Clinical relevance of H-RAS, K-RAS, and N-RAS mRNA expression in primary breast cancer patients.

PURPOSE: The RAS family comprises three proto-oncogenes (H-RAS, K-RAS, and N-RAS) and is among the most widely studied of oncogenes. The present study aimed at investigating the clinical relevance of mRNA levels of the three isoforms in a large group of breast cancer patients with a long-term follow-up. METHODS: 198 previously untreated patients were enrolled in the study. mRNA levels of K-RAS, H-RAS, and N-RAS were measured using microarray (Affymetrix HG-U133A). RESULTS: Elevated H-RAS levels were found significantly more frequently in patients with larger (p = 0.021) and ER-positive tumors (p = 0.048), while elevated K-RAS levels were associated with nodal positivity (p = 0.001) and HER2-positivity (p = 0.010). Patients with high N-RAS mRNA levels were more likely to be diagnosed with triple-negativity (p < 0.001) and higher grading (p = 0.001). Patients with high K-RAS levels were more likely to show an elevated H-RAS (p = 0.003). After a median follow-up of 183 months, patients with high N-RAS expression had significantly reduced overall survival (OS) compared with patients with low N-RAS (mean: 146.9 vs. 211.0 months; median 169.3 vs. not reached; p = 0.009). In patients with non-metastatic disease at the time of tissue sampling, mean disease-free survival (DFS) was 150.1 months for patients with high N-RAS versus 227.7 months with low N-RAS; median DFS was not reached (p = 0.004). The expression of H-RAS and K-RAS was not associated with DFS/OS. In the multivariable analysis, distant metastasis, HER2 positivity, and elevated N-RAS mRNA levels independently predicted reduced OS, while nodal status, HER2 status, and N-RAS predicted reduced DFS. CONCLUSIONS: Elevated N-RAS mRNA levels predict impaired clinical outcome; hypothetically, further exploration of the RAS signaling pathway might enable identifying potential targeted treatment strategies. The association between high N-RAS levels and the most aggressive among breast cancer subtypes, the triple-negative phenotype, for which targeted approaches are still lacking, underlines the need to further investigate the RAS family.

Disseminated Tumor Cells and Dormancy in Breast Cancer Progression.

Hematogenous dissemination of single cancer cells is a common phenomenon in patients with solid tumors. These cells may experience different fates: most will die during the process; some will grow into metastasis and some will persist in secondary homing sites for many years in a state referred to as dormancy. The mechanisms of this state are still not clear; single cancer cells can survive either by completely withdrawing from the cell cycle or by continuing to proliferate at a slow rate that is counterbalanced by cell death. Another hypothesis assumes that at least some of dormant tumor cells feature stem cell-like characteristics that may contribute to their extremely long half-lives and enhance chemotherapy resistance. Breast cancer is particularly known for prolonged periods of clinical freedom of disease (sometimes up to 20-30 years), followed by a distant relapse. In this chapter, we explore the relationship between the clinical phenomenon of tumor dormancy and the disseminated tumor cells and discuss the potential implications for treatment.

Clinical relevance of circulating tumor cells in ovarian, fallopian tube and peritoneal cancer.

PURPOSE: Presence of circulating tumor cells (CTCs) is associated with impaired clinical outcome in several solid cancers. Limited data are available on the significance of CTCs in gynaecological malignancies. The aims of the present study were to evaluate the dynamics of CTCs in patients with ovarian, fallopian tube and peritoneal cancer during chemotherapy and to assess their clinical relevance. METHODS: 43 patients with ovarian, fallopian tube and peritoneal cancer were included into this prospective study. Patients received chemotherapy according to national guidelines. CTC analysis was performed using the CellSearch system prior to chemotherapy, after three and six cycles. RESULTS: In 26% of the patients, ≥ 1CTC per 7.5 ml of blood was detected at baseline (17% of patients with de novo disease, compared to 35% in recurrent patients). Presence of CTCs did not correlate with other factors. After three cycles of therapy, CTC positivity rate declined to 4.8%. After six cycles, no patient showed persistent CTCs. Patients with ≥ 1 CTC at baseline had significantly shorter overall survival and progression-free survival compared to CTC-negative patients (OS: median 3.1 months vs. not reached, p = 0.006, PFS: median 3.1 vs. 23.1 months, p = 0.005). When only the subgroup with newly diagnosed cancer was considered, the association between CTC status and survival was not significant (OS: mean 17.4 vs. 29.0 months, p = 0.192, PFS: 14.3 vs. 26.9 months, p = 0.085). Presence of ≥ 1 CTC after three cycles predicted shorter OS in the entire patient cohort (p < 0.001). CONCLUSIONS: Hematogenous tumor cell dissemination is a common phenomenon in ovarian, fallopian tube and peritoneal cancer. CTC status before start of systemic therapy correlates with clinical outcome. Chemotherapy leads to a rapid decline in CTC counts; further research is needed to evaluate the clinical value of CTC monitoring after therapy.

Axillary ultrasound for prediction of response to neoadjuvant therapy in the context of surgical strategies to axillary dissection in primary breast cancer: a systematic review of the current literature.

PURPOSE: Data on the optimal treatment strategy for patients undergoing neoadjuvant therapy (NAT) who initially presented with metastatic nodes and convert to node-negative disease (cN+ → ycN0) are limited. Since NAT leads to axillary downstaging in 20-60% of patients, the question arises whether these patients might be offered less-invasive procedures than axillary dissection, such as sentinel node biopsy or targeted removal of lymph nodes marked before therapy. METHODS: We performed a systematic review of clinical studies on the use of axillary ultrasound for prediction of response to NAT and ultrasound-guided marking of metastatic nodes for targeted axillary dissection. RESULTS: The sensitivity of ultrasound for prediction of residual node metastasis was higher than that of clinical examination and MRI/PET in most studies; specificity ranged in large trials from 37 to 92%. The diagnostic performance of ultrasound after NAT seems to be associated with tumor subtype: the positive predictive value was highest in luminal, the negative in triple-negative tumors. Several trials evaluated the usefulness of ultrasound for targeted axillary dissection. Before NAT, nodes were most commonly marked using ultrasound-guided clip placement, followed by ultrasound-guided placement of a radioactive seed. After chemotherapy, the clip was detected on ultrasound in 72-83% of patients; a comparison of sonographic visibility of different clips is lacking. Detection rate after radioactive seed placement was ca. 97%. CONCLUSION: In conclusion, ultrasound improves prediction of axillary response to treatment in comparison to physical examination and serves as a reliable guiding tool for marking of target lymph nodes before the start of treatment. High quality and standardization of the examination is crucial for selection of patients for less-invasive surgery.

Determination of the androgen receptor status of circulating tumour cells in metastatic breast cancer patients.

BACKGROUND: The prognostic relevance of circulating tumour cells (CTCs) in metastatic breast cancer (MBC) patients has been confirmed by several clinical trials. However, predictive blood-based biomarkers for stratification of patients for targeted therapy are still lacking. The DETECT studies explore the utility of CTC phenotype for treatment decisions in patients with HER2 negative MBC. Associated with this concept is a plethora of translational projects aiming to identify potential predictive biomarkers. The androgen receptor (AR) is expressed in over 70% of hormone receptor-positive and up-to 45% of triple-negative tumours. Studies has indicated the promising nature of AR as a new therapy target with a clinical benefit rate for anti-AR treatment in MBC patients up to 25% The aim of this analysis was the characterization of CTCs regarding the expression of the AR using immunofluorescence. METHODS: MBC patients were screened for the HER2-status of CTCs in the DETECT studies. In a subset of CTC-positive patients (n = 67) an additional blood sample was used for immunomagnetic enrichment of CTCs using the CellSearch® Profile Kit prior to transfer of the cells onto cytospin slides. Establishment of immunofluorescence staining for the AR was performed using prostate cancer cell lines LNCaP and DU145 as positive and negative control, respectively. Staining of DAPI, pan-cytokeratin (CK) and CD45 was applied to identify nucleated epithelial cells as CTCs and to exclude leucocytes. RESULTS: Co-staining of the AR, CK and CD45 according to the above mentioned workflow has been successfully established using cell lines with known AR expression spiked into the blood samples from healthy donors. For this translational project, samples were analysed from 67 patients participating in the DETECT studies. At least one CTC was detected in 37 out of 67 patients (56%). In 16 of these 37 patients (43%) AR-positive CTCs were detected. In eight out of 25 patients (32%) with more than one CTC, AR-positive and AR-negative CTCs were observed. CONCLUSION: In 43% of the analysed CTC samples from patients with MBC the AR expression has been detected. The predictive value of AR expression in CTCs remains to be evaluated in further trials.

Cyclin-dependent kinase 4/6 inhibitors: what have we learnt across studies, therapy situations and substances.

PURPOSE OF REVIEW: Cyclin-dependent kinases (CDK) are key regulatory enzymes that control cell cycle and cell division. In the recent years, new therapeutic options selectively targeting CDK 4 and 6 have shown promising clinical activity in several solid tumors. Since 2015, three CDK 4/6 inhibitors have been approved for treatment of hormone receptor-positive HER2-negative metastatic breast cancer: palbociclib, ribociclib and abemaciclib. These drugs share a common mechanism of action and have been evaluated in studies with a similar design. The following review gives a clinical overview of the CDK 4/6 inhibitors in breast cancer therapy and highlight current study data with regard to their antitumor efficacy and toxicities. RECENT FINDINGS: In clinical trials in the first-line and later-line setting, palbociclib, ribociclib and abemaciclib in combination with endocrine therapy significantly prolonged progression-free survival. The most common adverse events during treatment with CDK 4/6 inhibitors are neutropenia, fatigue and gastrointestinal symptoms. SUMMARY: CDK 4/6 inhibitors represent a valuable treatment option for patients with metastatic hormone receptor-positive HER2-negative breast cancer. Although the clinical efficacy of the three agents seems similar, their toxicity profiles differ. Therefore, the choice of a CKD 4/6 inhibitor depends on patient's characteristics and individual preferences. VIDEO ABSTRACT: In the video, the author describes the content of the review and present the main topics discussed in the article (http://links.lww.com/COOG/A44).

The clinical relevance of serum vascular endothelial growth factor (VEGF) in correlation to circulating tumor cells and other serum biomarkers in patients with metastatic breast cancer.

PURPOSE: VEGF is one of the most important angiogenesis-stimulating cytokines and has been previously shown to be overexpressed in several solid cancers. The aim of the present study was to assess the clinical relevance of serum VEGF (sVEGF) in a large cohort of metastatic breast cancer patients and to explore the relationship between sVEGF and other blood-based biomarkers. METHODS: Two hundred fifty-three patients with metastatic breast cancer were enrolled in this prospective, multicentre study. Blood samples were collected before start of first-line or later-line treatment. sVEGF was quantified by a commercially available ELISA. Circulating tumor cells (CTCs) were detected using CellSearch and other biomarkers (EGFR, HER2, RAS p21, TIMP1, CAIX) by ELISA. RESULTS: Levels of sVEGF were determined in all patients, with a median concentration of 231 pg/ml. After a median follow-up of 19 months, median overall survival (OS) was 10.2 months in patients with sVEGF levels above the upper quartile (i.e. 367 pg/ml), while median OS has not been reached in patients with sVEGF < 367 pg/ml (p < 0.001). Median progression-free survival (PFS) was 4.8 months for patients with sVEGF ≥ 367 pg/ml versus 9.1 months with sVEGF levels < 367 pg/ml (p < 0.001). Patients with sVEGF levels ≥ 367 pg/ml and ≥ 5 CTCs had the shortest OS, while those with sVEGF < 367 pg/ml and non-elevated CTCs had the longest OS. CTCs, grading, line of therapy and RAS p21 were independent predictors of OS. sVEGF, line of therapy and CTCs were independent predictors of PFS in the multivariate analysis. CONCLUSIONS: Metastatic breast cancer patients with elevated levels of sVEGF have significantly worse clinical outcome. This finding supports the biological role of VEGF in breast cancer. TRIAL REGISTRATION: Current Controlled Trials ISRCTN59722891 (DETECT).

Elevated serum RAS p21 is an independent prognostic factor in metastatic breast cancer.

BACKGROUND: An important component of the RAS signalling pathway, the RAS p21 oncogene, is frequently hyperactivated in breast cancer. Its expression in tumor tissue has been linked to poor clinical outcome. This study was designed to evaluate the clinical relevance of RAS p21 levels in peripheral blood in a large cohort of metastatic breast cancer patients. METHODS: Two hundred fifty-one patients with metastatic breast cancer were enrolled in this prospective, multicentre, open-label, non-randomized study. Blood samples were collected before start of first-line or later-line treatment. RAS p21 was determined using a sandwich-type ELISA immunoassay. For the determination of the cutoff, blood samples from age-matched healthy controls were analyzed. A value above 452 pg/ml was regarded as elevated (mean + 2 x SD). In the univariate survival analysis, two other cutoffs were considered as well (50th and 75th percentile of patients, i.e. 229 pg/ml and 320 pg/ml). Circulating tumor cells (CTCs) were detected using the CellSearch system. RESULTS: 29 of 251 (12%) patients had RAS p21 levels above the cut-off level of 452 pg/ml. Clinical-pathological parameters, such as hormone receptor and HER2 status, line of therapy and CTC status, did not correlate with RAS p21 levels. Elevated RAS p21 was significantly associated with shorter progression-free and overall survival in the univariate analysis (median PFS: 3.9 months [95%-CI: 1.8-6.0] for patients with elevated RAS p21 levels versus 8.5 months [95%-CI: 7.4-9.5] with non-elevated levels [p = 0.01]; median OS: 7.1 months [95%-CI: 0.3-14.2] versus not reached [p = 0.002], respectively). When RAS p21 cutoffs other than 452 pg/ml were considered, elevated RAS p21 was significantly associated with OS but not with PFS. Classical clinical-pathological factors were included into a multivariate Cox regression analysis. In addition, factors previously shown to influence survival in a univariate analysis, such as serum HER2, CAIX and TIMP1, were included as well. In the multivariate analysis, RAS p21, presence of ≥5 CTCs per 7.5 ml blood, higher grading and higher line of therapy remained independent predictors of shorter OS. CONCLUSIONS: Metastatic breast cancer patients with elevated levels of circulating RAS p21 have significantly worse clinical outcome. Hypothetically, these patients might benefit from therapeutic strategies targeting RAS pathway. TRIAL REGISTRATION: Current Controlled Trials ISRCTN59722891 (DETECT); trial registration date: April, 17th 2010; the trial was registered retrospectively.

Pre-Analytical and Analytical Variables of Label-Independent Enrichment and Automated Detection of Circulating Tumor Cells in Cancer Patients.

Circulating tumor cells (CTCs) are promising tools for risk prediction and the monitoring of response to therapy in cancer patients. Within the EU/IMI CANCER-ID consortium, we validated CTC enrichment systems for future inclusion into clinical trials. Due to the known heterogeneity of markers expressed on CTCs, we tested the Parsortix® system (ANGLE plc) which enables label-independent CTC enrichment from whole blood based on increased size and deformability of these tumor cells compared to leukocytes. We performed extensive comparisons both with spiked-in blood models (i.e., MDA-MB-468 tumor cell line cells spiked at very low concentration into blood from healthy donors) and validated the protocol on actual clinical samples from breast, lung, and gastrointestinal cancer patients to define optimal conditions for CTC enrichment. Multiple parameters including cassette gap, separation pressure, and cell fixatives were compared in parallel. Also, the compatibility of blood collection tubes with whole genome amplification of isolated tumor cells was demonstrated and we furthermore established a workflow for semi-automated CTC detection using a quantitative cell imager. The established workflow will contribute to supporting the use of size-based CTC enrichment platforms in clinical trials testing the clinical validity and utility of CTCs for personalized medicine.

Liquid biopsy in metastatic breast cancer.

The spread of single tumor cells shed by the primary tumor has been observed in most solid carcinomas and is generally associated with poor clinical outcome. Tumor cells detected in the peripheral blood are commonly referred to as circulating tumor cells (CTCs) and are seen as possible precursors of metastatic disease. Beyond CTCs, circulating tumor DNA and non-coding RNA are increasingly the focus of translation cancer research. In metastatic breast cancer (MBC), elevated levels of CTCs have been confirmed as an independent prognostic factor. While detection of elevated counts after the start of systemic therapy predicts poor response, it is unclear which treatment strategy should be offered in the case of CTC persistence. Currently, the main potentials of blood-based diagnostics in BC are therapy monitoring and liquid biopsy-based treatment interventions. Recently, the first positive study on CTC-guided therapy choices in hormone receptor positive HER2 negative MBC was published. In the present review, we discuss the current data and potential clinical application of liquid biopsy in the metastatic setting.

Circulating and Disseminated Tumor Cells in Breast Carcinoma: Report from the Consensus Conference on Tumor Cell Dissemination during the 39th Annual Meeting of the German Society of Senology, Berlin, 27 June 2019.

Hematogenous dissemination of single tumor cells from the primary tumor is a common phenomenon in most solid malignancies. In breast cancer, presence of circulating tumor cells (CTCs) in the peripheral blood and disseminated tumor cells (DTCs) in bone marrow predicts poor clinical outcome, both in early and metastatic setting. Beyond that, persistence of CTCs/DTCs is associated with shorter relapse-free interval as well. Numerous studies have shown that these cells differ from tumor cells in the primary tumor with regard to hormone and HER2 receptor status and it has been hypothesized that some of them might be in fact cancer stem cells. Recently, the first positive study on CTC-based therapy interventions has been presented at the San Antonio Breast Cancer Symposium 2018, demonstrating that detection of CTCs may guide treatment decisions in metastatic HR-positive HER2-negative disease. In this review, we present the current state of evidence of tumor cell dissemination and discuss the implications for future trials.

Circulating and Disseminated Tumour Cells in Breast Carcinoma: Report from the Consensus Conference on Tumour Cell Dissemination during the 38th Annual Meeting of the German Society of Senology, Berlin, 14 June 2018.

Haematogenic dissemination of tumour cells in breast carcinoma is among the most intensively researched areas in translational oncology. Large meta-analyses have shown the prognostic relevance of the disseminated tumour cells in the bone marrow and circulating tumour cells in the peripheral blood in the adjuvant as well as metastatic setting. The current status of the research was discussed in detail during the annual meeting of the German Society of Senology in Berlin. The following conference report gives an overview of the clinical study landscape and the new methodological developments for improving the detection and phenotyping of the circulating and disseminated tumour cells.

Clinical Relevance of Collagen Protein Degradation Markers C3M and C4M in the Serum of Breast Cancer Patients Treated with Neoadjuvant Therapy in the GeparQuinto Trial.

Background: Remodeling of extracellular matrix through collagen degradation is a crucial step in the metastatic cascade. The aim of this study was to evaluate the potential clinical relevance of the serum collagen degradation markers (CDM) C3M and C4M during neoadjuvant chemotherapy for breast cancer. Methods: Patients from the GeparQuinto phase 3 trial with untreated HER2-positive operable or locally advanced breast cancer were enrolled between 7 November 2007, and 9 July 2010, and randomly assigned to receive neoadjuvant treatment with EC/docetaxel with either trastuzumab or lapatinib. Blood samples were collected at baseline, after four cycles of chemotherapy and at surgery. Cutoff values were determined using validated cutoff finder software (C3M: Low ≤9.00 ng/mL, high >9.00 ng/mL, C4M: Low ≤40.91 ng/mL, high >40.91 ng/mL). Results: 157 patients were included in this analysis. At baseline, 11.7% and 14.8% of patients had high C3M and C4M serum levels, respectively. No correlation was observed between CDM and classical clinical-pathological factors. Patients with high levels of CDM were significantly more likely to achieve a pathological complete response (pCR, defined as ypT0 ypN0) than patients with low levels (C3M: 66.7% vs. 25.7%, p = 0.002; C4M: 52.7% vs. 26.6%, p = 0.031). Median levels of both markers were lower at the time of surgery than at baseline. In the multivariate analysis including clinical-pathological factors and C3M levels at baseline and changes in C3M levels between baseline and after four cycles of therapy, only C3M levels at baseline (p = 0.035, OR 4.469, 95%-CI 1.115-17.919) independently predicted pCR. In a similar model including clinical-pathological factors and C4M, only C4M levels at baseline (p = 0.028, OR 6.203, 95%-CI 1.220-31.546) and tumor size (p = 0.035, OR 4.900, 95%-CI 1.122-21.393) were independent predictors of pCR. High C3M levels at baseline did not correlate with survival in the entire cohort but were associated with worse disease-free survival (DFS; p = 0.029, 5-year DFS 40.0% vs. 74.9%) and overall survival (OS; p = 0.020, 5-year OS 60.0% vs. 88.3%) in the subgroup of patients randomized to lapatinib. In the trastuzumab arm, C3M did not correlate with survival. In the entire patient cohort, high levels of C4M at baseline were significantly associated with shorter DFS (p = 0.001, 5-year DFS 53.1% vs. 81.6%) but not with OS. When treatment arms were considered separately, the association with DFS was still significant (p = 0.014, 5-year DFS 44.4% vs. 77.0% in the lapatinib arm; p = 0.023, 5-year DFS 62.5% vs. 86.2% in the trastuzumab arm). Conclusions: Collagen degradation markers are associated with response to neoadjuvant therapy and seem to play a role in breast cancer.

The prognostic relevance of urokinase-type plasminogen activator (uPA) in the blood of patients with metastatic breast cancer.

In breast cancer (BC), elevated levels of urokinase-type plasminogen activator (uPA) in tumor tissue have been confirmed as a strong prognostic factor in level-of-evidence-1 studies. The aim of the present study was to evaluate the clinical relevance of uPA levels in serum of metastatic BC patients and to compare uPA with other blood-based biomarkers. 252 patients were enrolled in this prospective, multicentre study. Blood samples were collected before begin of first-line or later-line systemic treatment. Serum uPA was quantified by a commercially available ELISA. Circulating tumor cells (CTCs) were detected using CellSearch; other biomarkers (EGFR, VEGF, HER2, RAS p21, TIMP1, CAIX) by ELISA. Using the ROC analysis, the optimal cut-off value (determined by the Youden index) of serum uPA was 2.52 ng/ml. Using this value, 26% of patients had elevated uPA levels. Patients with visceral metastasis and more than one metastatic site were significantly more likely to present with elevated uPA levels. CTC status, serum HER2, RAS p21, CAIX, TIMP1 and VEGF correlated significantly with uPA levels. Elevated uPA levels predicted shorter overall and progression-free survival in univariate analysis (median OS: 7.5 months [95%-CI 4.5-10.5 months] vs. not reached, p < 0.001; PFS: 4.8 [95%-CI: 3.1-6.5] vs. 9.1 [7.4-10.8] months, p < 0.001). In multivariate analysis, elevated uPA, presence of ≥5 CTCs, elevated RAS p21, higher grading and higher line of therapy were independent predictors of shorter OS, while elevated CTC counts, higher line of therapy and negative estrogen receptor status were independent predictors of shorter PFS. In conclusion, elevated uPA levels independently predict reduced overall survival and improved prognostication in patients with known CTC status. Whether high serum uPA might identify patients most likely to benefit from therapies targeting uPA, remains to be evaluated in future trials.

Metronomic Chemotherapy for Primary Non-Metastatic Breast Cancer - a Systematic Review of the Literature.

Conventional chemotherapy is based on the "maximum tolerated dose" principle and aims at administering high doses of cytotoxic drugs followed by a rest period necessary for the body to recover. In the last decades alternative strategies have been developed to avoid serious side effects of conventional treatment, among them the metronomic chemotherapy. Much like a metronome keeps steady rhythm, metronomic therapy is administered continuously in low doses for a long time. In metastatic breast cancer, metronomic therapy is a valid option in pretreated or vulnerable patients and its use has recently been incorporated into various guidelines. In early breast cancer, the role of metronomic treatment remains to be clarified. A systematic review of PubMed/MEDLINE, ClinicalTrials.gov, the European Clinical Trials Database (EudraCT) and the Cochrane Database was conducted. In the present review, we discuss the current evidence on metronomic chemotherapy in non-metastatic breast cancer.

Clinical Relevance of Serum HER2 and Circulating Tumor Cell Detection in Metastatic Breast Cancer Patients.

BACKGROUND/AIM: Presence of circulating tumor cells (CTCs) is associated with impaired survival in metastatic breast cancer (MBC). This study was designed to evaluate whether assessment of serum HER2 (sHER2) levels provide additional prognostic information in MBC. MATERIALS AND METHODS: Two hundred and fifty-three MBC patients were enrolled in this multicentre trial. CTCs were detected before the start of first- or later-line treatment using the CellSearch system. sHER2 was determined using ELISA. RESULTS: ≥5 CTCs were detected in 122 of 245 evaluable patients (49.8%). One hundred and nineteen of 251 patients (47%) had sHER2 levels above 15 ng/ml. Median overall survival (OS) was 16.3 months in patients with elevated sHER2; median OS in patients with non-elevated sHER2 has not been reached (p=0.001). Patients with ≥5 CTCs were more likely to present with elevated sHER2 (61% vs. 33% in those with <5 CTC; p<0.001). In patients with HER2-negative tumors, elevated sHER2 was associated with shorter OS and PFS; in HER2-positive patients with OS only. Including sHER2, CTC status and established prognostic factors into a multivariate analysis, only the presence of CTCs and higher-line of therapy remained independent predictors of OS. CONCLUSION: Elevated levels of sHER2 are associated with worse survival, irrespective of the HER2 status of the tumor. However, sHER2 does not provide additional prognostic information in patients with known CTC status.

Evaluation of serum epidermal growth factor receptor (EGFR) in correlation to circulating tumor cells in patients with metastatic breast cancer.

Overexpression of epidermal growth factor receptor in breast cancer is associated with estrogen receptor negativity, higher histological grade and larger tumors. The aim of the present study was to evaluate the clinical significance of serum EGFR (sEGFR) in relation to circulating tumor cells (CTCs) in metastatic breast cancer. 252 patients were enrolled in this prospective multicentre study. Blood was drawn before start of a new line of therapy. sEGFR was determined using a sandwich-type ELISA. CTCs were detected using CellSearch. sEGFR was determined in 48 healthy controls and 252 patients, with no significant differences between the two groups. Clinical-pathological parameters did not correlate with sEGFR, irrespective of the cutoff chosen. Patients with sEGFR levels above the 50th and 75th percentile were more likely to present with <5 CTCs per 7.5 ml blood (p = 0.007; p = 0.003). Patients with sEGFR ≥73 ng/ml had significantly longer overall survival than those with sEGFR <73 ng/ml (19.7 vs. 15.2 months; p = 0.007). In the multivariate analysis, presence of ≥5 CTCs, higher grading and higher line of therapy remained independent predictors of shorter OS, while only higher line of therapy and presence of ≥5 CTCs were independent predictors of shorter PFS.

Potential Role of Circulating Tumor Cell Detection and Monitoring in Breast Cancer: A Review of Current Evidence.

The phenomenon of hematogenous tumor cell dissemination in patients with solid tumors has been extensively explored over the last decades. Breast cancer research investigated at first disseminated tumor cells in the bone marrow; however, the focus soon moved to circulating tumor cells (CTCs) in the peripheral blood as blood is easily accessible without an invasive procedure. The prognostic significance of CTC presence has been shown in large studies both in adjuvant and metastatic setting and commercially available detection assays have been evaluated for monitoring in clinical trials. Beyond detection and enumeration of CTCs, the characterization of single tumor cells may enhance our knowledge on disease progression and thus optimize treatment choices.

Liquid Biopsy in Metastasized Breast Cancer as Basis for Treatment Decisions.

According to current guidelines, the additional biopsy of breast cancer metastases to analyze the receptor status for phenotype assessment is recommended. However, due to clinical difficulties in performing biopsies of metastatic lesions, the phenotype of the primary tumor most often determines the treatment decisions in metastatic breast cancer. Liquid biopsy allows the analysis of several circulating biomarkers like circulating tumor cells (CTCs) or circulating tumor DNA (ctDNA) in peripheral blood samples of cancer patients. Thus, it is an elegant and easily practicable technique that delivers information on the current disease status. Determination of the CTC phenotype regarding the hormone receptor and human epidermal growth factor receptor 2 (HER2) status might replace additional tissue biopsy for planning further therapy strategies. Liquid biopsy is a crucial step towards a more individualized cancer therapy. In contrast to the conventional concept of tissue biopsy, it offers an easy, less invasive acquisition of biomaterial. In addition, it allows multiple repetitions and real-time monitoring of metastasized disease in the clinical routine. However, the clinical utility of liquid biopsy still needs to be evaluated.

Circulating tumor cells in breast cancer--current status and perspectives.

The phenomenon of tumor cell dissemination through the blood stream has been known since the 19th century. Circulating tumor cells (CTCs) may be detected in peripheral blood of patients with breast cancer and may serve as a surrogate marker for minimal residual disease. Prognostic relevance of CTCs has already been demonstrated in early and metastatic breast cancer and commercially available detection systems are currently employed in various clinical trials. Since peripheral blood is an easily accessible compartment, serial reevaluation of CTCs is possible and may contribute to better therapy monitoring. Another potential of CTCs lies in the characterization of tumor cells. Expression profiles may differ between CTCs and primary tumor, which may result in different responses to treatment. Assessment of molecular features of CTCs may be an important step for the optimization of adjuvant and metastatic systemic therapy.