Prediction of response to neoadjuvant chemotherapy in breast cancer patients by circulating apoptotic biomarkers nucleosomes, DNAse, cytokeratin-18 fragments and survivin.

Biomarkers predicting response to neoadjuvant chemotherapy in locally confined breast cancer (LBC) are highly needed. We prospectively assessed serial blood levels of apoptotic biomarkers nucleosomes, DNAse activity, cytokeratin-18 fragments (M30) and survivin in 51 LBC patients and correlated them with response to neoadjuvant treatment and established tumor markers. As controls, we used 31 healthy subjects, 13 patients with benign diseases and 28 with metastatic breast cancer (MBC). Levels of nucleosomes and survivin were elevated in LBC and MBC while M30, CEA and CA 15-3 levels were only elevated in MBC. During neoadjuvant chemotherapy, LBC patients with no change of disease (N=13) had significantly higher pretherapeutic levels of nucleosomes than patients with remission (N=38). We conclude that apoptotic biomarkers bear valuable information for diagnosis and therapy response prediction in LBC patients.

Circulating immunogenic cell death biomarkers HMGB1 and RAGE in breast cancer patients during neoadjuvant chemotherapy.

Neoadjuvant chemotherapy in breast cancer patients aims at preoperative reduction of tumor volume for better resection results and prognosis. As not all patients respond to neoadjuvant therapy, predictive biomarkers are needed for more efficient individual management. In prospectively collected sera of 51 consecutive locally confined breast cancer (LBC) patients receiving preoperative, neoadjuvant chemotherapy, value level kinetics of soluble high mobility group box 1 (HMGB1), soluble receptor for advanced glycation end products (sRAGE) as well as the established breast cancer biomarkers CA 15-3 and carcinoembryonic antigen (CEA) were investigated and correlated with therapy response objectified by pathological staging at surgery. In addition, biomarkers were measured in sera of 30 healthy controls (HC), 13 patients with benign breast diseases, and 28 metastatic breast cancer (MBC) patients. Pretherapeutic levels of soluble HMGB1 were decreased in MBC, while sRAGE was already decreased in LBC. In contrast, CA 15-3 and CEA were strongly elevated in MBC, but not in LBC. Combination of sRAGE and CA 15-3 enabled best discrimination of LBC from HC (AUC 78.2 %; sens 58 % at 95 % spec), while CA15-3 and CEA discriminated best between MBC and all controls (AUC 90.9 %; sens 70 % at 95 % spec). In LBC patients undergoing neoadjuvant chemotherapy, nine patients achieved complete remission (CR), 29 achieved partial remission (PR), while 13 had no change of disease (NC). NC patients tended to have higher HMGB1 and lower sRAGE levels before therapy onset (p = 0.056 and p = 0.054), while CA 15-3 and CEA did not predict therapeutic outcome. Furthermore, kinetics of HMGB1 during therapy correlated with efficacy of the treatment (p = 0.053). Markers of immunogenic cell death are valuable for the diagnosis of MBC and early estimation of response to neoadjuvant therapy in LBC patients.

Circulating nucleosomes and biomarkers of immunogenic cell death as predictive and prognostic markers in cancer patients undergoing cytotoxic therapy.

INTRODUCTION: Immunogenic cell death markers are released from apoptotic and necrotic cells upon pathologic or therapeutic causes and stimulate the innate and adaptive immune system. Cell death products such as nucleosomes, damage-associated molecular pattern (DAMP) molecules such as the high-mobility group box 1 protein (HMGB1) and its receptor of advanced glycation end products (sRAGE) are supposed to play an essential role in driving this process. However, this immunogenic activation may have dual effects, either by sensitizing the immune system for more efficient tumor cell removal or by creating a favorable tumor microenvironment that facilitates tumor growth, proliferation and invasiveness. AREAS COVERED: Here, we review recent findings on the relevance of serum nucleosomes, DNAse activity, HMGB1 and sRAGE as biomarkers for the diagnosis, prognosis and therapy prediction in cancer disease. EXPERT OPINION: In comparison with healthy controls, cancer patients demonstrated elevated serum levels of nucleosomes and HMGB1 while sRAGE levels were decreased. During locoregional and systemic cytotoxic therapies, a high release of nucleosomes and HMGB1 as well as low release of sRAGE before and during the initial phase of the treatment was found to be associated with poor response to the therapy and patient survival. Therefore, immunogenic cell death markers are promising tools for the prognosis, therapy prediction and monitoring in cancer patients.

Apoptosis-related biomarkers sFAS, MIF, ICAM-1 and PAI-1 in serum of breast cancer patients undergoing neoadjuvant chemotherapy.

BACKGROUND: Neoadjuvant chemotherapy improves surgical options and prognosis in patients with operable breast cancer. Predictive biomarkers are needed to choose the most effective therapy and to avoid unnecessary toxicity. PATIENTS AND METHODS: We analyzed the courses of apoptosis-related serum biomarkers macrophage migration-inhibitory factor (MIF), soluble cell death receptor sFAS, soluble intercellular adhesion molecule (sICAM), plasminogen activator inhibitor 1 (PAI-1) as well as the oncological biomarkers carcino-embryonic antigen (CEA) and carbohydrate antigen 15-3 (CA15-3) in prospectively collected sera of 51 patients with locally confined breast cancer undergoing preoperative chemotherapy. As controls 31 healthy women, 13 patients with benign breast disease and 28 patients with metastasized breast cancer were included. RESULTS: sFAS, MIF, CEA and CA15-3 showed significantly higher serum concentrations in patients with metastasized breast cancer than in healthy and benign controls. Additionally, sFAS and MIF discriminated between locally confined breast cancer and healthy controls with an area under the curve (AUC) in receiver operating characteristic (ROC) curves of 73.4% and 70.7%. After neoadjuvant chemotherapy, 38 patients achieved complete (N=9) or partial (N=29) remission, while 13 patients had no change of disease. Pretherapeutic levels of MIF were considerably higher in non-responsive patients (p=0.082). In addition, post-therapeutic sICAM and CA15-3 levels were higher in patients without complete remission. CONCLUSION: Apoptosis-related biomarkers are valuable markers in breast cancer patients and show potential for early estimation of the efficiency of neoadjuvant chemotherapy.

Methods for isolation of cell-free plasma DNA strongly affect DNA yield.

Extracellular nucleic acids are present in plasma, serum, and other body fluids and their analysis has gained increasing attention during recent years. Because of the small quantity and highly fragmented nature of cell-free DNA in plasma and serum, a fast, efficient, and reliable isolation method is still a problem and so far there is no agreement on a standardized method. We used spin columns from commercial suppliers (QIAamp DNA Blood Midi Kit from Qiagen; NucleoSpin Kit from Macherey-Nagel; MagNA Pure isolation system from Roche Diagnostics) to isolate DNA from 44 plasma samples in parallel at laboratories in Berlin and Munich. DNA in all samples was quantified by real-time PCR on a LightCycler 480 using three different targets (GAPDH, ß-globin, ERV). The quantities of cell-free DNA for the different isolation methods and genes varied between medians of 1.6 ng/mL and 28.1 ng/mL. This considerable variation of absolute DNA values was mainly caused by the use of different isolation methods (p<0.0001). Comparable results were achieved by the use of the genes GAPDH and ERV while higher values were obtained by use of ß-globin. The laboratory site had only minor influence on DNA yield when manual extraction methods were used.