). Lobular histology, low tumor grade and luminal A tumors were associated with helical domain mutations (PIK3CAhel), while luminal B with kinase domain mutations (PIK3CAkin). The overall incidence of PIK3CA mutations was higher in luminal as compared to MAC and hormone receptor negative tumors (p = 0.004). Disease-free and overall survival did not significantly differ with respect to PIK3CA mutation presence and type. However, a statistically significant interaction between PIK3CA mutation status and PTEN low protein expression with regard to prognosis was identified.

Conclusions

The present study did not show any prognostic significance of specific PIK3CA mutations in a large group of predominantly lymph-node positive breast cancer women treated with adjuvant chemotherapy. Further analyses in larger cohorts are warranted to investigate possible differential effect of distinct PIK3CA mutations in small subgroups of patients.     

WT1 mutations in patients with Denys-Drash syndrome: a novel mutation in exon 8 and paternal allele origin

Denys-Drash syndrome (DDS) is characterized by early onset nephropathy, pseudohermaphroditism in males and a high risk for developing Wilms' tumour (WT). The exact cause of DDS is unknown but germline mutations in the Wilms' tumour suppressor gene (WT1) have recently been described in the majority of DDS patients studied. These mutations occur de novo and are clustered around the zinc finger (ZF) coding exons of the WT1 gene. Analysis of exons 2–10 of the WT1 gene in constitutional DNA from five patients with DDS was carried out using the polymerase chain reaction (PCR) and direct DNA sequencing. In four out of the five patients, heterozygous germline mutations were found: a novel point mutation in exon 8 (ZF₂) at codon 377 altering the wild-type histidine to arginine, and three previously described point mutations in exon 9 (ZF₃) in the codons corresponding to amino acids ³⁹⁴Arg and ³⁹⁶Asp. In one patient, no mutations could be demonstrated. In three patients where parental DNA was available, the mutations were shown to have occurred de novo. Furthermore, since tumour DNA in two of these cases had lost the wild-type allele, polymorphic markers from the short arm of chromosome 11 were used to determine the parental origin of the mutant chromosome. In both cases, the mutant chromosome was shown to be of paternal origin. Since the majority of published WT1 mutations in DDS patients alter a RsrII restriction site in exon 9, we were able to perform PCR-based diagnosis in a female patient with early renal insufficiency and normal external genitalia.     

Genomic rearrangements of the APC tumor-suppressor gene in familial adenomatous polyposis

Germline mutations of the adenomatous polyposis coli (APC) tumor-suppressor gene result in the hereditary colorectal cancer syndrome familial adenomatous polyposis (FAP). Almost all APC mutations that have been identified are single-nucleotide alterations, small insertions, or small deletions that would truncate the protein product of the gene. No well-characterized intragenic rearrangement of APC has been described, and the prevalence of this type of mutation in FAP patients is not clear. We screened 49 potential FAP families and identified 26 different germline APC mutations in 30 families. Four of these mutations were genomic rearrangements resulting from homologous and nonhomologous recombinations mediated by Alu elements. Two of these four rearrangements were complex, involving deletion and insertion of nucleotides. Of these four rearrangements, one resulted in the deletion of exons 11 and 12 and two others resulted in either complete or partial deletion of exon 14. The fourth rearrangement grossly altered the sequence within intron 14. Although this rearrangement did not affect any coding sequence of APC at the genomic DNA level, it caused inappropriate splicing of exon 14. These rearrangements were initially revealed by analyzing cDNAs and could not have been identified by using mutation detection methods that screened each exon individually. The identification of a rearrangement that did not alter any coding exons yet affected the splicing further underscores the importance of using cDNA for mutation analysis. The identification of four genomic rearrangements among 30 mutations suggests that genomic rearrangements are frequent germline APC mutations.     

Different familial adenomatous polyposis phenotypes resulting from deletions of the entire APC exon 15

Germline mutations of the adenomatous polyposis coli ( APC) gene cause familial adenomatous polyposis (FAP), an autosomal, dominantly inherited disease that predisposes patients to colorectal cancer. The APC gene is composed of 15 coding exons and encodes an open reading frame of 8.5 kb. The 3' 6.5 kb of the APCopen reading frame is encoded by a single exon, exon 15. Most identified APC mutations are at the 5' half of the APC open reading frame and are nucleotide substitutions and small deletions or insertions that result in truncation of the APC protein. Very few well-characterized gross alterations of APC have been reported. Patients with FAP typically develop hundreds to thousands of colorectal tumors beginning in their adolescence. A subgroup of patients with FAP who develop fewer tumors at an older age have what is called attenuated FAP (AFAP). Accumulating evidence indicates that patients carrying germline APC mutations in the first four coding exons, in the alternatively spliced region of exon 9, or in the 3' half of the coding region usually develop AFAP. We characterized two germline APC alterations that deleted the entire APC exon 15 as the result of 56-kb and 73-kb deletions at the APC locus. A surprising finding was that one proband had the typical FAP phenotype, whereas the other had a phenotype consistent with that of AFAP.     

Novel mutations of endothelin-B receptor gene in Pakistani patients with Waardenburg syndrome

Mutations in EDNRB gene have been reported to cause Waardenburg-Shah syndrome (WS4) in humans. We investigated 17 patients with WS4 for identification of mutations in EDNRB gene using PCR and direct sequencing technique. Four genomic mutations were detected in four patients; a G to C transversion in codon 335 (S335C) in exon 5 and a transition of T to C in codon (S361L) in exon 5, a transition of A to G in codon 277 (L277L) in exon 4, a non coding transversion of T to A at −30 nucleotide position of exon 5. None of these mutations were found in controls. One of the patients harbored two novel mutations (S335C, S361L) in exon 5 and one in Intronic region (−30exon5 A>G). All of the mutations were homozygous and novel except the mutation observed in exon 4. In this study, we have identified 3 novel mutations in EDNRB gene associated with WS4 in Pakistani patients.     

BRCA1 mutations and polymorphisms in a hospital-based consecutive series of breast cancer patients from Apulia, Italy

INTRODUCTION: Hereditary breast cancer has been partly attributed to germline mutations in the BRCA1 gene that are deleterious for BRCA1 protein activity. This paper analyzes the incidence and characteristics of detectable BRCA1 mutations and polymorphisms in a hospital-based consecutive series of breast cancer patients from southern Italy to investigate the incidence and the association of these molecular alterations with breast cancer biology and family history. METHODS: One hundred cases with familial characteristics were selected from a consecutive series of 511 patients with a first diagnosis of breast cancer. DNA from peripheral blood was screened for whole BRCA1 gene mutations utilizing dHPLC as a pre-screening analysis and automatic DNA sequencing for the identification of specific alterations. RESULTS: In the overall series of 511 patients, 100 had a family history of breast cancer and were investigated for BRCA1 mutations. Two types of BRCA1 mutations were identified, 5382insC in six cases and 4566delA in one case. The 5382insC mutation was present in two out of six cases with ovarian cancer while 4566delA in one case of male cancer. The most frequent missense polymorphisms were E1038G, P871L, K1183R in exon 11, S1613G, M1652I in exon 16 and D1778G in exon 22. Confirming what found in previous studies, patients in whom pathological BRCA1 mutations were detected had early-onset breast cancer (p=0.05), positive nodal status (p=0.05), lower ER (p=0.02) and PgR (p=0.01) content. Interestingly, the K1183R polymorphism and, less strongly, S1613G polymorphism were associated to mutational risk (K1183R: OR 0.1 p=0.03; S1613G: OR 2.7 p=0.08). CONCLUSION: Mutations in the BRCA1 gene are frequent also in our consecutive series of patients from southern Italy. An association between two detected single nucleotide polymorphisms (SNPs) and BRCA1 mutational risk was ascertained. Finally, we confirm the fact that peculiar clinical-pathological features seem to characterize patients with a family history of breast cancer and BRCA1 alterations.     

Identification of a Comprehensive Spectrum of Genetic Factors for Hereditary Breast Cancer in a Chinese Population by Next-Generation Sequencing

The genetic etiology of hereditary breast cancer has not been fully elucidated. Although germline mutations of high-penetrance genes such as BRCA1/2 are implicated in development of hereditary breast cancers, at least half of all breast cancer families are not linked to these genes. To identify a comprehensive spectrum of genetic factors for hereditary breast cancer in a Chinese population, we performed an analysis of germline mutations in 2,165 coding exons of 152 genes associated with hereditary cancer using next-generation sequencing (NGS) in 99 breast cancer patients from families of cancer patients regardless of cancer types. Forty-two deleterious germline mutations were identified in 21 genes of 34 patients, including 18 (18.2%) BRCA1 or BRCA2 mutations, 3 (3%) TP53 mutations, 5 (5.1%) DNA mismatch repair gene mutations, 1 (1%) CDH1 mutation, 6 (6.1%) Fanconi anemia pathway gene mutations, and 9 (9.1%) mutations in other genes. Of seven patients who carried mutations in more than one gene, 4 were BRCA1/2 mutation carriers, and their average onset age was much younger than patients with only BRCA1/2 mutations. Almost all identified high-penetrance gene mutations in those families fulfill the typical phenotypes of hereditary cancer syndromes listed in the National Comprehensive Cancer Network (NCCN) guidelines, except two TP53 and three mismatch repair gene mutations. Furthermore, functional studies of MSH3 germline mutations confirmed the association between MSH3 mutation and tumorigenesis, and segregation analysis suggested antagonism between BRCA1 and MSH3. We also identified a lot of low-penetrance gene mutations. Although the clinical significance of those newly identified low-penetrance gene mutations has not been fully appreciated yet, these new findings do provide valuable epidemiological information for the future studies. Together, these findings highlight the importance of genetic testing based on NCCN guidelines and a multi-gene analysis using NGS may be a supplement to traditional genetic counseling.     

Expression profiles of BRCA1 splice variants in asynchronous and in G1/S synchronized tumor cell lines

Disrupting the function of the BRCA1 gene by mechanisms other than germline mutations is suspected to occur in cases of sporadic breast/ovarian cancers. Using ribonuclease protection assay and multiplex RT-PCR, we examined the change of the total BRCA1 mRNA pool and the expression profile of four predominant BRCA1 splice variants in asynchronous and in G1/S synchronized tumor cell populations compared to normal breast cells. Experiments were carried out on MCF-7 and MDA-MB-231 breast cancer, OVCAR-5 ovarian cancer, and K562 leukemia cell lines. The ratio of the full length, the delta(11q), the delta(9,10), and the delta(9,10,11q) BRCA1 isoforms showed different expression patterns in the examined breast and ovarian tumor cell lines as compared to the leukemia cell line. This observation raises the possibility that the dysregulation of alternative splicing of the BRCA1 gene could be involved in tumor formation in the breast and the ovary, even in the absence of germline mutations.     

Inherited<Emphasis Type="Italic"> BRCA2</Emphasis> mutations in African Americans with breast and/or ovarian cancer: a study of familial and early onset cases

In order to identify the spectrum of BRCA2 mutations in African Americans, breast or ovarian cancer patients from 74 independent families at elevated risk of germline mutations were investigated. The entire coding regions and flanking introns of BRCA2 were screened for germline mutations by single-stranded conformation polymorphism, protein truncation test, or denaturing high performance liquid chromatography followed by DNA sequencing. Eight distinct protein-truncating mutations were detected in six female patients (average age of onset of breast cancer: 37.6 years) and two male patients, but not in 163 unrelated disease-free controls. Two (1993delAA, 8643delAT) of the eight pathogenic mutations observed in African Americans have not been previously described. The other six pathogenic mutations (1882delT, 1991delATAA, 2001delTTAT, 2816insA, 4075delGT, 4088delA) have been detected in Caucasians; only the 2816insA mutation has been reported previously in African Americans. There were no significant differences in the frequency of deleterious BRCA2 mutations in African Americans compared with Caucasians. Six rare variations, not previously reported, were identified in five breast cancer patients but not in 163 disease-free control subjects. Of 11 different polymorphisms identified in high-risk African-American breast cancer patients, four may be unique to African Americans. An intron 10 polymorphism observed in patients was not detected in 163 disease-free African-American control subjects; this difference is statistically significant. Since many different pathogenic mutations and variants of unknown significance are observed in African Americans, BRCA2 genetic testing in high-risk African-American families must include the entire coding and flanking non-coding regions of the gene.     

Low Prevalence of CHEK2 Gene Mutations in Multiethnic Cohorts of Breast Cancer Patients in Malaysia

CHEK2 is a protein kinase that is involved in cell-cycle checkpoint control after DNA damage. Germline mutations in CHEK2 gene have been associated with increase in breast cancer risk. The aim of this study is to identify the CHEK2 gene germline mutations among high-risk breast cancer patients and its contribution to the multiethnic population in Malaysia. We screened the entire coding region of CHEK2 gene on 59 high-risk breast cancer patients who tested negative for BRCA1/2 germline mutations from UKM Medical Centre (UKMMC), Hospital Kuala Lumpur (HKL) and Hospital Putrajaya (HPJ). Sequence variants identified were screened further in case-control cohorts consisting of 878 unselected invasive breast cancer patients (180 Malays, 526 Chinese and 172 Indian) and 270 healthy individuals (90 Malays, 90 Chinese and 90 Indian). By screening the entire coding region of the CHEK2 gene, two missense mutations, c.480A>G (p.I160M) and c.538C>T (p.R180C) were identified in two unrelated patients (3.4%). Further screening of these missense mutations on the case-control cohorts unveiled the variant p.I160M in 2/172 (1.1%) Indian cases and 1/90 (1.1%) Indian control, variant p.R180C in 2/526 (0.38%) Chinese cases and 0/90 Chinese control, and in 2/180 (1.1%) of Malay cases and 1/90 (1.1%) of Malay control. The results of this study suggest that CHEK2 mutations are rare among high-risk breast cancer patients and may play a minor contributing role in breast carcinogenesis among Malaysian population.     

Two percent of men with early-onset prostate cancer harbor germline mutations in the BRCA2 gene

Studies of families with breast cancer have indicated that male carriers of BRCA2 mutations are at increased risk of prostate cancer, particularly at an early age. To evaluate the contribution of BRCA2 mutations to early-onset prostate cancer, we screened the complete coding sequence of BRCA2 for germline mutations, in 263 men with diagnoses of prostate cancer who were 相似文献   

Sequencing analysis of SLX4/FANCP gene in Italian familial breast cancer cases

Breast cancer can be caused by germline mutations in several genes that are responsible for different hereditary cancer syndromes. Some of the genes causing the Fanconi anemia (FA) syndrome, such as BRCA2, BRIP1, PALB2, and RAD51C, are associated with high or moderate risk of developing breast cancer. Very recently, SLX4 has been established as a new FA gene raising the question of its implication in breast cancer risk. This study aimed at answering this question sequencing the entire coding region of SLX4 in 526 familial breast cancer cases from Italy. We found 81 different germline variants and none of these were clearly pathogenic. The statistical power of our sample size allows concluding that in Italy the frequency of carriers of truncating mutations of SLX4 may not exceed 0.6%. Our results indicate that testing for SLX4 germline mutations is unlikely to be relevant for the identification of individuals at risk of breast cancer, at least in the Italian population.     

Hereditary breast cancer in Middle Eastern and North African (MENA) populations: identification of novel, recurrent and founder BRCA1 mutations in the Tunisian population

Germ-line mutations in BRCA1 breast cancer susceptibility gene account for a large proportion of hereditary breast cancer families and show considerable ethnic and geographical variations. The contribution of BRCA1 mutations to hereditary breast cancer has not yet been thoroughly investigated in Middle Eastern and North African populations. In this study, 16 Tunisian high-risk breast cancer families were screened for germline mutations in the entire BRCA1 coding region and exon?Cintron boundaries using direct sequencing. Six families were found to carry BRCA1 mutations with a prevalence of 37.5%. Four different deleterious mutations were detected. Three truncating mutations were previously described: c.798_799delTT (916 delTT), c.3331_3334delCAAG (3450 delCAAG), c.5266dupC (5382 insC) and one splice site mutation which seems to be specific to the Tunisian population: c.212?+?2insG (IVS5?+?2insG). We also identified 15 variants of unknown clinical significance. The c.798_799delTT mutation occurred at an 18% frequency and was shared by three apparently unrelated families. Analyzing five microsatellite markers in and flanking the BRCA1 locus showed a common haplotype associated with this mutation. This suggests that the c.798_799delTT mutation is a Tunisian founder mutation. Our findings indicate that the Tunisian population has a spectrum of prevalent BRCA1 mutations, some of which appear as recurrent and founding mutations.