Prospective evaluation of serum tissue inhibitor of metalloproteinase 1 and carbonic anhydrase IX in correlation to circulating tumor cells in patients with metastatic breast cancer.

INTRODUCTION: Circulating tumor cells (CTCs) reflect aggressive tumor behavior by hematogenous tumor cell dissemination. The tissue inhibitor of metalloproteinase 1 (TIMP-1) plays a role in tissue invasion and is also involved in angiogenesis, abrogation of apoptosis and in chemoresistance. Carbonic anhydrase IX (CAIX) is a metalloenzyme involved in cell adhesion, growth and survival of tumor cells. The aim of the study was to investigate whether serum concentrations of TIMP-1 and CAIX are associated with the detection of CTC in metastatic breast cancer. METHODS: Blood was obtained in a prospective multicenter setting from 253 patients with metastatic breast cancer at the time of disease progression. Serum TIMP-1 and CAIX were determined using commercial ELISA-kits (Oncogene Science). CTC were detected with the CellSearch system (Veridex). RESULTS: Five or more CTCs were detected in 122 patients out of 245 evaluable patients (49.8%). Out of 253 metastatic patients 70 (28%) had serum TIMP-1 levels above 454 ng/mL. Serum CAIX was elevated above 506 ng/mL in 90 (35%) patients. Both serum markers had prognostic significance. Median progression free survival (PFS) was 7.2 months with elevated TIMP-1 vs. 11.4 months with non-elevated levels (p < 0.01). OS was 11.5 vs. 19.1 months (p < 0.01). Median PFS was 7.5 months with elevated CAIX vs. 11.7 months with non-elevated levels (p < 0.01), overall survival (OS) was 13.4 months vs. 19.1 months (p < 0.01). In patients with five or more CTCs, serum levels were above the cut-off for CAIX in 47% vs. 25% in those with less than five CTCs (p = 0.01). For TIMP-1, 37% patients with five or more CTCs had elevated serum levels and 17% of patients with less than five CTCs (p = 0.01). Including TIMP-1, CAIX, CTC and established prognostic factors in the multivariate analysis, the presence of CTCs, the therapy line and elevated CAIX remained independent predictors of OS. CONCLUSIONS: Elevated serum levels of the invasion markers TIMP-1 and CAIX in metastatic breast cancer are prognostic markers and are associated with the presence of CTCs. Whether increased secretion of TIMP-1 and/or CAIX might directly contribute to tumor cell dissemination remains to be elucidated in further investigations. TRIAL REGISTRATION: Current Controlled Trials: ISRCTN59722891.

Comparison of estrogen and progesterone receptor status of circulating tumor cells and the primary tumor in metastatic breast cancer patients.

OBJECTIVES: The expression of predictive markers including the estrogen (ER) and progesterone receptor (PR) expression can change during the course of the disease. Therefore, reassessment of these markers at the time of disease progression might help to optimize treatment decisions. Metastatic tissue may be difficult to obtain for repeated analysis. In this context, characterization of circulating tumor cells (CTCs) could be of relevance. It was the purpose of the present study (1) to reevaluate the ER/PR expression by CTCs and (2) to compare the hormone receptor status expression profile of CTCs with the primary tumor. METHODS: We evaluated 193 blood samples from metastatic breast cancer patients at the time of first diagnosis of metastatic disease or disease progression. All samples underwent immunomagnetic enrichment using the AdnaTest BreastCancerSelect (AdnaGen AG, Germany) within 4h after blood withdrawal followed by RNA isolation and subsequent gene expression analysis by reverse transcription and Multiplex-PCR in separated tumor cells using the AdnaTest BreastCancerDetect. CTCs were analyzed for the three breast cancer-associated markers: EpCAM, Muc-1, Her-2 and actin as an internal PCR control. Expression of the ER and PR was assessed in an additional RT-PCR. The analysis of PCR products was performed by capillary electrophoresis on the Agilent Bioanalyzer 2100. RESULTS: The overall detection rate for CTCs was 45% (87/193 patients) with the expression rates of 71% for EpCAM (62/87 patients), 73% for MUC1 (64/87 patients), 48% for HER2 (42/87 patients), 19% for ER (17/87 patients) and 10% for PR (9/87 patients), respectively. Comparisons with the primary tumor were only performed in CTC+ patients (n=87). In 48/62 (77%) patients with ER+ tumors, CTCs were ER- and 46/53 (87%) patients with PR+ tumors did not express PR on CTCs. Primary tumors and CTCs displayed a concordant ER and PR status in only 41% (p=0.260) and 45% (p=0.274) of cases, respectively. CONCLUSION: Most of the CTCs were ER/PR-negative despite the presence of an ER/PR- positive primary tumor. The predictive value of hormone receptor status expression profile of CTCs for palliative endocrine therapy has to be prospectively evaluated. STATEMENT: We recently demonstrated in more than 400 primary breast cancer patients that the expression profile between CTCs and the primary tumor with regard to ER/PR/HER2 positivity differs. The concordance rate between ER, PR and HER2 status of CTCs and the primary tumor was 29%, 25% and 53%, respectively (Fehm T et al., Breast Cancer Res Aug 10 2009, 11(4) pR59). Based on these results we studied blood samples of 193 metastatic breast cancer patients participating in the German DETECT study (1) to reevaluate the ER/PR expression by CTCs and (2) to compare the hormone receptor status expression profile of CTCs with the primary. As already shown for primary breast cancer, most of the CTCs were ER/PR-negative despite the presence of an ER/PR- positive primary tumor. In the metastatic setting the phenotype of CTC reflects the phenotype of metastatic disease. Therefore palliative treatment selected based on the expression profile may not be effective since the phenotype has changed during disease progression. To our knowledge, this study is one of the biggest to compare hormonal receptor expression on CTC and the primary tumor. We hope that our manuscript is suitable for publication in Gynecologic Oncology.

Management of patients with brain metastases receiving trastuzumab treatment for metastatic breast cancer.

BACKGROUND: With the more effective control of visceral metastases in patients with metastatic breast cancer (MBC), an increasing number of patients face brain metastases (BM). The aim of this retrospective analysis was to investigate the incidence and factors affecting the prognosis of patients with BM under trastuzumab treatment for MBC. PATIENTS AND METHODS: A total of 75 HER2positive patients treated with trastuzumab for MBC were included. RESULTS are discussed in the context of the current literature. RESULTS: Patients who developed BM (n = 29) had longer median progression-free survival (PFS) during first-line chemotherapy and longer overall survival (OS) after diagnosis of MBC than 46 patients without BM (PFS: 27 vs. 14 months, p = 0.039; OS: 46 vs. 18 months, p = 0.067). Median survival of patients with continuation of trastuzumab after diagnosis of BM was longer than survival of patients with discontinuation of trastuzumab treatment after BM (18 vs. 3 months, p = 0.006). Survival of patients who were treated with surgery and radiotherapy for BM was better compared with radiotherapy alone (9 vs. 5 months, p = not significant) or best supportive care (9 vs. 2 months, p = 0.049). CONCLUSIONS: Continuation of trastuzumab treatment as well as resection of BM seem to give further benefit in the treatment of patients with HER2-overexpressing MBC.