http://repub.eur.nl/res/aut/4732/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/22064/ 2010-01-01T00:00:00Z Abstract The purpose of this study is to investigate EZH2 in a large series of breast cancer patients for its prognostic and predictive value, and to evaluate its functional role in treatment response in vitro. EZH2 levels were measured using quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) in primary breast cancer specimens and related to clinicopathologic factors and disease outcome. EZH2 expression was downregulated with siRNAs in MCF7, to assess expression alterations of putative EZH2 downstream genes and to determine cell numbers after treatment with the anti-estrogen ICI 164384. In 688 lymph node-negative patients who did not receive adjuvant systemic therapy, EZH2 was not significantly correlated with metastasis-free survival (MFS). In 278 patients with advanced disease treated with first-line tamoxifen monotherapy, the tertile with highest EZH2 levels was associated with the lowest clinical benefit (OR = 0.48; P = 0.02) and with a shorter progression-free survival (PFS) in both univariate (HR = 1.80; P < 0.001) and multivariate analysis, including traditional factors (HR = 1.61; P = 0.004). In vitro, EZH2 silencing in MCF7 caused a 38% decrease in cell numbers (P < 0.001) whereas ICI 164384 treatment resulted in a 25% decrease (P < 0.001) compared to controls. Combining EZH2 silencing with ICI treatment reduced cell numbers with 67% (P < 0.001) compared to control conditions. EZH2 downregulation was associated with an almost two-fold upregulation of the estrogen receptor alpha (ER) (P = 0.001). In conclusion, EZH2 has no prognostic value in breast cancer. High levels of EZH2 are associated with poor outcome to tamoxifen therapy in advanced breast cancer. Downregulated EZH2 leads to upregulation of the ER and better response to anti-estrogens. http://repub.eur.nl/res/pub/30212/ 2008-09-01T00:00:00Z Purpose: We previously discovered an extracellular matrix (ECM) gene cluster associated with resistance to first-line tamoxifen therapy of patients with metastatic breast cancer. In this study, we determined whether the six individual ECM genes [collagen 1A1 (COL1A1), fibronectin 1 (FN1), lysyl oxidase (LOX), secreted protein acidic cysteine-rich (SPARC), tissue inhibitor of metalloproteinase 3 (TIMP3), and tenascin C (TNC)] were associated with treatment response, prognosis, or both. Experimental Design: In 1,286 primary breast tumors, mRNA expression (quantitative realtime PCR) was related to clinicopathologic factors and disease outcome in univariate and multivariate analysis including traditional factors. Results: TIMP3, FN1, LOX, and SPARC expression levels (continuous variables) were significantly associated with distant metastasis-free survival (MFS) in 680 lymph node-negative untreated patients (P < 0.03). Using a calculated linear prognostic score, these patients were evenly divided into five prognostic groups with a significant difference in 10-year MFS of ∼40% between the two extreme prognostic groups. Furthermore, high TNC expression as continuous variable was associated with (a) shorter MFS in 139 estrogen receptor-positive and lymph node-positive patients who received adjuvant tamoxifen therapy (hazard ratio, 1.53; P = 0.001), and (b) no clinical benefit (odds ratio, 0.81; P = 0.035) and shorter progression-free survival (hazard ratio, 1.19; P = 0.002) in 240 patients in whom recurrence was treated with tamoxifen as first-line monotherapy. These results were also significant in multivariate analyses. Conclusion: FN1, LOX, SPARC, and TIMP3 expression levels are associated with the prognosis of patients with breast cancers, whereasTNC is associated with resistance to tamoxifen therapy. Further validation and functional studies are necessary to determine the use of these ECM genes in decisions regarding treatment and whether they can serve as targets for therapy. http://repub.eur.nl/res/pub/36221/ 2007-02-20T00:00:00Z Purpose: A HOXB13-to-IL17BR expression ratio was previously identified to predict clinical outcome of breast cancer patients treated with adjuvant tamoxifen. However, this ratio may predict a tumor's response to tamoxifen, its intrinsic aggressiveness, or both. Patients and Methods: We have measured the HOXB13 and IL17BR expression levels by real-time polymerase chain reaction in 1,252 primary breast tumor specimens. Expression levels were normalized to housekeeper gene levels and related to clinicopathologic factors for all patients. The primary objective of this study was to determine the relationship of a HOXB13-to-IL17BR ratio with tumor aggressiveness and/or with response to tamoxifen therapy in estrogen receptor (ER) -positive disease. We selected ER-positive tumors, and clinical end points for the HOXB13-to-IL17BR ratio were disease-free survival (DFS) in patients with primary breast cancer (N = 619) and progression-free survival (PFS) in patients with recurrent breast cancer treated with first-line tamoxifen monotherapy (N = 193). The odds ratio (OR) and hazard ratio (HR) and their 95% CI were calculated, and all P values were two-sided. Results: The HOXB13-to-IL17BR ratio was significantly associated with DFS and PFS. In multivariate analysis, HOXB13-to-IL17BR ratio expression levels were associated with a shorter DFS for node-negative patients only. Corrected for traditional predictive factors, the dichotomized HOXB13-to-IL17BR ratio was the strongest predictor in multivariate analysis for a poor response to tamoxifen therapy (OR = 0.16; 95% CI, 0.06 to 0.45; P < .001) and a shorter PFS (HR = 2.97; 95% CI, 1.82 to 4.86; P < .001). Conclusion: High HOXB13-to-IL17BR ratio expression levels associate with both tumor aggressiveness and tamoxifen therapy failure. http://repub.eur.nl/res/pub/14026/ 2006-07-31T00:00:00Z BACKGROUND: The treatment of ovarian cancer is hindered by intrinsic or acquired resistance to platinum-based chemotherapy. The aim of this study is to determine the frequency of mismatch repair (MMR) inactivation in ovarian cancer and its association with resistance to platinum-based chemotherapy. METHODS: We determined, microsatellite instability (MSI) as a marker for MMR inactivation (analysis of BAT25 and BAT26), MLH1 promoter methylation status (methylation specific PCR on bisulfite treated DNA) and mRNA expression of MLH1, MSH2, MSH3, MSH6 and PMS2 (quantitative RT-PCR) in 75 ovarian carcinomas and eight ovarian cancer cell lines RESULTS: MSI was detected in three of the eight cell lines i.e. A2780 (no MLH1 mRNA expression due to promoter methylation), SKOV3 (no MLH1 mRNA expression) and 2774 (no altered expression of MMR genes). Overall, there was no association between cisplatin response and MMR status in these eight cell lines.Seven of the 75 ovarian carcinomas showed MLH1 promoter methylation, however, none of these showed MSI. Forty-six of these patients received platinum-based chemotherapy (11 non-responders, 34 responders, one unknown response). The resistance seen in the eleven non-responders was not related to MSI and therefore also not to MMR inactivation. CONCLUSION: No MMR inactivation was detected in 75 ovarian carcinoma specimens and no association was seen between MMR inactivation and resistance in the ovarian cancer cell lines as well as the ovarian carcinomas. In the discussion, the results were compared to that of twenty similar studies in the literature including in total 1315 ovarian cancer patients. Although no association between response and MMR status was seen in the primary tumor the possible role of MMR inactivation in acquired resistance deserves further investigation. http://repub.eur.nl/res/pub/10121/ 2003-01-01T00:00:00Z PURPOSE: In recent studies, we showed that TP53 gene mutation or high levels of cytosolic vascular endothelial growth factor (VEGF) in estrogen receptor (ER)-alpha-positive primary breast tumors predict a poor disease outcome for patients treated with first-line tamoxifen for advanced disease. Mutant TP53 may up-regulate VEGF, whereas, on the other hand, wild-type TP53 may decrease VEGF production. EXPERIMENTAL DESIGN: In the present study, we aimed to assess the combined predictive value of TP53 gene mutation and VEGF status of 160 advanced breast cancer patients with ER-positive tumors who were treated with tamoxifen (median follow-up from start of tamoxifen treatment, 64 months). To assess TP53 gene mutation status, the entire open reading frame was sequenced; for VEGF status, an ELISA was used. RESULTS: In univariate analysis, both TP53 gene mutation (28% of the tumors) and a VEGF level above the median value were significantly associated with a short progression-free survival, post-relapse overall survival, and a poor rate of response to tamoxifen. In Cox multivariate regression analysis including the traditional predictive factors, the addition of TP53 gene mutation and VEGF status, alone or in combination, significantly predicted a poor efficacy of tamoxifen treatment. When the two factors were combined, a significantly decreased odds ratio was seen for the rate of response (odds ratio, 0.27). Similarly, an increased hazard ratio (HR) was seen for progression-free survival (HR, 2.32) and post-relapse overall survival (HR, 1.68) in the group with mutant TP53 and high VEGF compared with the group with both risk factors absent. CONCLUSIONS: Combined TP53 gene mutation status and high VEGF levels of ER-positive primary breast tumors independently predict a poor course of the disease of patients with advanced breast cancer treated with tamoxifen. These patients, having unfavorable tumor characteristics, might benefit more from other types of (individualized) treatment protocols. http://repub.eur.nl/res/pub/9358/ 2000-01-01T00:00:00Z TP53 has been implicated in regulation of the cell cycle, DNA repair, and apoptosis. We studied, in primary breast tumors through direct cDNA sequencing of exons 2-11, whether TP53 gene mutations can predict response in patients with advanced disease to either first-line tamoxifen therapy (202 patients, of whom 55% responded) or up-front (poly)chemotherapy (41 patients, of whom 46% responded). TP53 mutations were detected in 90 of 243 (37%) tumors, and one-fourth of these mutations resulted in a premature termination of the protein. The mutations were observed in 32% (65 of 202) of the primary tumors of tamoxifen-treated patients and in 61% (25 of 41) of the primary tumors of the chemotherapy patients. TP53 mutation was significantly associated with a poor response to tamoxifen [31% versus 66%; odds ratio (OR), 0.22; 95% confidence interval (CI), 0.12-0.42; P < 0.0001]. Patients with TP53 gene mutations in codons that directly contact DNA or with mutations in the zinc-binding domain loop L3 showed the lowest response to tamoxifen (18% and 15% response rates, respectively). TP53 mutations were related, although not significantly, to a poor response to up-front chemotherapy (36% versus 63%; OR, 0.34; 95% CI, 0.09-1.24). In multivariate analysis for response including the classical parameters age and menopausal status, disease-free interval, dominant site of relapse, and levels of estrogen receptor and progesterone receptor, TP53 mutation was a significant predictor of poor response in the tamoxifen-treated group (OR, 0.29; 95% CI, 0.13-0.63; P = 0.0014). TP53-mutated and estrogen receptor-negative (<10 fmol/mg protein) tumors appeared to be the most resistant phenotype. Interestingly, the response of patients with TP53 mutations to chemotherapy after tamoxifen was not worse than that of patients without these mutations (50% versus 42%; OR, 1.35, nonsignificant). The median progression-free survival after systemic treatment was shorter for patients with a TP53 mutation than for patients with wild-type TP53 (6.6 and 0.6 months less for tamoxifen and up-front chemotherapy, respectively). In conclusion, TP53 gene mutation of the primary tumor is helpful in predicting the response of patients with metastatic breast disease to tamoxifen therapy. The type of mutation and its biological function should be considered in the analyses of the predictive value of TP53.