OL Oncology Letters 1792-1074 1792-1082 D.A. Spandidos 10.3892/ol.2018.8538 OL-0-0-8538 Articles Multicenter cross-sectional screening of the BRCA gene for Chinese high hereditary risk breast cancer populations WeiHongyi 1 WangMinghao 1 OuJianghua 2 JiangWeihua 2 TianFuguo 3 ShengYuan 4 LiHengyu 4 XuHong 5 ZhangRuishan 5 GuanAihua 6 WangChangqing 6 JiangHongchuan 7 RenYu 8 HeJianjun 8 LiuJian 9 HuangWeiwei 9 LiaoNing 10 CaiXiangjun 11 MingJia 12 LingRui 13 XuYan 14 HuChunyan 14 ZhangJianguo 15 GuoBaoliang 15 OuyangLizhi 16 ShuaiPing 17 LiuZhenzhen 18 ZhongLing 1 JingRuilin 19 ZengZhen 1 ZhangMeng 19 ZhangTing 1 XuanZhaoling 19 TanXuanni 1 LiangJunbin 19 PanQinwen 1 ChenLi 1 ZhangFan 1 FanLinjun 1 ZhangYi 1 YangXinhua 1 LiJingbo 1 ChenChongjian 19 JiangJun 1 Department of Breast Surgery, Southwest Hospital, The Third Military Medical University, Chongqing 400038, P.R. China Department of Breast Surgery, The Affiliated Tumor Hospital of Xinjiang Medical University, Urumqi, Xinjiang 830011, P.R. China Breast Surgery, Shanxi Cancer Hospital, Taiyuan, Shanxi 030013, P.R. China Department of Thyroid and Breast Surgery, Changhai Hospital, Second Military Medical University, Shanghai 200433, P.R. China Department of Breast Surgery, Liaoning Cancer Hospital and Institute, Shenyang, Liaoning 110042, P.R. China Department of Breast Surgery, Jilin Cancer Hospital and Institute, Changchun, Jilin 130000, P.R. China Department of General Surgery, Beijing Chaoyang Hospital Affiliated to Capital Medical University, Beijing 100020, P.R. China Department of Breast Surgery, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi 710061, P.R. China Department of Medical Oncology, Fujian Provincial Cancer Hospital, Fuzhou, Fujian 350014, P.R. China Breast Department, Cancer Center, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong 510080, P.R. China Department of Hepatobiliary and General Surgery, PLA No. 202 Hospital, Shenyang, Liaoning 110812, P.R. China Department of Breast, Thyroid, and Pancreas Surgery, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, P.R. China Depatement of Thyroid, Breast and Vascular Surgery, Xijing Hospital, Forth Military University, Xi'an, Shaanxi 710032, P.R. China Department of Breast Surgery, Daping Hospital, Research Institute of Surgery, Third Military Medical University, Chongqing 400038, P.R. China Department of Breast Surgery, The Second Affiliated Hospital of Harbin Medical College, Harbin, Heilongjiang 150086, P.R. China Department of Breast Surgery, Hunan Cancer Hospital, Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan 4100013, P.R. China Health Management Center, Sichuan Provincial People's Hospital, Chengdu, Sichuan 610072, P.R. China Department of Breast Surgery, Henan Cancer Hospital, Zhengzhou, Henan 450008, P.R. China Annoroad Gene Technology (Beijing) Co., Ltd., Beijing 100176, P.R. China Correspondence to: Dr Jun Jiang, Department of Breast Surgery, Southwest Hospital, The Third Military Medical University, 30 Gaotanyan Street, Chongqing 400038, P.R. China, E-mail: jcbd@medmail.com.cn Dr Chongjian Chen, Annoroad Gene Technology (Beijing) Co., Ltd., 88 Kechuang Six Street, Beijing 100176, P.R. China, E-mail: cchen@annoroad.com 06 2018 18 04 2018 15 6 9420 9428 11082017 25102017 Copyright © 2018, Spandidos Publications 2018

Due to lack of systematic reviews, BRCA, DNA Repair Associated (BRCA) mutations in the Chinese population are not completely understood. The following study investigates the prevalence and type of BRCA mutations in Chinese patients with high hereditary risk of breast cancer (BC). Patients Drwere recruited from 14 cities between October 2015 and February 2016, and were selected based on family and personal medical history. BRCA mutations were analyzed by collecting blood samples from all participants. 437 BC patients were included. A total of seventy-six (17.4%) mutation carriers were identified with no geographic difference. The mutation rate in the early-onset BC patients was lower compared to family history of breast/ovarian cancer (OC), bilateral BC, male BC, BC&OC or meeting ≥2 criteria (9.2 vs. 21.7, 24.0, 22.2, 16.7 and 24.3%, respectively, P=0.007). A total of 61 mutation sites were identified (BRCA1 32, BRCA2 29) including 47.5% novel sites and extra 10 variants of uncertain significance. A total of five sites were repeated in more than one unrelated patient. A total of 11 sites were associated with hereditary breast and ovarian cancer syndrome, two of which were confirmed by family pedigrees. Compared with BRCA− patients, patients with BRCA1 mutation tended to be triple-negative BC (P<0.001), whereas patients with BRCA2 mutation were more likely to be hormone receptor positive BC (P=0.02). The present study provides a general BRCA mutation profile in the Chinese population. The prevalence of BRCA mutation in BC patients with high hereditary risk is lower compared with Western populations. Chinese mutation type is different with Western people, without obvious founder mutation.

 breast cancer BRCA1 BRCA2 chinese population Introduction

Even though, the incidence of breast cancer (BC) in China is lower compered to Western countries, since the 1990s it has increased twice as fast as the global rate (1) with very early onset (2,3). In China, BC patients <50 years old account for 46% of the total cases (4), with the peak prevalence observed in the 45 to 55 years old age (5–8). Meanwhile, the highest mortality is observed in even younger age i.e. in the 30 to 44 years old age (5). Therefore, in order to reduce the number of BC patients, and the growing burden of this disease, it is urgent to promote BC prevention in China.

BRCA (Breast Cancer Susceptibility Gene) is associated with the majority of hereditary BC, which accounts for about 5 to 10% of all cases of breast cancers (9). Meanwhile, the resemblance in clinical and pathologic features between sporadic triple-negative BC (TNBC) and BRCA1 mutant BC implies that mechanism behind BRCA germline mutant tumors is strongly associated with somatic mutation of sporadic BC (10). However, a lack of basic information about the prevalence and spectrum of BRCA mutations hinders research progress on the etiology (6) and risk evaluation of model of breast cancer prevention in China. Additionally, because of the large presence of ethnic-specific contexts (11–14), the Western risk evaluation models do not apply well to China. Therefore, the following paper assembled a wide-range clinic-based cross-sectional study of hereditary risk among BC patients, who were representative for BRCA mutations study. The patients were from different parts of mainland China, which was important for determining the prevalence and types of BRCA mutations, as well as to provide basic information for further studies of BRCA and BC prevention models in China.

 Materials and methods <sec> <title>Study population

The present study has been approved by center medical ethics committee (Ethics Committee of the First Affiliated Hospital of Third Military Medical University, PLA), and was successfully registered with Chinese Clinical Trial Registry (ChiCTR), which is the international clinical trial registration platform.

In the time period from October 2015 to February 2016, 445 patients diagnosed with BC were recruited from 18 tertiary general clinics located in North, South and Northwest China, which account for 3/4 of Chinese national territory (Fig. 1). Among these patients, three refused to participate in the study, while for other five the recruitment criteria couldn't be confirmed. Pathological examination was used to confirm the cancer diagnoses and breast cancer subtype. Furthermore, twenty-seven participants were additionally excluded from the clinical data analysis due to incomplete or illegible clinical information. Where it was possible, family members of probands with confirmed BC were recruited to participate.

After reading and signing the informed consent, participants were interviewed in order to provide medical details including past medical history, present BC diagnosis age, tumor size, states of axillary lymph nodes and metastasis, and details of family cancer history. Consequently, fresh peripheral venous blood (5 ml) was collected from each participant, and transferred into a coded Ethylene Diamine Tetraacetie Acid (EDTA) tube at 4°C. During the same day, blood samples were sent to Annoroad Gene Technology (Beijing) Co., Ltd, for further analyses (BRCA genetic testing). Each participant received a sealed file filled with BRCA test result (excluding their clinical records). Genetic consultation was provided in order to better explain the BRCA test results. The participants did not encounter any financial costs (including genetic consultation) for their participation in the present study.

 Recruitment criteria

The recruitment criteria adhered to the breast cancer diagnosis and treatment guidelines and specifications (Chinese Cancer Society, V2015). Participants who met one of the following criteria were included in the study: i) BC with onset age ≤35 years old (early-onset BC group), ii) ≥1 relativea (either sex) from the same side of the family as BC patient and (or) ovarian cancer (BC/OC) diagnosed at any age (BC/OC family history group), iii) two primary BC cancersb [bilateral BC (BBC) group], iv) male BC patients (MBC group), v) BC with OC (BC&OC group) and vi) meeting ≥2 criteria above simultaneously (mixed group). aIncluding first, second and third degree relative with no age limitation. bTwo primary BC, bilateral BC excluding metastatic contralateral BC or unilateral BC including two or more different types of cancers.

In case the patients failed to sign the informant consent, they were excluded from the present study.

 BRCA gene analysis

BRCA1 (MIM:113705) and BRCA2 (MIM:600185) testing was performed using Annoroad Gene Technology (Beijing, China) on an Illumina HiSeq 2500 platform (Illumina, San Diego, CA, USA). Genomic DNA was first extracted from peripheral blood white cells using the QIAGEN DNeasy Blood and Tissue Kit (Qiagen, Shanghai, China). The genomic DNA was then fragmented by a bioruptor sonication device (Diagenode, Leige, Belgium). The construction and capture of the DNA library followed the standard protocols from Illumina (Illumina, San Diego, CA, USA) and Roche (Roche, Shanghai, China). A Qubit 3.0 Fluorometer (Invitrogen, San Francisco, CA, USA) and Bioanalyzer (Agilent, Santa Clara, CA, USA) were used to determine the quantity of the library. Finally, the library was sequenced on one lane using 100 paired-end (2x100 bp) strategies.

 Variant nomenclature

Reference sequences used for BRCA1 and BRCA2 analyses were GenBank NM_007294.2 (BRCA1) and NM_000059.3 (BRCA2). Mutation nomenclature was described according to Human Genome Variation Society (v2.0) (15).

 Statistical analysis

Medians were used with interquartile ranges of abnormally distributed data for continuous variables (diagnosed ages of patients) and rank test for analyses. Proportions were shown for categorical variables. Comparisons of mutation rates and proportions were analyzed by Chi-square test of unordered categorical variable. Univariate and multivariate logistic analysis were used to examine the relationships between hormone receptors (HRs, including estrogen receptor ER, progesterone receptor PR) and human epidermal growth factor receptor 2 (HER2) and BRCA mutation states with odd ratios (OR) and 95% confidence intervals (95% CI).

All P-values were two-sided. P<0.05 was considered to indicate a statistically significant difference. All data were analyzed using PASW Statistics 22.0 (SPSS, Inc., Chicago, IL, USA).

 Results <sec> <title>Study population

From a total of 437 BC patients enrolled in the study, almost half came from north China (Table I). Forty percent of early-onset BCs (173/437) and 30% of patients with BC/OC family history (163/437) were observed in more than half of the participants (Table II). The median age of 437 BC patients was 35.0 (31.0, 46.0) years. The median age of mutation carriers was higher compared to non-carriers [41.0 (34.0, 47.0) vs. 35.0 (30.0, 46.0), P=0.002].

 Mutation frequency

Seventy-six (17.4%) BRCA mutation carriers were identified, 31 (15.4%) of which were from patients from North China, 30 (19.9%) from patients from South China, and 15 (17.6%) from patients from Northwest China. No significant difference in gene mutation rates was found between different region areas (Table I). Furthermore, the early-onset patient rate (9.2%) was significantly different in relation to remaining 5 groups (P=0.007) (Table II); while no significant difference was found between the 5 groups (P>0.05).

 BRCA mutation status

According to the American College of Medical Genetics and Genomics (ACMG) (16), a total of 61 deleterious mutation points (29 in BRCA1, 32 in BRCA2) were observed in 76 carriers (Table III), and consequently classified into the already ‘known’ (Table IV) and the ‘novel’ (Table V) mutations. Briefly, 72% of novel variations were found in BRCA2 (Tables IV and V). Moreover, five mutations of 61 were observed in more than one unrelated patients from different areas (Fig. 2 and Table IV).

Thirty-four points (55.7%) were frame shift, followed by 17 (27.9%) nonsenses, 7 splices (11.5%), 2 pathogenic missenses (3.3%), and 1 synonymous mutation (1.6%). Extra 10 missenses were found as variants of uncertain significance (VUS) (Table VI), accounting for 14.1% of all the variants including 61 deleterious mutation.

Eleven different mutations in 10 families were related to hereditary breast and ovarian cancer syndrome (HBOC), four in BRCA2 were novel (Table VII). Moreover, none of the identified mutations were shared between the families. BRCA1 c.190T>C&BRCA2 c.9090dup were found in one same family, and were carried by a proband from maternal and paternal line respectively. The proband's BC was inherited from maternal HBOC (Fig. 3A). In BRCA1 c.5431C>T hereditary family, all the middle-age females had BC/OC, but all the male carriers of four general relatives were healthy (Fig. 3B).

 Clinical analysis

410 precise tumor node metastasis TNM results were obtained. No significant difference was found among wild type BRCA (BRCA−), BRCA1 and BRCA2 mutation patients (Table VIII).

Univariate analysis demonstrated higher expression of ER negative (ER−) and PR negative (PR−) in BRCA1 group compared to BRCA− group (72.6% vs. 35.1%, P<0.0001; 82.4% vs. 40.6%, P<0.0001). Conversely, higher expression of ER positive (ER+) and PR positive (PR+) were found in BRCA2 group compered to BRCA− group patients (93.5% vs. 64.9%, P=0.001; 87.1% vs. 59.4%, P=0.002). Moreover, BRCA1 and BRCA2 groups were both more frequently HER2 negative (HER2−) compared to BRCA− group (97.1, 96.8% vs. 74.3%, P=0.003 and 0.005 respectively). Based on the multivariate analysis, PR− and HER2− were the independent risk factors for BRCA1 mutation, HER2− alone for BRCA2 (Tables IX and X). Then, compared with BRCA−, BRCA1 mutation tended to be TNBC (68.6% vs. 24.8%, P<0.0001), while BRCA2 mutation had higher proportion of HRs positive (HRs+) BC (93.5% vs. 75.2%, P=0.002) (Table XI).

 Discussion

The results from the present study, which is to our knowledge, the largest screening study ever performed in China, reveal that the total BRCA mutation rate is 17.4% for breast cancer patients at risk of hereditary BRCA mutation across China with no observed geographical differences.

One of the main components of the present study has to do with 21.7% of mutation rate which lies in BC/OC family history subgroup. This finding is in line with a Korean study conducted across 36-centers (22.3%) (17). The mutation rate appears generally lower compared to Western countries (23~35.3%) (18–22), but higher compared to Peking or Shanghai regions (10.5~18.2%) (23–25). Lower prevalence of BRCA mutation is in line with comparisons of BC incidence with the Western countries. It clearly suggests essential distinction in BRCA mutation between Asian and Euromerican people that goes well beyond different study design biases. Furthermore, thus far observed domestic inconsistencies may be caused by the limitations related to areas and criteria. The present study covers most of the Chinese regions, i.e. areas with huge concentrations of Chinese populations. Moreover, the present study does not impose the BC onset age limitation for family cancer history, which allows for wider screening rang. However, some families with late-onset hereditary BC/OC that are really in need of BRCA testing may be excluded from testing due to young cutoff diagnose age established by BRCA testing guideline. Most of all, our results are representative of the real data on hereditary risk for breast cancer patients in China.

The sporadic early-onset (≤35 years) BC mutation rates (9.2%) are nearly twice higher compared to those obtained by the recent studies from China (5%, ≤40 years) (24) and from Western countries (5.9%, <36 years) (26), but lower compared to other 5 risk factor groups in this study. The observed discrepancy may come from different sample sizes and study populations. Moreover, different age limitations for early-onset suggest that cutoff age may impact the mutation rates in younger patients. Currently, the age of 45 is set as upper limitation of young BC for BRCA testing according to Chinese BC Treatment Guideline (27). Nevertheless, increasing trend of breast cancer incidence in younger patients in China (6) may imply more patients with sporadic early-onset from the whole population. Ten years interval, between the ages 35 and 45, may actually double the difference on sensitivity and specificity for BRCA screening in Chinese patients with breast cancer. Additionally, besides onset age, family history, bilateral BC, with OC and similar should also be considered in the early-onset BC to improve BRCA test indications (24). Consequently, we suggest re-evaluation of the early-onset age for BRCA test to increase sensitivity of BRCA mutation screening and to fit current BC epidemiology in China.

BRCA mutation sites in our study suggest special features of BRCA mutation in China. Four sites are hereditary, and nearly half are novel. Also, 14.1% VUS were slightly higher than Asian data (13.6%) obtained from a study conducted by Hall et al. Interestingly, their data were already more than 2.5 times higher compared to European VUS (28). Then, BRCA1 c.2572C>T reappeared twice in two unrelated patients from Chongqing and Shanxi Provinces, respectively. Up to now, it has been found in unrelated patients from five different provinces in China (29). Nevertheless, it has not been frequently reported in international studies, meaning it might be one of founder mutation candidates among Chinese populations. BRCA1 c.5470_5477del is another highly repeated site found three times in our study. Internationally, the site has been previously identified in Korean sample (30), while nationally it has been reported in Shanghai (31), Beijing (32), Zhejiang, and Liaoning (33). Thus far, it has been found in 12 unrelated carriers from six different areas of China, including HBOC patients (33.3%) (32), sporadic BC patients (33.3%) (31,33) and early-onset BC (33) or BBCpatients (33.3%) (the present study). Also, three of the patients (from Zhejiang, Shanghai, and Liaoning) share the same haplotype (33). To our knowledge, it has only once been reported in native Asian patient (34), therefore, it is strongly associated with Asian founder mutation. However, Kwong et al have recognized BRCA2 c.3109C>T as a founder mutation in southern Chinese population in Hong Kong (35,36). 81.5% of probands in the study were immigrants from Guangdong. It was repeated in two unrelated patients from Chongqing, southwest of China, but it did not appear in our population sample from southern China. Further large scale unselected population epidemic research is necessary to clarify this.

Two confirmed HBOC families have special features. The proband who carried two mutations is the youngest patient in her family, thus it is not possible to ignore the effects of paternal inherited BRCA2 c.9090dup (novel) on the proband given her uncle has lung cancer (Fig. 3A). BRCA1 c.5431C>T pathogenicity shows a gender trend: It appears harmful for women, and silent for men (Fig. 3B).

All the information on the mutation sites reported above suggest quite different BRCA mutation spectrum and features that exit in Chinese BC patients compared with well-known white populations.

Similar to other studies, we came to the conclusion that BRCA1 mutation is concerned with TNBC. However, BRCA2 mutations tend to be HRs+ BC, which is inconsistent with results from other studies (37,38). The different results show heterogeneous of BRCA2 mutant BC beyond different study populations and regions (39,40). The present study suggests that BRCA2 mutant BC may responds better to endocrine therapy due to high proportion of HRs+ tumor with the same background of hereditary risk of BC compared with BRCA− and BRCA1 mutant BC.

In summary, this study provides a general BRCA mutation profile in China, which enhances the prevalence of BRCA mutations in non-white populations. The BRCA screening provides a distinguishing BRCA mutation profile in China, which compared to the West reveals lower mutation prevalence, and special mutation spectrum. Cutoff ages for diagnosis of early-onset and BC/OC family history should be re-evaluated based on population screening data to improve BRCA test indications. BRCA2 mutation suggests the best response to endocrine therapy among BRCA mutant and BRCA− BCs in this selected hereditary risk population. However, further studies are necessary to confirm precise BRCA mutation situation in China.

 Acknowledgements

The authors would like to thank Zefei Jiang, Mengmeng Zhang, Shaohua Wei, Yuanyuan Wang, Dawei Yun, Huiquan Jiang, Yang Li, Yimeng Hu, Yong Yang and other medical staff for their involvement in the study design, BRCA lab test, information collection, data analysis and manuscript writing. The present study was supported by the China Health Promotion Foundation.

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The regional distribution of subjects in China. The four regions of China's territory are indicated with red lines. Areas where patients came from are indicated with blue.

The regional distribution of unrelated subjects carrying the same mutations. The areas of BRCA1 c.2572C>T carriers are indicated with dark blue; BRCA1 c.4801A>T in green; BRCA1 c.5470_5477del in yellow; BRCA2 c.2806_2809del in light blue; and the areas of BRCA2 c.3109C>T carriers are indicated in red.

Family pedigrees of BRCA1 c.190T>C&BRCA2 c.9090dup and BRCA1 c.5431C>T. Cancer patients with mutations are indicated with solid symbols, while the healthy carriers are indicated with half solid symbols. The late individuals are indicated with bias crossing symbols. Probands are indicated with arrows. Unknown BRCA state is indicated question mark. Ages under symbols indicate cancer diagnosed age for patients and current age for healthy carriers. BC, breast cancer; OC, ovarian cancer; HHC, liver cancer; LC, lung cancer. (A) Family pedigree BRCA1 c.190T>C&BRCA2 c.9090dup. IIe is a lung cancer patient who rejected to have BRCA testing. (B) Family pedigree of BRCA1 c.5431C>T. Ia is a female breast cancer patient without BRCA1 c.5431C>T.

The geographic mutation rates (n=437).

Area N BRCA+a (%) P-value
North China 201 31 (15.4) 0.5
South China 151 30 (19.9)
Northwest China 85 15 (17.6)
Total 437 76 (17.4)

Both BRCA1 and BRCA2 mutations.

The mutation rates in the recruited groups (n=437).

Recruit Criteria N BRCA+a (%) P-value
EOb 173 16 (9.2)
BC/OC FH 129 28 (21.7)
BBC 50 12 (24.0)
MBC 9 2 (22.2) 0.007
BC&OC 6 1 (16.7)
Fixed 70 17 (24.3)
Total 437 76 (17.4)

EO, early-onset breast cancer; BC/OC FH, breast and/or ovarian cancer family history; BBC, bilateral breast cancer; MBC, male breast cancer; BC&OC, breast cancer with ovarian cancer.

Both BRCA1 and BRCA2 mutations.

Patients were diagnoses with BC onset breast cancer, BC/OC FH, breast and/or ovarian cancer family history; BBC, bilateral breast cancer.

Deleterious and novel mutations (n=61).

Gene Known (%) Novel (%) Total
BRCA1 21 (65.6) 8 (27.6) 29
BRCA2 11 (34.4) 21 (72.4) 32
Total 32 (52.5) 29 (47.5) 61

BRCA known deleterious mutation sites (n=32).

Gene Location Exon Mutation type AA change Probands
BRCA1 c.190T>Cb 4 M p.Cys64Arg HOBC
c.212G>Ac 4 M p.Arg71Lys HBOC
c.212+1G>Tb Intr S – MBC/BCFH
c.441+1G>A Intr S – EO
c.1660G>Ta 10 N p.Glu554Ter BCFH
c.1674del 10 FS p.Gly559fs MBC
c.2014A>Tb 10 N p.Lys672Ter BCFH
c.2572C>Td 10 N BBC
c.3329dup 10 FS p.Gln1111fs EO
c.3400G>T 10 N p.Glu1134Ter BBC/BCFH
c.3472G>T 10 N p.Glu1158Ter EO
c.3607C>T 10 FS p.Arg1203Ter BCFH
c.3626T>G 10 N p.Leu1209Ter BBD
c.3640G>Tb 10 N p.Glu1214Ter BCFH
c.4065_4068del 10 FS p.Asn1355fs HBOC
c.4484+1G>A Intr S – BCFH
c.4801A>Td 15 N p.Lys1601Ter HBOC
c.5251C>T 19 N p.Arg1751Ter EO
c.5278-1G>C Intr S – BBD
c.5431C>T 22 N p.Gln1811Ter HBOS
c.5470_5477deld 23 FS p.Ile1824fs BBD/BCFH
BRCA2 c.961C>T 10 N p.Gln321Ter EO
c.1310_1313delb 10 FS p.Lys437Ilefs BCFH
c.1399A>T 10 N p.Lys467Ter EO
c.2806_2809deld 11 FS p.Asp936fs EO
c.3109C>Ta,d 11 FS p.Gln1037Ter BBD/BCFH
c.5682C>Aa 11 N p.Tyr1894Ter BCFH
c.7007G>Ta 14 S p.Arg2336Leu MBC
c.8504C>G 20 N p.Ser2835Ter BCFH
c.8517C>A 20 N p.Tyr2839Ter BCFH
c.9100C>T 23 N p.Gln3034Ter EO
c.9117G>A 23 Syn p.Pro3039= EO

BBC, bilateral breast cancer; BCFH, breast cancer family history; EO, early-onset breast cancer; FS, frameshift; HBOC, heredity breast and ovarian cancer syndrome family; Intr, Intron; MBC, male breast cancer; M, missense; N, nonsense; S, splice; Syn, synonymous.

Once found in Chinese people.

Hereditary mutation from the same family.

Deleterious missense mutation was carried in unrelated probands.

Mutation was carried in unrelated probands.

Novel variations (n=29).

Gene Location Exon Mutation type AA change Probands
BRCA1 c.1934del 10 FS p.Ser645fs EO
c.2957del 10 FS p.Ile986fs HBOC
c.3294del 10 FS p.Leu1098fs BCFH
c.3621del 10 FS p.Lys1207fs BBD
c.3859del 10 FS p.Glu1287fs BCFH
c.4013del 10 FS p.Lys1338fs EO
c.4676-1G>T Intr S – EO
c.5156del 18 FS p.Val1719fs BCFH
BRCA2 c.31del 2 FS p.Phe11fs EO
c.767_771del 9 FS p.Thr256fs EO
c.988del 10 FS p.Lys330fs BCFH
c.3364del 11 FS p.Gly1122fs EO
c.426-2A>T Intr S – HBOC
c.4410_4413del 11 FS p.Ile1470fs BCFH
c.5480del 11 FS p.Ile1827fs BCFH
c.5495del 11 FS p.Ser1832fs MBC/BCFH
c.5599_5602del 11 FS p.Tre1867fs EO
c.5718_5719del 11 FS p.Asn1906fs BBC
c.5753dela 11 FS p.His1918fs BCFH
c.6288_6289dela 11 FS p.Pro2096fs BCFH
c.6462_6465del 11 FS p.Tyr2154fs BBC/EO
c.6552del 11 FS p.Glu2184fs BCFH
c.6698_6699insTTTT 11 FS p.Ala2233fs HBOC
c.7178_7179del 14 FS p.Met2393fs BCFH
c.8019_8020insAT 18 N p.Lys2673fs BBD
c.8039_8040del 18 FS p.Asp2680fs EO
c.8367_8369 TAC>A 19 FS p.Tyr2789Ter EO
c.8400_8402delinsAAAA 19 FS p.Phe2801fs EO
c.9090dupa,b 23 FS p.Thr3030fs EO

BBC, bilateral breast cancer; BCFH, breast cancer family history; EO, early-onset breast cancer; FS, frameshift; HBOC, heredity breast and ovarian cancer syndrome family; Intr, Intron; MBC, male breast cancer; M, missense; N, nonsense; S, splice.

More than one carriers from the same family.

Inherited from paternal family, which doesn't have breast or ovarian cancer family history.

Information of VUS (n=10).

Gene Location AA change Proband Family history
BRCA1 c.446A>C p.E149A p.(Glu149Ala) EO –
c.1669A>C p.T557P p.(Thr557Pro) MBC (TNBC) Uterine cancer
c.4580A>T p.E1527V p.(Glu1527Val) BBC –
c.5156T>C p.V1719A p.(Val1710Ala) BBC –
c.5498T>A p.V1833E p.(Val1833Glu) MBC –
BRCA2 c.6875A>C p.E2292A p.(Glu2292Ala) BBC (TNBC) –
c.7811T>C p.L2604P p.(Leu260Pro) EO –
c.7967T>C p.L2656P p.(Leu2656Pro) EO –
c.8162T>C p.L2721P p.(Leu2721Pro) BBC one BC sister
c.9374T>C p.L3125P p.(Leu3125Pro) BBC (TNBCs) –

EO, early-onset breast cancer; MBC, male breast cancer; BBC, bilateral breast cancer; TNBC, triple negative breast cancer; VUS, variants of uncertain significance.

Familial mutations information (n=11).

Gene Location Family history features Chinesea
BRCA1 c.190T>Cb HBOC of maternal hereditary N
c.1660G>T BC sisters Y
c.2014A>T BCs of maternal history N
c.212+G>T BCs of maternal history Y
c.3640G>T BC sisters N
c.5431C>T HBOC of paternal hereditary N
BRCA2 c.988del BC sisters N (Novel)
c.1310_1313del BCs maternal history N
c.5753del BCs maternal history N (Novel)
c.6288_6289del BCs maternal history N (Novel)
c.9090dupb Paternal line N (Novel)

BC, breast cancer; HBOC, heredity breast and ovarian cancer syndrome family; Y, yes; N, no.

Whether mutation was once reported in Chinese.

Carried by the same proband.

TNM in three BRCA groups (n=410).

TNM (%)
Gene I (n=126) II (n=199) III (n=67) IV (n=18) P-trend
BRCA− 99 (28.9)a 171 (49.9) 57 (16.6) 16 (4.7) 0.33
BRCA1 17 (48.6) 14 (40) 3 (8.6) 1 (2.9)
BRCA2 10 (31.3) 14 (43.8) 7 (21.9) 1 (3.1)

TNM, tumor metastasis node.

Three carcinoma in situ patients were included.

HRs and HER2 comparisons between BRCA1 and BRCA− groups (n=376a).

Molecular markers BRCA1 (%) BRCA−b (%) Univariate P-value Multivariate P-value OR (95%CI)
ER
  <1% 24 (70.6) 120 (35.1) <0.0001 0.73 1.2 (0.4–3.3)
  ≥1% 10 (29.4) 222 (64.9) –
PR
  <1% 28 (82.4) 139 (40.6) <0.0001 0.003 6.3 (1.9–20.6)
  ≥1% 6 (17.6) 203 (59.4) –
HER2
  − 33 (97.1) 254 (74.3) 0.003 0.01 12.7 (1.7–95.6)
  + 1 (2.9) 88 (25.7) –

HRs, hormone receptors; ER, estrogen receptor; PR, progesterone receptor; HER2, humane epidermal growth factor receptor 2; OR, odd ratio; 95% CI, 95% confidence interval.

One BRCA- and 1 BRCA1 mutant participants were excluded because of unclear HER2 state.

Wild type BRCA.

HRs and HER2 comparisons between BRCA2 and BRCA- groups (n=373a).

Molecular markers BRCA2 (%) BRCA−b (%) Univariate P-value Multivariate P-value OR (95%CI)
ER
  <1% 2 (6.5) 120 (35.1) 0.001 0.09 0.2 (0.4–1.3)
  ≥1% 29 (93.5) 222 (64.9) –
PR
  <1% 4 (12.9) 139 (40.6) 0.002 0.41 0.6 (0.2–2.2)
  ≥1% 27 (87.1) 203 (59.4) –
HER-2
  − 30 (96.8) 254 (74.3) 0.005 0.03 9.4 (1.3–71.0)
  + 1 (3.2) 88 (25.7)

HRs, hormone receptors; ER, estrogen receptor; PR, progesterone receptor; HER2, humane epidermal growth factor receptor 2.

One BRCA- participant was excluded because of the unclear HER2 state.

Wild type BRCA.

Molecular types comparison in three BRCA groups (n=409a).

Gene TNBC (%) HR+b (%) P-value
BRCA-c 85 (24.8) 258 (75.2) –
BRCA1 24 (68.6) 11 (31.4) <0.0001
BRCA2 2 (6.5) 29 (93.5) 0.02
Total 111 298

TNBC, triple negative breast cancer; HRs+, Hormone receptors positive.

Thirteen participants were excluded because of HER2 overexpress type with no BRCA mutation carriers.

ER positive or/and PR positive regardless of HER2 state.

Wild type BRCA.