中国上海家族性乳腺癌BRCA1和BRCA2基因的突变

目的研究上海地区家族性乳腺癌中BRCA1／BRCA2基因的突变位点及携带情况。方法研究对象来自35个汉族家族性乳腺癌家系，家系中至少有一个一级亲属乳腺癌患病史。共35例患者，其中13例发病年龄≤加岁。由静脉血提取基因组DNA，对BRCA1／BRCA2基因的全部编码序列进行扩增。扩增产物突变分析先由变性高效液相色谱分析进行筛查，之后进行DNA直接测序证实。结果在BRCA1基因中发现有4个突变位点，其中2个为新发现位点——拼接点突变（IVS17-1G〉T；IVS21＋1G〉C）；另两个为已报道的致病突变位点——移码突变（1100delAT；5640delA）。BRCA2基因的1个致病突变位点位于11号外显子上，为移码突变（5802delAATT）。另外，共发现有12个新的单核苷重复多态位点，都未引起氨基酸编码改变；其中，8个在BRCA1基因上，4个在BRCA2基因上。在家族性乳腺癌中，BRCA1突变频率（11．4％）高于BRCA2基因（2．9％）。结论新发现的2个BRCA1基因的拼接点突变可能是中国上海人群家族性乳腺癌的特有突变位点；在我国上海地区人群中，BRCA1基因突变起着比BRCA2基因更大的作用；该研究丰富了中国人群中BRCA基因的突变谱，并为未来的临床基因检测提供了筛查模式。     

CHEK2基因c.1100delC与中国人遗传性乳腺癌易感性的关联研究

目的研究上海地区非BRCA1／2基因突变的遗传倾向乳腺癌中CHEK2基因c．1100delC突变的携带情况及可能的作用。方法研究对象来自114例遗传倾向性乳腺癌，包括家族性乳腺癌76例，其中8例发病年龄低于40岁；38例单纯早发性乳腺癌（发病年龄〈40岁）。对照组为121名无乳腺癌的健康女性，静脉血中提取基因组DNA，对CHEK2基因的第10～14外显子进行长片段PCR扩增，PCR产物再进行含突变的第10外显子的扩增。突变分析全部由DNA直接测序进行鉴定。结果研究人群和对照人群中都没有发现c．1100delC的突变；在3例（3／114，2．6％）家族性乳腺癌中发现邻近c．1100delC的新的错义突变位点1111C〉T（p-His371 Tyr），对照组中则无此突变发现。结论CHEK2基因c．1100delC突变可能是中国人群罕见的突变位点，在中国人乳腺癌遗传易感性中的作用非常有限；1111C〉T可能与中国上海地区遗传倾向乳腺癌低度外显的易感性有关，需要进行进一步研究确认。     

中国乳腺癌患者BRCA1基因的频发突变5589del8

目的研究在中国大陆乳腺癌人群中是否存在BRCAI／2基因突变的“热点”。方法研究对象为来自全国4个乳腺癌医疗中心的177例家族性和早发性乳腺癌患者和426例散发性乳腺癌患者,根据前期研究中已发现的BRCAI／2基因突变位点,应用变性高效液相色谱分析和DNA测序技术对这些患者进行已知位点的突变检测。结果在前期研究的70例家族性和早发性乳腺癌患者和本研究的177例患者（共247例）中,共发现3例BRCAl5589del8突变的携带者,在426例散发性乳腺癌患者中也发现了2例BRCAl5589del8突变的携带者。单倍型分析的结果显示这5例患者具有相近甚至相同的单倍型。结论BRCAl5589del8突变是中国人群中BRCAl基因的频发突变,它是否是中国人群中的“始祖突变”仍需进一步研究证实。     

Clustering of variations and haplotype analysis in the highly variable region of exon 11 of BRCA1 in Chinese women with sporadic breast cancer

Studies on mutations of BRCA1 gene in Oriental populations, especially in Chinese are sparse. To evaluate the contribution of BRCA1 mutations to sporadic breast cancer in Chinese, mutations in exon 11 from nucleotide positions 3533 to 3682(U14680), a highly variable region of BRCA1 gene, were screened by single-strand conformation polymorphism (SSCP) and DNA sequencing in 329 genomic DNAs from 95 Chinese women with sporadic breast cancer, 29 with benign breast disease and 50 controls. As results, 6 single nucleotide changes were found, and 5 of them (3545A>G, 3551G>T, 3607C>T, 3646T>A and 3661T>C) were newly discovered. The 3545A>G and 3607C>T were germline variations, and the other 3 were somatic variations. In addition, four new haplotypes were defined. Since these newly discovered nucleotide changes were only detected in patients, our results suggest that BRCA1 may also play a role in the development of sporadic breast cancer in Chinese populations.     

Mutational analysis of BRCA1 and BRCA2 genes in Chinese ovarian cancer identifies 6 novel germline mutations

Germline mutations in the BRCA1 and BRCA2 genes predispose women to breast and ovarian cancer. An incidence of 5% and 3.3% respectively has been reported of BRCA1 and BRCA2 mutations in women with ovarian cancer unselected for family history. The contribution of BRCA1 and BRCA2 mutations to ovarian cancer in Chinese women is unknown. A total of 60 samples of ovarian cancer diagnosed in Chinese unselected for age or family history were analyzed for BRCA mutations using the protein truncation test. The entire coding exon of BRCA1 of 53 cases and that of exon 11 of BRCA2 of 43 cases were successfully screened. Six germline (11.3%) mutations (633C>T, 1080delT, 1129delA, 2371-2372delTG, 3976-3979delGTGA, and IVS 22+7 A>G) were detected in BRCA1. One germline mutation (3337C>T) (2.1%) was detected in BRCA2. None of these seven cases were associated with strong family history of breast and/or ovarian cancer. Five out of our six BRCA1 mutations and the one BRCA2 mutation identified are novel. Our 11.3% incidence of BRCA1 mutations in ovarian cancer found amongst Chinese with insignificant family history is apparently higher than that previously reported in other populations. It suggests that BRCA1 mutation may play a significant role in the development of sporadic ovarian cancer in Chinese women.     

BRCA1 Germ‐Line Mutations and Tumor Characteristics in Eastern Chinese Women with Familial Breast Cancer

Although several studies detected the BRCA1 germ‐line mutations in Chinese women with familial breast cancer, most of them did not employ conventional full gene sequencing, especially in eastern China. In addition, the clinicopathological features of BRCA1‐associated breast cancer in Chinese women were not well investigated. In this study, we screened the complete coding regions and exon‐intron boundaries of BRCA1 by polymerase chain reaction (PCR)‐sequencing assay. Immunohistochemistry analyses were performed on tumor samples to detect the expression of estrogen receptor (ER), progesterone receptor (PR), P53, and human epidermal growth factor receptor‐2 (HER‐2). Breast cancer patients having one or more affected relatives referred from the Zhejiang Cancer Hospital, eastern China during 2008–2011 were selected for the study. A total of 62 familial breast cancer patients received the BRCA1 germ‐line mutation screening. Five deleterious mutations were detected in this cohort. The mutation rate was 11.3% (7/62). We found two novel mutations (3414delC and 5,280 C > T) and two recurrent mutations (5,273 G > A and 5589del8). BRCA1 mutation tumors tended to be negative for ER, PR, and HER‐2, and exhibited high histological grade compared with tumors without BRCA1 mutations. Our study suggests that recurrent mutations may exist in eastern Chinese women with familial breast cancer and PCR‐sequencing assay is a useful tool to screen these mutations. It also suggests that BRCA1‐associated breast cancers in Chinese women exhibit an aggressive phenotype. Anat Rec, 2013. © 2012 Wiley Periodicals, Inc.     

Mutation analysis of BRCA1 gene in African-American patients with breast cancer

An estimated 7% of all breast cancers and 10% of all ovarian cancers are associated with inherited mutations in BRCA1 and BRCA2 genes. The mutations of a breast cancer-susceptible gene, BRCA1, confers increased risk of breast cancer in young women. Numerous studies have reported specific mutations in the BRCA1 and BRCA2 genes in the white population. However, there are very few studies on African-American and other ethnic minority groups. The goal of this study is to identify whether African-American patients with breast cancer carry some common mutations reported in other ethnic groups and whether they carry some novel mutations. We screened hot-region mutations on exons 2, 5, 11, 16, and 20 of BRCA1 gene in 54 African-American patients with breast cancer by NIRCA and SSCP methods. Our data revealed one novel frameshift mutation (3331 insG) and three missense sequence variants (A3537G, A3667G, and C4009T) on exon 11. Each sequence change was confirmed by automatic DNA sequencing. One rare sequence variant, A3537G, has been revealed in high frequency (3/54). Our data suggested that African-American patients with breast cancer carry some unique BRCA1 gene mutations.     

BRCA1 mutations in a selected series of breast/ovarian cancer patients.

Germline mutations in the BRCA1 gene have been associated with familial breast/ ovarian cancer in large families showing high penetrance of the disease. Little is known, however, about the contribution of BRCA1 mutations to breast/ovarian cancer in small families with few affected members or in isolated early onset cases. Therefore we examined the BRCA1 gene in 63 breast/ovarian cancer patients who either came from small families with as few as one affected first degree relative, or in patients who had no family history but had developed breast cancer under 40 years of age. Using the protein truncation test, we were able to identify three unique BRCA1 germline mutations (4.8%). Two of the probands had only one affected first degree and several second degree relatives and the third had three affected first degree relatives including two sisters who, when tested, were also found to carry the mutation. There was no family history of ovarian cancer in any of the three families.     

Contribution of the BRCA1 and BRCA2 mutations to breast cancer in Tunisia

Hereditary breast cancer accounts for 3–8% of all breast cancers, with mutations in the BRCA1 and BRCA2 genes responsible for up to 30% of these. To investigate the prevalence of BRCA1 and BRCA2 gene mutations in breast cancer patients with affected relatives in Tunisia, we studied 36 patients who had at least one first degree relative with breast and/or ovarian cancer Thirty-four 34 patients were suggestive of the BRCA1 mutation and two were suggestive of the BRCA2 mutation, based on the presence of male breast cancer detected in their corresponding pedigrees. Four mutations in BRCA1 were detected, including a novel frame-shift mutation (c.211dupA) in two unrelated patients and three other frameshift mutations – c.4041delAG, c.2551delG and c.5266dupC. Our study is the first to describe the c.5266dupC mutation in a non-Jewish Ashkenazi population. Two frameshift mutations (c.1309del4 and c.5682insA) were observed in BRCA2. Nineteen percent (7/36) of the familial cases had deleterious mutations of the BRCA1 or BRCA2 genes. Almost all patients with deleterious mutations of BRCA1 reported a family history of breast and/or ovarian cancer in the index case or in their relatives. Our data are the first to contribute to information on the mutation spectrum of BRCA genes in Tunisia, and we give a recommendation for improving clinical genetic testing policy.     

新疆地区维吾尔族乳腺癌BRCA1基因突变分析

目的研究新疆地区维吾尔族乳腺癌患者BRCA1基因突变情况及突变位置。方法选取70例维吾尔族乳腺癌根治标本,对照组为32例维汉族乳腺良性病变（纤维腺病及纤维腺瘤）及乳腺癌旁非癌组织;应用PCR-单链构象多态性和DNA序列测定的方法检测BRCA1基因突变。结果（1）70例维吾尔族乳腺癌中发现9例BRCA1突变的12个新位点。（2）70例维吾尔族乳腺癌BRCA1的突变率为12.86%（9/70）,22例维吾尔族早发性乳腺癌（≤35岁）BRCA1突变率为31.82%（7/22）。维吾尔族早发性乳腺癌BRCA1突变率（7/22）高于维吾尔族晚发性乳腺癌（2/48）,差异有统计学意义（χ^2=10.295,P〈0.01）。（3）70例维吾尔族乳腺癌中发现9例BRCA1基因核苷酸多态性位点,其中8例多态性位点均为3232A〉G。（4）2例双侧乳腺癌中均检测出BRCA1基因的突变。结论BRCA1突变可能与新疆维吾尔族乳腺癌尤其是维吾尔族早发性乳腺癌及双侧乳腺癌的发生密切相关。     

Contribution of BRCA1 and BRCA2 germline mutations to the incidence of early-onset breast cancer in Cyprus

In Cyprus, the prevalence of breast cancer associated with BRCA1 and BRCA2 mutations in young women is unknown. In this study, we present the results of mutational analysis of the BRCA1 and BRCA2 genes in 26 Cypriot women diagnosed with breast cancer by the age of 40. The entire coding regions, including splice sites, of the BRCA1 and BRCA2 genes were sequenced using cycle sequencing. We identified four pathogenic mutations: two in BRCA1 [c.1840A>T (K614X), c.5310delG (5429delG)] and two in BRCA2 [c.3531-3534delCAGC (3758del4), c.8755delG (8984delG)] in six of 26 unrelated patients. The BRCA2 mutation c.3531-3534delCAGC (3758del4) is novel and the BRCA1 mutation c.1840A>T (K614X) is reported for the first time in Cypriot patients. The BRCA2 Cypriot founder mutation c.8755delG (8984delG) was detected in three unrelated patients. Additionally, we identified one novel BRCA1 missense mutation, two novel polymorphisms and three novel intronic variants of which BRCA1 c.4185+3A>G (IVS12+3A>G) may be pathogenic. Of the six BRCA1/2 mutation carriers, only four had a family history. These results show that the prevalence of BRCA1 and BRCA2 mutations in Cypriot women diagnosed with early-onset breast cancer is high. We conclude that Cypriot women with early-onset breast cancer should be offered BRCA1/2 testing irrespective of their family history.     

Founder mutation in the BRCA1 gene in Malay breast cancer patients from Singapore

The mutation spectrum of the BRCA1 gene among ethnic groups from Asia has not been well studied. We investigated the frequency of mutations in the BRCA1 gene among Malay breast cancer patients from Singapore, independent of family history. By using the protein truncation test (PTT) and direct sequencing, BRCA1 mutations were detected in 6 of 49 (12.2%) unrelated patients. Four novel missense mutations in exon 11, T557A (1788A>G), T582A (1863A>G), N656S (2086A>G) and P684S (2169C>T) were identified in one patient. Two patients had missense mutations in exon 23, V1809A (5545T>C), which has been previously detected in individuals from Central and Eastern Europe. Three unrelated patients had the deleterious 2846insA frameshift mutation in exon 11. Methylation specific PCR (MSP) of the promoter region of the BRCA1 gene detected hypermethylation of tumor DNA in an additional 2 patients. Haplotype analysis using the microsatellite markers D17S855, D17S1323 and D17S1325 revealed a common haplotype for the three unrelated patients and their three relatives with the 2846insA mutation. These findings strongly suggest that the 2846insA mutation, the most common deleterious mutation in this study, may possibly be a founder mutation in breast cancer patients of Malay ethnic background.     

Novel germline mutations in the BRCA1 and BRCA2 genes in Indian breast and breast-ovarian cancer families

The two major hereditary breast/ovarian cancer predisposition tumor suppressor genes, BRCA1 and BRCA2 that perform apparently generic cellular functions nonetheless cause tissue-specific syndromes in the human population when they are altered, or mutated in the germline. However, little is known about the contribution of BRCA1 and BRCA2 mutations to breast and/or ovarian cancers in the Indian population. We have screened for mutations the entire BRCA1 and BRCA2 coding sequences, and intron-exon boundaries, as well as their flanking intronic regions in sixteen breast or breast and ovarian cancer families of Indian origin. We have also analyzed 20 female patients with sporadic breast cancer regardless of age and family history, and 69 unrelated normal individuals as control. Thus a total of 154 samples were screened for BRCA1 and BRCA2 mutations using a combination of polymerase chain reaction-mediated site directed mutagenesis (PSM), polymerase chain reaction-single stranded conformation polymorphism assay (PCR-SSCP) and direct DNA sequencing of PCR products (DS). Twenty-one sequence variants including fifteen point mutations were identified. Five deleterious pathogenic, protein truncating frameshift and non-sense mutations were detected in exon 2 (c.187_188delAG); and exon 11 (c.3672G>T) [p.Glu1185X] of BRCA1 and in exon 11 (c.5227dupT, c.5242dupT, c.6180dupA) of BRCA2 (putative mutations - four novel) as well as fourteen amino acid substitutions were identified. Twelve BRCA1 and BRCA2 missense variants were identified as unique and novel. In the cohort of 20 sporadic female patients no mutations were found.     

Mutation analysis of BRCA1 and BRCA2 from 793 Korean patients with sporadic breast cancer

To investigate the role of BRCA1 and BRCA2 mutations in Korean patients with sporadic breast cancer, 793 breast cancer patients were analyzed by denaturing high performance liquid chromatography and direct sequencing. The 793 breast cancer patients enrolled in this study had no family history of affected first- or second-degree relatives with breast and/or ovarian cancer. Seventy-nine different sequence variations were identified, of which 34 were novel. Fifteen deleterious mutations were detected in 20 out of 793 patients (2.5%): 11 frameshift mutations and 4 nonsense mutations (seven in BRCA1 and eight in BRCA2), and no recurrent or founder mutations were observed in BRCA mutation screening. However, three mutations (K467X, 3972delTGAG, and R2494X in BRCA2) were identified in other studies of the Korean population. Of 793 patients, the clinicopathological information was obtained in 135 patients, who included 20 deleterious mutation-positive and 115 deleterious mutation-negative groups. The median age at diagnosis, histologic type, histologic grade and T stage did not show statistically significant difference between these two groups. BRCA-mutation-associated tumors showed lower estrogen receptor, progesterone receptor, and HER-2/neu but higher p53 expression. Although poor prognostic features were noted in BRCA-associated tumors, we did not find statistically significant differences. The present study will be helpful in the evaluation of the need for the genetic screening of germline BRCA mutations and reliable genetic counseling for sporadic breast cancer patients.     

Identification of fifteen novel germline variants in the BRCA1 3'UTR reveals a variant in a breast cancer case that introduces a functional miR-103 target site

Mutations in the BRCA1 gene confer a substantial increase in breast cancer risk, yet routine clinical genetic screening is limited to the coding regions and intron-exon boundaries, precluding the identification of mutations in noncoding and untranslated regions (UTR). As 3'UTR mutations can influence cancer susceptibility by altering protein and microRNA (miRNA) binding regions, we screened the BRCA1 3'UTR for mutations in a large series of BRCA-mutation negative, population and clinic-based breast cancer cases, and controls. Fifteen novel BRCA1 3'UTR variants were identified, the majority of which were unique to either cases or controls. Using luciferase reporter assays, three variants found in cases, c.* 528G>C, c.* 718A>G, and c.* 1271T>C and four found in controls, c.* 309T>C, c.* 379G>A, c.* 823C>T, and c.* 264C>T, reduced 3'UTR activity (P < 0.02), whereas two variants found in cases, c.* 291C>T and c.* 1139G>T, increased 3'UTR activity (P < 0.01). Three case variants, c.* 718A>G, c.* 800T>C, and c.* 1340_1342delTGT, were predicted to create new miRNA binding sites and c.* 1340_1342delTGT caused a reduction (25%, P = 0.0007) in 3'UTR reporter activity when coexpressed with the predicted targeting miRNA, miR-103. This is the most comprehensive identification and analysis of BRCA1 3'UTR variants published to date.     

BRCA1 and BRCA2 in Indian breast cancer patients

Incidence of breast cancer in Indian women is not as high as in Western countries, nonetheless age-adjusted incidence rates (AAR) have risen from 17.9 to 24.9 per 100,000 from 1965 to 1985. Although these rates are still approximately one quarter to one third of incidence rates in North America and Europe, respectively, due to the large population of women at risk, nearly 80,000 new cases were diagnosed in India in 2000. Although identification of BRCA1 and BRCA2 has greatly increased our understanding of breast cancer genetics in populations of Western European descent, the role of these genes in Indian populations remains unexplored. Analysis of a series of 20 breast cancer patients from North India with either family history of breast and/or ovarian cancer (2 or more affected first degree relatives) or early age of onset (<35 years) led to identification of two novel splice variants (331+1G>T; 4476+2T>C) in BRCA1 (10%). In addition, two BRCA2 missense variants were each identified in more than one patient (two unrelated individuals each) and likely represent population-specific polymorphisms.     

BRCA1 and BRCA2 mutations in women with familial or early-onset breast/ovarian cancer in the Czech Republic

Germline mutations in BRCA1 and BRCA2 account for majority of hereditary breast and ovarian cancer. The complete coding sequence analysis of both genes was carried out in 197 breast/ovarian cancer patients from high-risk families and 53 patients with sporadic breast/ovarian cancer. In summary, 59 mutations (16 different) in BRCA1 and 29 mutations (17 different) in BRCA2 were identified in unrelated breast and/or ovarian index cases. Using the BIC Database numbering, the most frequently found mutations in BRCA1 were c.5385dupC (22 cases), c.3819_3823delGTAAA (8 cases) and c.300T>G (6 cases). The most frequently found mutations in BRCA2 were c.8138_8142delCCTTT (7 cases) and c.8765_8766delAG (7 cases). Altogether, these 5 mutations represented 56.8% of all detected mutations. A broad spectrum of other mutations was detected including four novel mutations (c.2881delA in BRCA1; and c. 6677_6678delAA, c.6982dupT and c.8397_8400dupTGGG in BRCA2). Deleterious mutations were found in 80 (40.6%) of 197 high risk-families, in 6 (37.5%) of 16 patients with sporadic bilateral breast, ovarian or both cancers and in 2 (6.2%) of 32 women with sporadic early-onset unilateral breast cancer. No mutation was detected in 5 cases of sporadic early-onset unilateral ovarian cancer.     

Recurrent BRCA1 and BRCA2 germline mutations in ovarian cancer: a founder mutation of BRCA1 identified in the Chinese population

Previous mutational analysis for BRCA gene mutations in sporadic ovarian cancer occurring in Chinese patients in Hong Kong identified six germline BRCA1 mutations and one germline BRCA2 mutation, six of which were novel (Khoo et al., 2000). Knowledge of BRCA gene mutations in the Chinese population is relatively scant. In this study, we focussed on whether any of these mutations could be recurrent in our Chinese population, making use of archival paraffin embedded tissue. A consecutive series of 214 ovarian cancer cases, half of Southern Chinese origin from Hong Kong whilst the other half of Northern Chinese origin from Beijing were used for the study. We identified one further novel mutation, 1081delG, in BRCA1. This was found to occur in two unrelated individuals with shared haplotype as revealed by allelotype analysis, thus demonstrating founder effect. Two other recurrent mutations were also identified, the 2371-2372delTG mutation in BRCA1 and the 3337C>T mutation in BRCA2 recurring in two and three unrelated individuals respectively, giving an overall prevalence 4.7% of recurrent BRCA mutations in ovarian cancer in the Southern Chinese population. Most importantly, all our recurrent mutation carriers were identified from Southern Chinese patients from Hong Kong whilst such mutations were absent in samples from the Northern Chinese. Our findings indicate possible heterogeneity in the BRCA genotype between Northern and Southern Chinese. The identification of a founder mutation and two recurrent mutations moreover, has important implications towards screening strategies for breast and ovarian cancer among Chinese of southern ancestral origin who are now dispersed throughout the world.