Thymidine kinase and thymidylate synthase in advanced breast cancer: response to tamoxifen and chemotherapy

Thymidylate synthase (TS) is a crucial target for 5-fluorouracil (5-FU) in the de novo pathway of pyrimidine synthesis, which is necessary for DNA synthesis. Thymidine kinase (TK) plays a key role in the complementary or alternative salvage pathway of pyrimidine synthesis in acute or pathological tissue stress. In the present study, the activity levels of TS and TK were determined in 257 primary breast tumors of patients who received tamoxifen as first-line systemic therapy after diagnosis of advanced disease. In 155 (60%) responding patients, the median response duration was 23 months for tumors with low TK activity, 15 months for tumors with intermediate TK activity, and 13 months for tumors with high TK activity (P = 0.003). In Cox multivariate analysis corrected for classical predictive factors including estrogen receptor and progesterone receptor, patients with intermediate and high levels of TK activity in their tumors showed a rapid disease progression (P = 0.0002) and an early death (P = 0.002) after start of tamoxifen treatment. Tumor TS activity levels were not significantly associated with the efficacy of tamoxifen treatment. In 121 patients who became resistant to tamoxifen or additional endocrine treatments and who received 5-FU-containing polychemotherapy, tumor TK activity was not significantly related to the efficacy of chemotherapy. Of the 13 patients with low tumor TS activity, only 1 (8%) responded favorably, whereas 46% (43 of 93) of those with intermediate and 73% (11 of 15) of those with high TS activity responded (P = 0.001). In Cox multivariate regression analysis in which TS was the only significant variable, intermediate and high TS activities were associated with a slow disease progression (P = 0.005) and prolonged survival (P = 0.016) on chemotherapy. In conclusion, for patients with recurrent breast cancer, high tumor TK activity is a significant marker of poor clinical outcome on tamoxifen therapy. Elevated tumor TS activity predicts a favorable outcome for 5-FU-containing polychemotherapy when applied after tumor progression on endocrine therapy

The prognostic value of polymorphonuclear leukocyte elastase in patients with primary breast cancer

A variety of serine proteases, including urokinase-type plasminogen activator (uPA), plasmin,and polymorphonuclear leukocyte elastase (PMN-E), have been implicated in the processes of tumor cell invasion and metastasis. Besides degrading of matrix proteins, PMN-E has been shown to be able to cleave and inactivate plasminogen activator inhibitor-1 (PAI-1), the main inhibitor of uPA, and alpha2-antiplasmin, the natural inhibitor of plasmin, thus enabling an uncontrolled matrix degradation by the fibrinolytic enzymes. Because only limited data are available on a relationship between the tumor level of PMN-E and prognosis in primary breast cancer patients, in the present study we have measured with an ELISA the levels of PMN-E (in complex with alpha1-proteinase inhibitor) in cytosolic extracts of 1143 primary breast tumors. Levels of complexed PMN-E have been correlated with the lengths of metastasis-free survival (MFS), relapse-free survival, and overall survival, and a comparison was made with data previously obtained for uPA and PAI-1. Our results show that patients with a high PMN-E level in their primary tumor had a rapid relapse and an early death compared with patients with a low tumor level of PMN-E. This held true for node-negative and node-positive subgroups of patients as well. The relationship of PMN-E with a poor prognosis was especially obvious during short-term follow-up (0-60 months). In Cox multivariate regression analysis, corrected for the traditional prognostic factors, PMN-E was an independent prognostic factor, and high levels of PMN-E were associated with a poor MFS [hazard ratio (HR), 1.63; 95% confidence interval (CI), 1.23-2.16; P < 0.001], relapse-free survival (HR, 1.45; 95% CI, 1.10-1.89; P = 0.01), and overall survival (HR, 1.64; 95% CI, 1.20-2.23; P = 0.003). Furthermore, in all three multivariate models, PMN-E still added significantly to the model after the additional inclusion of the uPA. PMN-E was an independent prognostic factor for MFS even in the multivariate analysis including the traditional clinical prognostic factors and the strong established biochemical prognostic factors uPA and PAI-1. Our present study suggests that PMN-E is associated with breast cancer metastasis, and knowledge of the tumor PMN-E status might be helpful in selecting the appropriate individualized (adjuvant) treatment for patients with breast cancer

Multivariate meta-analysis of individual participant data helped externally validate the performance and implementation of a prediction model

Objectives Our aim was to improve meta-analysis methods for summarizing a prediction model's performance when individual participant data are available from multiple studies for external validation. Study Design and Setting We suggest multivariate meta-analysis for jointly synthesizing calibration and discrimination performance, while accounting for their correlation. The approach estimates a prediction model's average performance, the heterogeneity in performance across populations, and the probability of "good" performance in new populations. This allows different implementation strategies (e.g., recalibration) to be compared. Application is made to a diagnostic model for deep vein thrombosis (DVT) and a prognostic model for breast cancer mortality. Results In both examples, multivariate meta-analysis reveals that calibration performance is excellent on average but highly heterogeneous across populations unless the model's intercept (baseline hazard) is recalibrated. For the cancer model, the probability of "good" performance (defined by C statistic ≥ 0.7 and calibration slope between 0.9 and 1.1) in a new population was 0.67 with recalibration but 0.22 without recalibration. For the DVT model, even with recalibration, there was only a 0.03 probability of "good" performance. Conclusion Multivariate meta-analysis can be used to externally validate a prediction model's calibration and discrimination performance across multiple populations and to evaluate different implementation strategies

The clinical significance of androgen receptors in breast cancer and their relation to histological and cell biological parameters

To analyse the clinical significance of the presence of androgen receptors (AR) in breast carcinomas, clinical and histological parameters of 153 primary breast carcinomas (median follow-up 46 months) were examined. Oestrogen (ER) and progesterone receptor (PR) levels were determined in cytosol preparations using enzyme immunoassay assays and in cryostat sections by immunohistochemistry. AR and Ki-67 levels were only determined immunohistochemically. Data were analysed by uni- and multivariate models. 94/153 (61%) breast carcinomas were ER+ PR+ AR+, while 14 cases were only positive for AR. All grade III tumours (n = 17) were steroid receptor negative and 14 (76%) of these cases demonstrated high Ki-67 values suggestive of more aggressive behaviour. Strikingly, 14 ductal carcinomas negative for ER and PR were positive for AR. In univariate analysis, AR as well as ER, tumour size, lymph node status, grade and Ki-67 proved to be significant prognostic factors for disease free survival (DFS). Multivariate analysis, however, showed lymph node status, tumour size and ER status to be the only independent prognostic factors for DFS within this model. We conclude that simple histological and cell biological parameters, including AR, can be used to select high- and low-risk patients at the time of primary surgery and can provide valuable information on treatment options

RNA expression of breast cancer resistance protein, lung resistance-related protein, multidrug resistance-associated proteins 1 and 2, and multidrug resistance gene 1 in breast cancer: correlation with chemotherapeutic response

PURPOSE: The aim of this study was to investigate whether expression of particular drug resistance genes in primary operable breast cancer correlates with response to first-line chemotherapy in advanced disease. EXPERIMENTAL DESIGN: We determined mRNA levels of BCRP, LRP, MRP1, MRP2, and MDR1 in 59 primary breast tumor specimens of patients who

Bcar1/p130Cas protein and primary breast cancer: prognosis and response to tamoxifen treatment

BACKGROUND: The product of the Bcar1/p130Cas (breast cancer resistance/p130Crk-associated substrate) gene causes resistance to antiestrogen drugs in human breast cancer cells in vitro. To investigate its role in clinical breast cancer, we determined the levels of Bcar1/p130Cas protein in a large series of primary breast carcinomas. METHODS: We measured Bcar1/p130Cas protein in cytosol extracts from 937 primary breast carcinomas by western blot analysis. The levels of Bcar1/p130Cas protein were tested for associations and trends against clinicopathologic and patient characteristics, the lengths of relapse-free survival and overall survival (n = 775), and the efficacy of first-line treatment with tamoxifen for recurrent or metastatic disease (n = 268). RESULTS: Bcar1/p130Cas levels in primary tumors were associated with age/menopausal status and the levels of estrogen receptor and progesterone receptor. In univariate survival analysis, higher Bcar1/p130Cas levels were associated with poor relapse-free survival and overall survival (both two-sided P =.04; log-rank test for trend). In multivariate analysis, a high level of Bcar1/p130Cas was independently associated with poor relapse-free survival and overall survival. The response to tamoxifen therapy in patients with recurrent disease was reduced in patients with primary tumors that expressed high levels of Bcar1/p130Cas. In multivariate analysis for response, Bcar1/p130Cas was independent of classical predictive factors, such as estrogen receptor status, age/menopausal status, disease-free interval, and dominant site of relapse. CONCLUSION: Patients with primary breast tumors expressing a high level of Bcar1/p130Cas protein appear to experience more rapid disease recurrence and have a greater risk of (intrinsic) resistance to tamoxifen therapy. Thus, measurement of Bcar1/p130Cas may provide useful prognostic information for patients with primary or metastatic breast cancer

GATA3 mRNA expression, but not mutation, associates with longer progression-free survival in ER-positive breast cancer patients treated with first-line tamoxifen for recurrent disease

In breast cancer, GATA3 mutations have been associated with a favorable prognosis and the response to neoadjuvant aromatase inhibitor treatment. Therefore, we investigated whether GATA3 mutations predict the outcome of tamoxifen treatment in the advanced setting. In a retrospective study consisting of 235 hormone-naive patients with ER-positive breast cancer who received tamoxifen as first-line treatment for recurrent disease, GATA3 mutations (in 14.0% of patients) did not significantly associate with either the overall response rate (ORR) or with the length of progression-free survival (PFS) after the start of tamoxifen therapy. Interestingly, among 148 patients for whom both mutation and mRNA expression data were available, GATA3 mutations associated with an increased expression of GATA3. However, only 23.7% of GATA3 high tumors had a mutation. Evaluation of the clinical significance of GATA3 mRNA revealed that it was associated with prolonged PFS, but not with the ORR, also in multivariate analysis. Thus, GATA3 mRNA expression, but not GATA3 mutation, is an independent predictor of prolonged PFS in ER-positive breast cancer patients who received first-line tamoxifen for recurrent disease. Besides GATA3 mutation, other mechanisms must exist that underlie increased GATA3 levels

BCAR4 induces antioestrogen resistance but sensitises breast cancer to lapatinib

Background: High BCAR4 and ERBB2 mRNA levels in primary breast cancer associate with tamoxifen resistance and poor patient outcome. We determined whether BCAR4 expression sensitises breast cancer cells to lapatinib, and identifies a subgroup of patients who possibly may benefit from ERBB2-targeted therapies despite having tumours with low ERBB2 expression. Methods :Proliferation assays were applied to determine the effect of BCAR4 expression on lapatinib treatment. Changes in cell signalling were quantified with reverse-phase protein microarrays. Quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) of ERBB2 and BCAR4 was performed in 1418 primary breast cancers. Combined BCAR4 and ERBB2 mRNA levels were evaluated for association with progression-free survival (PFS) in 293 oestrogen receptor-α (ER)-positive patients receiving tamoxifen as first-line monotherapy for recurrent disease.Results:BCAR4 expression strongly sensitised ZR-75-1 and MCF7 breast cancer cells to the combination of lapatinib and antioestrogens. Lapatinib interfered with phosphorylation of ERBB2 and its downstream mediators AKT, FAK, SHC, STAT5, and STAT6. Reverse transcriptase-PCR analysis showed that 27.6% of the breast cancers were positive for BCAR4 and 22% expressed also low levels of ERBB2. The clinical significance of combining BCAR4 and ERBB2 mRNA status was underscored by the finding that the group of patients having BCAR4-positive/ERBB2-low-expressing cancers had a shorter PFS on tamoxifen treatment than the BCAR4-negative group. Conclusion :This study shows that BCAR4 expression identifies a subgroup of ER-positive breast cancer patients without overexpression of ERBB2 who have a poor outcome and might benefit from combined ERBB2-targeted and antioestrogen therapy

Identification of a putative protein-profile associating with tamoxifen therapy-resistance in breast cancer

Tamoxifen-resistance is a major cause of death in patients with recurrent breast cancer. Current clinical parameters can correctly predict therapy response in only half of the treated patients. Identification of proteins that associate with tamoxifen-resistance is a first step towards better response prediction and tailored treatment of patients. In the present study we aimed to identify putative protein biomarkers indicative of tamoxifen therapy-resistance in breast cancer, using nanoLC-FTICR MS. Comparative proteome analysis was performed on ~5,500 pooled tumor cells obtained through laser capture microdissection from two independently processed data sets (n=24 and n=27) of tamoxifen therapy-sensitive and -resistant tumors. Peptide and protein identifications were acquired by matching mass and elution time features to information in previously generated accurate mass and time tag reference data bases. A total of 17,263 unique peptides were identified that corresponded to 2,556 non-redundant proteins identified with >=2 peptides. From this total, 1,713 protein

High miR-26a and low CDC2 levels associate with decreased EZH2 expression and with favorable outcome on tamoxifen in metastatic breast cancer

For patients with metastatic breast cancer, we previously described that increased EZH2 expression levels were associated with an adverse outcome to tamoxifen therapy. Main objective of the p