Y Xie

Association between CYP2D6 *10 genotype and survival of breast cancer patients receiving tamoxifen treatment

BACKGROUND Human cytochrome P450 2D6 (CYP2D6) genotype may affect the efficacy of tamoxifen treatment in Caucasian women with breast cancer. The most common polymorphism of CYP2D6 in Chinese women is variant 10 (188 C to T). PATIENTS AND METHODS Tamoxifen and 4-hydroxytamoxifen (4OHtam) were measured in the serum of 37 women with breast cancer who were receiving tamoxifen treatment. The association between CYP2D6 *10 genotype and survival was determined in a cohort of 293 women with breast cancer who received tamoxifen (n = 152) or who did not (n = 141). RESULTS The serum 4OHtam concentrations were significantly lower in women with the CYP2D6 *10 homozygous variant T/T genotype than in those with the homozygous wild-type C/C genotype (P = 0.04). Among tamoxifen-treated women, women with the T/T genotype had a significantly worse disease-free survival (DFS) than those with the C/C or C/T genotype, and the T/T genotype remained an independent prognostic factor of DFS in multivariate analysis (hazard ratio = 4.7; 95% confidence interval = 1.1-20.0; P = 0.04). Among women who did not receive tamoxifen, there was no significant association between CYP2D6 *10 genotype and survival. CONCLUSION In tamoxifen-treated patients, women with the CYP2D6 *10 T/T genotype have a lower 4OHtam level in the serum and a worse clinical outcome.

Promoter methylation of BRCA1 in triple-negative breast cancer predicts sensitivity to adjuvant chemotherapy

BACKGROUND BRCA1 function is inactivated through BRCA1 promoter methylation in a substantial number of triple-negative breast cancers. We investigated the impact of BRCA1-methylation status on the efficacy of adjuvant chemotherapy in patients with triple-negative breast cancer or with non-triple-negative breast cancer. METHODS BRCA1 promoter methylation was assessed in 1163 unselected breast cancer patients. Methylation was evaluated using a methylation-specific PCR (MSP) assay. RESULTS In the subgroup of 167 triple-negative breast cancer patients who received adjuvant chemotherapy, patients with BRCA1-methylated tumors had a superior 10-year disease-free survival (DFS)(78% versus 55%, P = 0.009) and 10-year disease-specific survival (DSS) (85% versus 69%, P = 0.024) than those with BRCA1-unmethylated tumors, and BRCA1 methylation was an independent favorable predictor of DFS and DSS in a multivariate analysis in this subgroup [DFS: hazard ratio (HR) = 0.45; 95% confidence interval (CI) 0.24-0.84; P = 0.019; DSS: HR = 0.43; 95% CI = 0.19-0.95; P = 0.044]. In contrast, in 675 non-triple-negative breast cancer patients who received adjuvant chemotherapy, BRCA1 methylation was an unfavorable predictor of DFS and DSS in univariate analysis (DFS: HR = 1.56; 95% CI 1.16-2.12; P = 0.003; DSS: HR = 1.53; 95% CI = 1.05-2.21; P = 0.026). CONCLUSIONS Triple-negative breast cancer patients with BRCA1-methylated tumors are sensitive to adjuvant chemotherapy and have a favorable survival compared with patients with BRCA1-unmethylated triple-negative tumors.

Protein phosphatase PP6 is required for homology-directed repair of DNA double-strand breaks

DNA double-strand breaks (DSBs) are among the most lethal lesions associated with genome stability which, when destabilized, predisposes organs to cancers. DSBs are primarily fixed either with little fidelity by non-homologous end joining (NHEJ) repair or with high fidelity by homology-directed repair (HDR). The phosphorylated form of H2AX on serine 139 (g-H2AX) is a marker of DSBs. In this study, we explored if the protein phosphatase PP6 is involved in DSB repair by depletion of its expression in human cancer cell lines, and determined PP6 expression in human breast cancer tissues by immunohistochemistry staining. We found that bacterially-produced PP6c (the catalytic subunit of PP6)-containing heterotrimeric combinations exhibit phosphatase activity against g-H2AX in the in vitro phosphatase assays. Depletion of PP6c or PP6R2 led to persistent high levels of g-H2AX after DNA damage and a defective HDR. Chromatin immunoprecipitation assays demonstrated that PP6c was recruited to the region adjacent to the DSB sites. Expression of PP6c, PP6R2, and PP6R3 in human breast tumors was significantly lower than those in benign breast diseases. Taken together, our results suggest that g-H2AX is a physiological substrate of PP6, and PP6 is required for HDR and its expression may harbor a protective role during the development of breast cancer.

Association between the HER2 Ile655Val Polymorphism and Response to Trastuzumab in Women with Operable Primary Breast Cancer

BACKGROUND Human epidermal growth factor receptor 2 (HER2) Ile655Val polymorphism may affect the efficacy of trastuzumab treatment of breast cancer. PATIENTS AND METHODS HER2 Ile655Val polymorphism was determined in 4167 patients with primary breast cancer using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. We investigated the associations between the HER2 Ile655Val polymorphism and clinical outcomes in women with HER2-negative breast cancer and with HER2-positive breast cancer who received trastuzumab or who did not. RESULTS At a median follow-up of 44 months, HER2 Ile655Val polymorphism was not significantly associated with survival either in the entire study population of 4167 patients or in 2976 HER2-negative breast cancer patients. Among 816 HER2-positive patients who received adjuvant chemotherapy and/or endocrine therapy without trastuzumab treatment, patients with the Val/Ile or the Val/Val genotype had a significantly worse disease-free survival (DFS) and distant DFS (DDFS) than those with the Ile/Ile genotype (DFS, adjusted hazard ratio [HR] 1.5; 95% confidence interval [CI] 1.0-2.3; P = 0.037; DDFS, adjusted HR 1.9; 95% CI 1.2-2.9 P = 0.005). In contrast, among 212 HER2-positive patients who received chemotherapy in combination with trastuzumab treatment, patients with the Val/Ile or the Val/Val genotype had a significantly better DFS and DDFS than those with the Ile/Ile genotype (5-year DFS, 100% versus 83%; P = 0.008; 5-year DDFS, 100% versus 89%; P = 0.031). CONCLUSIONS HER2 Ile655Val polymorphism affects the function of HER2 gene only restricted in HER2-positive breast cancers. HER2-positive breast cancer patients with the Val variant have an aggressive phenotype, but are sensitive to trastuzumab treatment.

Prevalence of BRCA1 mutations and responses to neoadjuvant chemotherapy among BRCA1 carriers and non-carriers with triple-negative breast cancer

BACKGROUND The frequency of BRCA1 germline mutations among Chinese women with triple-negative breast cancer is unclear, and the association between BRCA1 mutations and the response to neoadjuvant chemotherapy in women with triple-negative breast cancer has not been determined. PATIENTS AND METHODS Nine hundred and fifty-six triple-negative breast cancer patients were treated at our institute between 2003 and 2012; we tested the BRCA1/2 mutations for 956 patients and 953 patients in this cohort, respectively. Among the 956 patients, 652 patients received neoadjuvant chemotherapy. RESULTS In this cohort, 7.1% (68/956) and 2.3% (22/953) of patients carried a BRCA1 or BRCA2 mutation, respectively. The BRCA1/2 mutation rates were 10.5% and 3.0% among the patients who were diagnosed at or before the age of 50 in this cohort, respectively. The pCR (pathologic complete response) rate was 31.6% in the 652 patients who received neoadjuvant chemotherapy. BRCA1 carriers had a significantly higher pCR rate than non-carriers (BRCA1 carriers versus non-carriers, 53.8% versus 29.7%, P < 0.001). Among women treated with anthracycline with or without taxane regimens, the pCR rate was 57.1% for BRCA1 carriers, 29.0% for non-carriers (P < 0.001); among women treated with taxane regimens, the pCR rate was 40.0% for BRCA1 carriers, 32.9% for non-carriers (P = 0.73). At a median follow-up of 43 months, the recurrence-free survival was similar between BRCA1 carriers and non-carriers among the 947 patients of this study (adjusted hazard ratio = 0.92; 95% confidence interval: 0.45-1.90; P = 0.82). CONCLUSIONS Chinese women with triple-negative breast cancer who are diagnosed at or before age of 50 are candidates for BRCA1 genetic testing. Among triple-negative breast cancer patients, BRCA1 carriers are more likely to respond to neoadjuvant anthracycline-based regimens than are non-carriers.

HER2 and response to anthracycline-based neoadjuvant chemotherapy in breast cancer

BACKGROUND The predictive role of human epidermal growth factor receptor 2 (HER2) to adjuvant anthracycline-based chemotherapy remains controversial. Here, we investigated the association between HER2 status and pathological response in breast cancer patients who received neoadjuvant anthracycline-based regimens. PATIENTS AND METHODS Women (n = 538) with operable primary breast cancer received neoadjuvant anthracycline-based chemotherapy. Pathological complete response (pCR) was defined as no invasive breast tumor cells in breast after completion of neoadjuvant chemotherapy. HER2 status was determined by immunohistochemistry and/or by fluorescence in situ hybridization in core biopsy breast cancer tissue obtained before initiation of neoadjuvant chemotherapy. RESULTS In this cohort of 538 patients, 23.9% of patients achieved a pCR in their breast. HER2-positive tumors had a lower rate of pCR than did HER2-negative tumors (14.7% versus 25.7%, P = 0.013); negative HER2 status remained as an independent favorable predictor of pCR after adjusted for age, estrogen receptor, progesterone receptor, tumor size, chemotherapy cycles, and tumor grade in a multivariate analysis (odds ratio = 3.14; 95% confidence interval = 1.60-6.16, P = 0.001). Furthermore, patients with a pCR had a higher 3-year disease-free survival (DFS) rate than did patients without a pCR (P = 0.007). CONCLUSION Women with HER2-negative breast cancers rather than HER2-positive tumors benefit from anthracycline-based neoadjuvant chemotherapy.

Expression of ER-a36, a novel variant of estrogen receptor a, and resistance to tamoxifen treatment in breast cancer.

CTRC-AACR San Antonio Breast Cancer Symposium: 2008 Abstracts Abstract #3037 Background: Recently, a 36 kDa variant of estrogen receptor a (ER-a66), ER-a36, has been identified and cloned. ER-a36 predominantly localizes on the plasma membrane and in the cytoplasm and mediates a membrane-initiated “nongenomic” signaling pathway. In this study, we investigated the association between ER-a36 expression and tamoxifen resistance in breast cancer patients. Methods: ER-a36 protein expression in tumors from 710 breast cancer patients with a median follow-up of 7.9 years was assessed using immunohistochemistry (IHC) assay. Survival curves were compared using the log-rank test and multivariate analysis was performed using Cox model. All statistical tests were two-sided. Results: Among the patients with ER-a66 positive tumors who received tamoxifen treatment (n=307), overexpression of ER-a36 was associated with poorer disease-free survival (DFS) and disease-specific survival (DSS) and remained as an unfavorable independent factor of survival in multivariate analyses (DFS: HR=2.27; 95% CI= 1.40 to 3.68; P =. 001; DSS: HR=2.42; 95% CI= 1.37 to 4.28; P = .002). In contrast, among patients with ER-a66 positive tumors who did not receive tamoxifen (n=129), ER-a36 expression was not associated with survival, indicating a correlation between ER-a36 expression and tamoxifen resistance. Furthermore, ER-a36 expression was not associated with survival in ER-a66 negative tumors whether the patients received tamoxifen (n=73) or not (n=149). Our in vitro experiments with MCF7/ER36 cells also confirmed that high ER-a36 expression resulted in tamoxifen resistance. Conclusions: Patients with ER-a66 positive tumors that also express high levels of ER-a36 are less likely to benefit from tamoxifen treatment. ER-a36 is an important predictive marker for tamoxifen therapy in ER-a66 positive breast cancer patients. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3037.

Associations between RAD51D germline mutations and breast cancer risk and survival in BRCA1/2-negative breast cancers

Background RAD51D is involved in DNA double-strand break repair by homologous recombination and plays an important role in the maintenance of genomic stability. The associations between RAD51D germline mutations and breast cancer risk and survival are not fully elucidated. Patients and methods RAD51D germline mutations were determined using a multigene panel in 7657 unselected breast cancer patients who were negative for BRCA1/2 germline mutations. The RAD51D recurrent mutation p.K91fs was screened in 7947 healthy controls by Sanger sequencing. Results A total of 29 cases (0.38%) carried deleterious RAD51D germline mutations among this cohort of 7657 unselected breast cancer patients. The RAD51D recurrent mutation p.K91fs was identified in 18 cases (0.24%) of these 7657 patients. In contrast, the p.K91fs mutation was found in 8 of 7947 healthy controls with a frequency of 0.10%. The RAD51D p.K91fs mutation was significantly associated with increased breast cancer risk in unselected breast cancer [odds ratio = 2.34, 95% confidence interval (CI) 1.02-5.38; P = 0.040]. RAD51D mutation carriers were diagnosed at a younger age (P = 0.006) and were more likely to be triple-negative breast cancer (P = 0.003), estrogen receptor negative (P = 0.005) and high-grade cancers (P = 0.023) than noncarriers. Furthermore, RAD51D mutation carriers had a significantly worse recurrence-free survival [unadjusted hazard ratio (HR) = 3.00, 95% CI 1.56-5.80; P = 0.001] and distant recurrence-free survival (unadjusted HR = 2.54, 95% CI 1.14-5.67; P = 0.023) than noncarriers. Conclusion The RAD51D recurrent mutation, p.K91fs, confers a moderately increased breast cancer risk, and RAD51D mutation carriers have an unfavorable survival compared with noncarriers.

Abstract 2988: A PP6-phosphatase complex is required for homology-directed repair of DNA double-strand breaks

DNA double-strand breaks (DSBs) are among the most lethal lesions associated with genome stability which, when destabilized, predisposes organs to cancers. DSBs are primarily fixed either with little fidelity by non-homologous end joining (NHEJ) repair or with high fidelity by homology-directed repair (HDR). The phosphorylated form of H2AX on serine 139 (g-H2AX) is a marker of DSBs. It is dephosphorylated by PP2A-like protein phosphatases (PP2A, PP4, and PP6). Here, we identify three novel PP6 subunits. Several PP6c (the catalytic subunit of PP6)-containing heterotrimeric complexes show phosphatase activities against g-H2AX in the in vitro phosphatase assays using bacterially-produced proteins, providing direct evidence that the PP6 holoenzyme is a PP6c-containing heterotrimeric complex. Furthermore, depletion of PP6c, PP6R2, or MyPT1 leads to defective HDR, and expression of phosphatase-inactive PP6c fails to rescue this defect. Chromatin immunoprecipitation assays demonstrated that PP6c is recruited to the region adjacent to the DSB sites. Expression of PP6R2, but not PP6c or MyPT1, is associated with improved overall survival in patients with breast cancer at five years (n=100, p=0.038). Taken together, our results suggest that the PP6-PP6R2-MyPT1 complex is required for HDR and PP6R2 is a potential therapeutic target. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2988.

Abstract P1-01-27: Level III and Interpectoral Lymph Nodes Involvement in Breast Cancer with Positive Axillary Lymph Nodes after Neoadjuvant Chemotherapy

Background: In the era of sentinel node biopsy (SNB), mainly aimed at local control, the axillary dissection (AD) was performed for breast cancer with positive node. It was argued that only level I/II lymph nodes dissection might be inadequate because nodes involvement was found in level III and interpectoral region when axillary node was positive. It is necessary to assess the extent of AD after modern preoperative axillary staging. The aim of this study was to investigate the incidence and associated factors of node involvement in level III/interpectoral region. Methods: A consecutive series of 338 core needle biopsy confirmed T0-2 invasive breast cancer cases were included in this study. Axillary node metastases were proved by ultrasound guided needle biopsy (NB) if ultrasonographic abnormal node was detected prior to SNB or by SNB if no abnormal node was detected. Cases of negative NB but positive SNB with image abnormal node were excluded. Prior to AD included level III and interpectoral lymph nodes, 4 to 8 cycles of neoadjuvant chemotherapy with anthrocyclin and/or taxane regimen were completed for each case. The chi-square test was used to determine the relation between level III/interpectoral lymph nodes metastases and clinicopathological factors. Multivariate logistic regression was analyzed for covariate selection. Results: A median of 19 axillary nodes was harvested per case (range: 5-46, average: 19.2). The pathologic complete response rate of axillary nodes was 35.3% (70/198) in NB positive subgroup. Level III and interpectoral lymph nodes were harvested in 76.9% (260/338) of cases (range: 1-8, average: 1.9, median: 1) and 49.7% (168/338) of cases (range: 1-10, average: 1.6, median: 1), respectively. The incidence of positive level III and interpectoral lymph nodes were 8.9% (30/338) and 8.9% (30/338), respectively. Node involvement of level III/interpectoral region was found in 13.3% (45/338) of these cases. The incidence of node involvement in level III/interpectoral region of NB positive subgroup (14.6%, 29/198) was not significantly higher compared with SNB positive subgroup (11.4%, 16/140, P=0.391). Increasing tumor size was significantly correlated with increasing likelihood of node involvement in level III/interpectoral region (T0-1: 6.3% vs T2: 16.7%, P=0.008). In SNB positive T0-1 subgroup, there was no positive node found in level III/interpectoral region. Multivariate analysis showed that tumor size was the only independent factors predicting node involvement in level III/interpectoral region (OR=3.488, 95%CI:1.427-8.528, P=0.006). Conclusions: The incidence of node involvement in level III/interpectoral region was 13.3% of nodes positive T0-2 breast cancer treated with neoadjuvant chemotherapy. Tumor size may be the predictor of node involvement in this region. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P1-01-27.