RESUMO
INTRODUCTION: The calibrated automated thrombography (CAT) assay is emerging as a reliable tool for real time estimation of thrombin generation (TG) potential. As a time-dependent colorimetric assessment of thrombin quantity generated per sample, it measures the amount of thrombin-cleaved fluorogenic substrate produced, and so is regarded as a better overall indicator of the clotting efficiency and function of the haemostatic process than one stage clotting-time based assays. AIM: We already recognise that the pathways underlying the thrombotic phenotype for different malignancies may be driven by different factors of the coagulation cascade with TG has a common denominator. Two such malignancies with high venous thromboembolism (VTE) incidence are Multiple myeloma (MM) and Pancreatic cancer (PC). Understanding the underlying variations in these two distinct cancer models using patient samples and cell lines might potentially allow individual approaches to identifying thrombotic risk and relevant prevention strategies. MATERIALS AND METHODS: Citrated blood samples were taken from healthy controls, pre-surgical pancreatic cancer and pre-chemotherapy multiple myeloma patients enrolled into ongoing clinical trials. The clotting ability was tested using platelet free plasma (PPP) on a one-step clotting time-based (CT) assay and the TG profiles were evaluated on a Thrombinoscope™ software (Thrombinoscope BV, Maastricht, Netherlands). Solid tumour cells of pancreatic cancer and malignant haematological cell lines were used at various cell concentrations for the CAT assay, which was performed with the addition of platelet-free control plasma or control plasma deficient in coagulation factors VII and XII. RESULTS: At TF concentration conditions of 1 pmol/L, the peak height of thrombin generated on thrombogram curves strongly correlated with CT of patient samples. Compared to healthy controls, pancreatic cancer had higher thrombin peaks (P), shorter lag times (LG), and an overall stronger TG profile than MM. Pancreatic cancer cell lines exhibited higher concentration-dependent TG profiles in control plasma than haematological cell lines, with higher peaks, endogenous thrombin potential (ETP), shorter lag times (LG) and faster times-to-peak (ttPeak). CONCLUSIONS: This study demonstrates that the CAT assay is a useful predictor of the thrombotic phenotype in cancer patients as it gives a more comprehensive overall coagulation profile than one stage CT-based assays. It shows that for patient samples and cell lines, the similarities and differences that exists in the TG potential, significantly depends on specific coagulation factors present in the intrinsic or extrinsic arms of the clotting cascade.
RESUMO
INTRODUCTION: IMiD-based regimens are now widely considered standard of care in the treatment of Multiple myeloma (MM) patients (Kumar et al., 2008). One of the major adverse events noted in many MM clinical studies in patients treated with combination regimens including Thalidomide (Thal), Lenanlidomide (Len) and dexamethasone or chemotherapy was the development of hrombosis (Carrier et al., 2011). AIM: We have postulated that one mechanism for venous thromboembolism (VTE) occurrence may be through chemotherapy damage to endothelium (Date et al., 2013) and much recent work has centred on the study of soluble dysfunction markers to predict this event. In states of endothelial dysfunction soluble antigen concentrations of circulating endothelial activation markers sCD106 and sCD54 have been shown to increase (Burger and Touyz, 2012), and sCD106 was recently associated with inferior survival in newly diagnosed MM patients treated with Thal- or Len-based therapies (Terpos et al., 2013). MATERIALS AND METHODS: Serum from newly diagnosed and relapsed MM patients were collected before, during and after prescribed chemotherapy courses (minimum of 4 cycles). Levels of endothelial activation markers sCD106 and sCD54 were evaluated by quantitative ELISA (Platinum ELISA kits; eBioscience, Hatfield, UK). RESULTS: The percentage mean±SD change in the serum concentration of sCD106 increased by 25.8% after the first cycle of chemotherapy (T2; n=10) relative to T1 prior to chemotherapy administration. These levels subsequently decreased after the second cycle of chemotherapy (T3; n=9) and after completion (T4; n=5), but were still higher than baseline levels (15.5 and 15.3% increase in comparison to baseline, respectively). In contrast, the mean±SD percentage change in sCD54 relative to T1 were similar after the first cycle (T2; n=10) and the second cycle (T3; n=9), but increased by 15.0% after completion of chemotherapy (T4; n=5). Additionally, a statistical correlation was found to exist between the serum concentration of sCD106 and sCD54 (Pearson's correlation coefficient r=0.84, p <0.0005) for all analysed samples (n=39). CONCLUSIONS: In this study, the increase in serum levels of both sCD106 and sCD54 after IMiD-based chemotherapy implies a disruptive effect of these combination regimens on the vascular endothelium. This agrees with previous studies where serum levels of sCD106 were shown to be significantly elevated in chronic lymphocytic leukaemia patients on Len indicating Len-induced endothelial dysfunction, and associated with subsequent DVT development (Aue et al., 2011). Our study confirms the potential significance of these biomarkers in demonstrating chemotherapy-induced endothelial damage. Correlation with VTE however is difficult as most MM patients on chemotherapy receive thromboprophylaxis as per international guidelines.