ALBERT

Description: Abstract 3396 Introduction The risk of venous thromboembolic event (VTE) in patients with cancer is particularly increased compared to normal population. It seems that this risk depends largely on the characteristics of the tumor, such as its site or its stage of evolution, and the anti-neoplasic treatment. The capacity of thrombin generation and D-dimers levels are two biological markers proposed for the stratification of the risk of VTE. We have analyzed thrombin generation and D-dimers levels in patients with breast cancer as markers of a hypercoagulable state, depending on the stage and the period of the tumor evolution. Materials and methods It is a prospective study carried out at the day hospital of the carcinology department of the University Hospital H. Bourguiba in Sfax-Tunisia, including patients with breast cancer at different stages of evolution and different period elapsed since cancer diagnosis. At the time of inclusion, a venous citrated blood sampling (3.2%) was made. The test of thrombin generation was carried out according to the technique of Calibrated Automated Thrombogram assay (CAT®, Diagnostica Stago, Asnières, France). The parameters of the thrombogram were analyzed: endogenous thrombin potential (ETP), peak of thrombin (peak) and mean rate index (MRI). D-dimers were measured using the STA®- Liatest® D-Di assay (Diagnostica-Stago). Results Sixty one patients were included. Their average age is 51.8 ± 10.9 years old. Depending on the stage of cancer disease, 3 sub-groups of patients were distinguished: early local stage (T1, n=16), advanced local stage (T2-T4; n=25) and metastatic stage (M; n=20). Considering the time passed since diagnosis, we have different periods : inferior to 6 months (n=26), 6 to 12 months (n=7), 12 to 36 months (n=15) and more than 36 months (n=13). The analysis of the different parameters of the thrombogram depending on the cancer stage revealed that patients with an advanced local stage and a period elapsed since cancer diagnosis inferior to 6 months had significantly higher values of ETP and thrombin peak (1708±247 nM.min and 379±80 nM) compared to those with an older cancer (1404±308 nM.min, p

Description: Abstract 2292 Introduction: The clinical course of sickle cell disease (SCD) is punctuated by episodic vascular occlusive events. The possibility that activation of the clotting system plays a contributory role in these complications is supported by abundant clinical data during both steady-state disease and pain crisis. Hydroxyurea therapy induces fetal haemoglobin, improves laboratory parameters and reduces acute clinical complications of SCD, but despite an abundance of evidence for coagulation and platelet activation, it remains incompletely defined whether these changes contribute to the reduced thrombin generation. This study is designed to evaluate coagulation profiles of patients with SCA in steady state and to determine whether hypercoagulable state is modified or not in patients on hydroxyurea therapy. Patients and Methods: We studied erythrocyte derived microparticles (Ed-MP) and platelet derived microparticles (Pd-MP) expressing or not expressing phosphatidylserine (PS) in patients with steady state SCD and we evaluated their specific procoagulant activity and their impact on thrombin generation process. A total of 92 steady state SCD patients were included in the study, of which 19 were under treatment with hydroyurea. The control group consisted of 30 healthy age and sex matched controls. Microparticles in whole blood were assessed using flow cytometry. Ed-MP and Pd-MP were identified using an anti-CD235 and CD41 monoclonal antibodies and annexin V. Thrombin generation in platelet poor plasma (PPL) was measured by CAT assay using PPP-reagent 5pM (Thrombinoscope, The Netherlands). Procoagulant phospholipid dependent activity in plasma was assessed by the Procoag-PPL assay (Diagnostica Stago, France). Thrombomodulin (TM) levels were measured by enzyme-linked immunosorbent assay (Elisa) Asserachrom thrombomodulin (Diagnostica Stago, Asnieres, France). Results: Hydroxyurea treated patients had lower counts of leukocytes, reticulocytes and platelets and an increased mean hemoglobin concentration as compared to non treated patients. Leukocyte and reticulocytes counts of treated patients were higher than those of controls. Platelets counts did not differ between treated and untreated patients. Patients on treatment with hydroxyurea had significantly lower levels of Ed-MP/PS+ and Ed-MP compared to untreated patients. The concentration of Pd-MP/PS+ and Pd-MP were not significantly different between hydroxyurea treated and non treated patients. The Ed-MP/PS+ showed a significant inverse correlation with Hb F (p

Description: Background: Despite reduced coagulation factors, it was recently described that patients with cirrhosis have an increased thrombotic tendency. Therefore, assays predicting the risk of bleeding or thrombosis are needed. Aims: As the Tissue Factor (TF) pathway plays a major role in the initiation of the coagulation, we evaluated the equilibrium between Tissue Factor activity (TFa) and Tissue Factor Pathway Inhibitor (TFPI) and their relationship with the thrombin generation (TG) assay in 42 cirrhotic patients. TG was realized in the absence and presence of thrombomodulin (TM), since it was previously evidenced a resistance to TM in cirrhotic patients. Methods: Citrated blood samples of 42patients with confirmed cirrhosis classified according to the Child score (A,B,C), free of familial history of thromboembolism or thrombophilia and not treated by anticoagulant were analyzed. TG was triggered by 5 pM of TF in the presence/absence of thrombomodulin (4 µm), and 4 µM of phospholipids (Cat Assay, Stago,France). TFa was determined by a home-made assay, free TFPI (fTFPI) by the Asserachrom Free TFPI (Diagnostica Stago, France). Protein C (PC) and S (PS) quantified by Staclot Protein C and S assays (Diagnostica Stago, France). Results were compared to a group of 30 healthy subjects (CT). Results: Main results are summarized in the table. For TG assay, only the Lag-Time (LT) and the Endogenous Thrombin Potential (ETP) were indicated.Table 1.nTFa (pM)fTFPI (ng/mL)Thrombin generationPC (%)PC (%)without TMwith TMRatioLT (min)ETP (nM/min)LT (min)ETP (nM/min)ETP+TM/ ETP-TMCT300.24 ± 0.1114.2 ± 3.52.46 ± 0.241659 ± 2882.59 ± 0.21008 ± 2730.60 ± 0.08126 ± 1482 ± 11A120.49** ± 0.7213.8 ± 5.32.82 ± 0.551338** ± 2803.12 ± 0.64862 ± 2220.66 ± 0.1658.9*** ± 10.954.6*** ± 11.6B192.73 ** ± 5.116.5 ± 6.452.82 ± 0.711349* ± 4252.93 ± 0.6998 ± 2190.73 ** ± 0.1146.4***, ° ± 11.148.7*** ± 17.8C114.23 **, °°° ± 5.7620.3 ± 11.52.61 ± 0.611574 ± 3892.99 ± 0.681216 ± 3000.78 **, ° ± 0.0735.6***, °°° ± 1349.3*** ± 18* p〈 0.05; ** p 〈 0.01 versus CT; ° p 〈 0.01 °°°; p 〈 0.001 versus Child A (non-parametric Mann-Whittney test) TFa was significantly (p 〈 °0.01) increased in comparison with CT, and patients with the most severe disease (Child C) have higher levels (p 〈 0.001) than patients with the lowest Child score (A). In contrast, fTFPI levels were not significantly different from CT, whatever the Child score was. A significant decrease of ETP (without TM) was observed for cirrhotic patients with the lowest Child scores (A and B) in comparison with CT, whereas the lag-time (with and without TM) and ETP with TM were not different from CT. Therefore, the ratios of ETP with TM/ETP without TM increased significantly, indicating a resistance to the anticoagulant activity of TM, in relation to the severity of the disease. This resistance was in part explained by the decrease levels of PC since there was a significant negative correlation (r = - 0.35, p = 0.03) between PC levels and the ratio ETP + TM/ETP - TM. PC levels were also inversely correlated to ETP levels, in the absence (r = - 0.54, p = 0.0006) or presence of TM (r = - 0.53, p = 0.0006). PC levels were also inversely correlated to TFa levels (r = - 0.39, p = 0.01), suggesting that PC decrease could be related to a defective synthesis, but also possibly by a consumption due to the activation of the coagulation cascade by the TF. In contrast, no significant correlation were observed for PS. Conclusion: In addition toaresistance to the anticoagulant activity of the PC pathway, cirrhotic patients with the more severe disease stage (Child C) have high levels of TF activity which is not counterbalanced by TFPI. This could contribute to the higher prevalence of thrombotic disease in these patients. The origin of TF remains to be established, but could come from Kuppfer cells in reaction to the liver injury. These results may have clinical implication for the treatment or prophylaxis of thrombosis in cirrhotic patients. Disclosures Van Dreden: Diagnostica Stago: Employment.

Description: Background Clopidogrel is cornerstone treatment for atherothrombotic patients. The incidence of recurrent thrombotic events raises up to 10% of patients on treatment with clopidogrel. Intestinal absorption via the glycoproteine ABCB1, bioactivation by the cytochrome P450 (CYP) system, binding to P2Y12 receptor and inhibition of platelet aggregation are critical steps for the efficiency of clopidogrel treatment. Prospective independent clinical trials showed that the risk of recurrent thrombotic complications is associated with the presence of common polymorphisms of genes encoding ABCB1 (rs1045642C〉T) or the CYP2C19 isoenzyme (*2_rs4244285G〉A, *3_rs4986893G〉A, *4_rs28399504A〉G) or the inefficiency of clopidogrel to decrease intracellular VASP or the presence of high residual platelet reactivity (HRPR). Aim To evaluate the available pharmacogenetic, pharmacological and functional assays for the diagnosis of the resistance to clopidogrel treatment on the same population of patients with atherothrombosis. Materials and Methods A cohort of 94 out-patients with atherothrombosis receiving clopidogrel treatment (75 mg o.d) for more than 3 months were included. Polymorphisms ABCB1, CYP *2, *3, *4, *17 were assessed with rPCR method. VASP was measured with flow cytometry assay and poor response (PR) to the treatment was diagnosed if VASP index was 〉 50. Light Transmission Platelet Aggregation (LTPA) triggered by ADP (5 ìM and 10 ìM) was assessed on citrated platelet rich plasma. Multielectrode aggregometry (WBMEA) was performed on citrated whole blood using Multiplate instrument and ADP reagent (ADP-test®). The cut-off point for the diagnosis of HRPR on clopidogrel treatment was the lower value of the maximum aggregation or the AUC observed in the upper quintile of ADP triggered LTPA or WBMEA measurements. Results The frequency of heterozygous for CYP*2, *3, *4 and ABCB 1 polymorphisms was 28%, 0%, 1%, and 51% respectively. The frequency of the respective homozygocity was 2%, 0%, 0%, and 20%. Patients heterozygous for the CYP*2 G〉A polymorphism showed significantly higher VASP index as compared to the wild type but LTPA and WBMEA were not significantly different. VASP index, LTPA and WBMEA were not significantly different in patients heterozygous or homozygous for the ABCB1 C〉T polymorphism as compared to the wild type. The VASP index was significantly correlated with the LTPA (r=0.44, p

Description: Introduction The optimization of the antithrombotic treatment with LMWH (enoxaparin), direct and indirect anti-Xa ( Apixaban, fondaparinux), in patients with adenocarcinoma of the pancreas, or human breast carcinoma is a challenging issue. The understanding of their mechanism of action in cancer-induced hypercoagulability might provide evidence for treatment optimization. To this aim, we studied the influence of pancreas adenocarcinoma cells (BXPC3), or human breast carcinoma cells (MCF7) on the antithrombotic activity of enoxaparin, fondaparinux and apixaban. Materials and Methods Cells were cultured and adhered in 96-well plates (50cells/µl). Thrombin generation (TG) of normal platelet poor plasma spiked with clinically relevant concentrations of enoxaparin, fondaparinux and apixaban which was added in wells carrying cancer cells was assessed with the Calibrated Automated Thrombogram assay. In the control experiment TG was triggered using PPP-Reagent 5 pM TF. The lag-time of thrombin generation, the maximum concentration of thrombin (Peak), and the mean rate index (MRI) of the propagation phase of thrombin generation were analysed [MRI=Peak/(ttPeak-lag-time)]. The IC50 of the studied compounds were calculated by extrapolation from the concentration-response curve, and compared. Results All studied antithrombotic agents inhibited in a concentration dependent manner the thrombin generation. The three studied agents significantly inhibited TG at plasma concentrations usually achieved at doses for thromboprophylaxis. However the presence of cancer cells and their type was determinant for the antithrombotic potency of the studied drugs. Comparison on the basis of IC50 showed that the presence of either BXPC3 or MCF7 cells did not significantly modify the antithrombotic potency of enoxaparin as compared to the control experiment. In contrast the inhibitory effect of fondaparinux and apixaban on thrombin generation was partially reversed when TG was triggered by BXPC3 since the IC50 were significantly increased as compared to the control experiment. The presence of MCF7 cells did not significantly modify the antithrombotic activity of apixaban and fondaparinux. Conclusions The LMWH enoxaparin and the specific FXa inhibitors apixaban and fondaparinux demonstrate potent inhibitory capacity on thrombin generation triggered by cancer cells. The type of cancer cells is determinant for the antithrombotic efficiency of the specific factor Xa inhibitors independently of their mechanism of action (i.e. antithrombin dependent or independent inhibition of FXa). In contrast the type of cancer cells does not significantly influence the potency of enoxaparin. This is probably due to the presence of the trace amounts of anti-IIa activity. The present study stresses out that the impact of the type of cancer cells on the antithrombotic activity of the specific Xa inhibitors should not be neglected. These data have to be taken into consideration for the design of dose-finding studies of the direct orally active FXa inhibitors in patients with different histological types of cancer. Disclosures: No relevant conflicts of interest to declare.

Description: Background: Multiple myeloma (MM) and plasma cell dyscrasias (PCD) are associated with increased risk of venous thromboembolism (VTE) which is further enhanced by treatments with immunomodulatory agents, melphalan or steroids. The optimization of VTE prevention in patients with MM is an unmet need. According to Virchow's triad, blood hypercoagulabilty is one of the three conditions required for thrombosis. Elaboration of a risk assessment model, which includes biomarkers of hypercoagulability, could lead to better identification of MM patients eligible for pharmacological thromboprophylaxis. Aims: We conducted a longitudinal observational study, to explore the relationship of MM with cellular and plasma hypercoagulability aiming to identify the most relevant biomarkers which could be used in the RAM for VTE. Materials and Methods: Newly diagnosed patients (n=72) with PCD were recruited from July 2014 to May 2015. The control group (CG) was consisted of 30 healthy age and sex-matched individuals. A systematic compression ultrasound was performed at baseline and 6 months post-therapy. Blood samples were obtained at time of diagnosis and at 3-6 months post therapy. Samples of platelet poor plasma were assessed for thrombin generation (TG) with the PPP-Reagent® (5pMTF and 4μM phospholipids), P-selectin, D-dimers (D-Di), activated FVII (FVIIa), Tissue Factor (TFa), fibrin monomers (FM), and procoagulant phospholipid dependent clotting time (PPL-ct). The upper and lower normal limits (UNL and LNL) were calculated by the mean±2sd. We present herein the data obtained at the inclusion of the patients. Results: Forty-three patients had MM, 16 had asymptomatic MM (AMM), 13 had MGUS. The median age was 68 years (40-84 years); 44% of patients were males. The median time to follow up was 6 months (range: 2-12 months). Patients with PCD as compared to the CG had significantly shorter PPL-ct, which it has been shown that it is inversely correlated with the concentration of platelet derived procoagulant microparticles. PCD patients had also lower Endogenous thrombin potential (ETP), shorter time to thrombin Peak (ttPeak) and lower mean rate index (MRI) of the propagation phase of TG as compared to the controls. The levels of P-selectin were not significantly different between the two groups. Patients with MM as compared to MGUS and AMM patients had significantly higher levels of TFa, FVIIa, FM, D-Di, Peak and MRI (Table 1). Among MM patients 72% had PPL-ct below the LNL, 62% and 28% had TFa and FM above the UNL respectively, 4% had MRI above the UNL and 29% had MRI below the LNL. Among patients who received therapy, 46% also received thromboprophylaxis with either aspirin (76%, n=22) or LMWH (24%, n=5); all were MM patients who received IMiD-based therapies. During the follow-up period the rate of VTE events in MM patients was 7% (all MM patients under treatment). Conclusion: In patients with PCD increased procoagulant microparticles of cellular origin is a generalized phenomenon. In addition, patients with MM present significant TF pathway activation and increased in vivo TG. A significant part, but not all of the patients present strong signs of plasma hypercoagulability. The finding of high inter-individual variability of TG underlines the heterogeneity of blood coagulation alterations in MM patients. The data of the prospective part of this study will allow to validate the clinical significance of this finding. Table 1. Biomarkers of plasma and cellular hypercoagulablity in newly diagnosed patients with PCD. ETP: endogenous thrombin potential; MRI: mean rate index of the propagation phase of thrombin generation. FM: fibrin monomers Parameters Controls (n=30) MGUS (n=13) AMM (n=16) MM (n=43) PPL (sec) 62.8±8.6 38.1±8.2* 41.3±6.8* 39.9±13.7* P-Selectin (pg/ml) 62660±10390 35127±12462 29697.7±9090.45 36223±13638 FTa (ng/ml) 0.3±0.1 2.1±1.4* 1.9±1.3* 7.5±23.3*$ FVIIa (U/ml) 50.9±10.6 60.2±27.5 64.4±48.6 102.1±203.5*$ FM (μg/ml) 3.5±0.8 8.1±8.4* 5.7±1.7* 17.7±38.3*$ D-Di (μg/ml) 0.3±0.1 1.1±1.1 0.6±0.5 1.8±2.8*$ Thrombin generation Lagtime (min) 2.5±0.4 3.4±1.2 3.43±0.71 4.2±2.3*$ ETP (nM/min) 1496±191 1029±205 1045.87±288.9 1202±516$ Peak (nM) 289±36 168±60 174±64 216±74$ ttPeak (min) 5.3±0.7 7.3±1.8 7.17±1.23 7.1±2.7 MRI 110±24 48±24 52±28 82±44$ *p

Description: Introduction: A major problem associated with breast cancer chemotherapy is the subsequent development of resistance to chemotherapeutic agents (multidrug résistance-MDR). Hypercoagulability, increased risk of venous thromboebolism (VTE) and high rate of failure of the antithrombotic treatment with LMWHs have been observed in patients with disease resistant to the chemotherapy. Aim of the study: In the present study we investigated if acquisition of MDR by breast cancer cells MCF7 is associated with modification of their procoagulant potency and induces alteration of the efficacy of the antithrombotic agents. We evaluated the capacity of wild type MCF7 cells (MCF7/WT) and doxorubicin resistant cells (MCF7/DR) to trigger thrombin generation (TG) and to modify the antithrombotic activity of the LMWH enoxaparin, and the specific direct and indirect FXa inhibitors (apixaban and fondaparinux). Materials and Methods: Pre-treatment of MCF-7 cells for several weeks with increasing concentrations of doxorubicin, induced the acquisition of chemo-resistance phenotype documented by the expression the MDR1-Pgp. Tissue factor (TF) and MDR1-Pgp expression by MCF7/WT and MCF7/DR cells were assessed by flow cytometry and western blot assays. Reverse transcriptase - polymerase chain reaction (RT-PCR) for TF mRNA expression was also performed. Thrombin generation of normal platelet poor plasma (PPP) in the presence of MCF7 cells was assessed with the Calibrated Automate Thrombogram® assay (Diagnostica Stago). TG in the presence of MCF7 cells was also assessed in PPP spiked with clinically relevant concentrations of enoxaparin, apixaban or fondaparinux. The efficiency of the studied agents in the presence of MCF7 cells was compared to that in the control experiment where TG was triggered by PPP-Reagent® 5 pM TF. The antithrombotic agents were compared on the bases of the concentration which inhibited 50% TG (IC50). Results: The MCF7/DR cells showed higher expression of TF as compared to MCF7/WT. The TF expression by MCF7/DR was correlated with the expression of the MDR1/P-gp. The MCF7/DR cells significantly enhanced TG as compared to MCF7/WT cells. The three studied agents significantly inhibited TG at plasma concentrations usually achieved at doses for thromboprophylaxis. The presence resistant MCF7 cells did not significantly modify the antithrombotic potency of fondaparinux. The inhibitory effect of enoxaparin and apixaban on thrombin generation was partially reversed when TG was triggered by MCF7/DR as compared to that triggered by MCF7/WT or in the control experiment. Table 1. Modification of the inhibitory potency of the antithrombotic agents by MCF7 cells on thrombin generation in function of the acquisition of MDR phenotype. IC50 for MRI PPP + TF/PPL PPP + MCF7/WT PPP+ MCF7/DR fondaparinux (anti-Xa IU/mL) 0.21 ± 0.03 0.28 ± 0.06 0.30 ± 0.04 enoxaparin (anti-Xa IU/mL) 0.10 ± 0.02 0.12 ± 0.05 0.23 ± 0.08* apixaban (ng/ml) 14 ± 0.9 19 ± 3.1 26 ± 1.4* *p

Description: Abstract 1119 Introduction Diagnosis of heparin-induced thrombocytopenia (HIT) in intensive care unit (ICU) patients represents a major challenge mainly because both the use of unfractionated heparin and the presence of thrombocytopenia are quite common. Despite the existence of several laboratory tests, accurate and prompt HIT diagnostics remains difficult. The ideal combination of an immunological and a functional test is restrictedto specialized laboratories, due to the complexity of the latter. We are in need of an easy-to-perform, widely accessible, rapid and reliably assay. Aim of the study To prospectively evaluate the performance of the latelar-flow immunoassay STic HIT Expert® (Diagnostica Stago, France) for the detection in ICU patients suspected for HIT. Patients-methods Seventy two patients (40 males/32 females) hospitalized in ICU from January to June 2012 were included. Thirty one patients presented with sepsis, 27 underwent extracorporeal circulation (ECC), 21 were hemodialysed and 3 patients were receiving chemotherapy. Sixty one patients were treated with unfractionated heparin and 11 patients received low molecular weight heparin (LMWH). A 4T's score was performed for all patients. All samples were tested in polyspecific ELISA (Zymutest Hyphen Biomed, Neuville-Sur-Oise, France), STic HIT Expert® (Diagnostica Stago, France) and serotonin release assay. In case of a positive polyspecific ELISA, IgG, IgM et IgA isotypes were also performed. Sensitivity, specificity, positive and negative predictive values (PPV an NPV) of STic HIT Expert® were determined against SRA. Results All three tests (polyspecific ELISA, STic HIT Expert®, SRA) were negative in forty patients and had a low HIT suspicion (4T's score: 0–4). In 10 out of 72 patients polyspecific all immunological tests and SRA were positive and HIT suspicion was intermediate or high (4T's score: 4–7). In 9 patients, ELISA tests and STic HIT Expert® were positive but SRA was negative. These patients had a low HIT suspicion (4T's score: 1–4) and underwent ECC (6 out of 9), were hemodialyzed (3 out of 9) or complicated by sepsis (2 out of 9). On the other hand 13 out of 72 patients had ELISA tests positive but STic HIT Expert® and SRA negative. The prevalence of sepsis was high in these patients (8 out of 13), 3 patients underwent ECC and one patient was hemodialysed. STic HIT Expert®, polyspecific and IgG specific ELISA had an excellent sensitivity and negative predictive value at 100%. Moreover STic HIT Expert® was associated with a smaller number of false positive results than the ELISAs. (Table) Conclusion STic HIT Expert® has an excellent performance with a high negative predictive value (100%) and a satisfactory specificity (85%). Less false positive results are detected with STic HIT Expert® than with polyspecific and IgG specific ELISAs. Moreover the test offers a shorter turnaround than ELISA tests and is an easy-to-use single sample test. These characteristics could help avoid HIT over diagnosis in ICU patients. Disclosures: No relevant conflicts of interest to declare.

Description: Background . The protection rights of LMWHs expired or are expiring, so the extent and nature of the studies required to obtain a market authorization for LMWH copies is an issue of debate. The actual situation, which is characterized by the increasing number of enoxaparin copies available worldwide, makes the definition of criteria for the biological similarity between LMWHs copies and the original product a real need. Aim . The present study, conducted in vitro, offers an analytical approach for the comparison of branded enoxaparin and its copies using a standard pharmacological assay (the measurement of the specific anti-Xa activity) and the calibrated automated thrombogram performed in human platelet rich or platelet poor plasma (PRP and PPP) as well as in an experimental model of hypercoagulability induced by cancer cells. The study aims to provide an integrated rational for the determination of the criteria for the similarity and sameness of branded and copies of enoxaparin. Methods . The studied copies of enoxaparin were Cutenox®, Dilutol®, Enoxa®, Fibrinox®, Loparin®, Lupenox®, Novex®, Noxprin®, Versa®. Samples of PPP and PRP from 15 volunteers were spiked with 2-20 µg/ml of branded (Lovenox®) or copies of enoxaparin. The specific anti-Xa activity in PPP was measured with the Rotachrom® assay. Thrombin generation in PPP and PRP in the presence of tissue factor or pancreatic cancer cells BXPC3 was assessed with the Calibrated Automated Thrombogram. Among thrombogram parametersn the Mean Rate Index (MRI) of TG propagation phase of thrombin generation was analysed. The concentrations inhibiting 50% (IC50) the MRI were calculated. Results . The specific inhibitory potency of enoxaparin copies against factor Xa ranged from 0.072 to 0.088 anti-Xa IU/μg, being lower as compared to that of the branded enoxaparin (0.095 anti-Xa IU/μg). the branded enoxaparin induced a concentration dependent inhibition of thrombin generation. The IC50 values revealed significant differences among the studied copies of enoxaparin and between each one of them and the branded enoxaparin. The potency of each copy of enoxaparin to inhibit thrombin generation as compared to the branded product varied in the three experimental systems. the presence of platelets or pancreatic cancer cells in human plasma induces significant modifications of the inhibitory potency of enoxaparin copies on thrombin generation, which distinguish them not only from the branded enoxaparin but also among them. Conclusion . the copies of enoxaparin which respond to the criteria of sameness to the branded enoxaparin according to the pharmacological criteria endorsed by health authorities demonstrate a significant variability regarding their inhibitory potency on thrombin generation. The presence of platelets and cancer cells significantly influence the variability of the antithrombotic efficiency of the enoxaparin copies as compared to the branded product. The findings of the present study underline the need for the elaboration of functional criteria which evaluate the global antithrombotic capacity of enoxaparin copies in order to evaluate their potential sameness with the branded drug. Disclosures No relevant conflicts of interest to declare.

Description: Background: Research has focused on the implications of platelet, endothelial cell and blood coagulation activation in the risk of venous thromboembolism (VTE) in patients with multiple myeloma (MM). The reciprocal interaction between cancer cells and blood coagulation renders biomarkers of hypercoagulability as potential candidates for the evaluation of resistance to treatment in patients with newly diagnosed multiple myeloma (NDMM). Aims: The aim of the current analysis of the ROADMAP-MM study was to identify among a large number of hypercoagulability biomarkers in NDMM patients the most clinically relevant for the inclusion in treatment-resistance assessment tools. Methods: The ROADMAP-MM-CAT study (NCT03405571) was an investigator initiated, prospective, non-interventional trial. Newly diagnosed, treatment naïve symptomatic patients with multiple myeloma (based on 2014 IMWG Criteria) (n=144) were enrolled prior to treatment initiation and response to treatment was assessed at 3 months. Selected biomarkers , such as STA-Procoag-PPL®, factor VIIa factor V, antithrombin, fibrin monomers, thrombomodulin, free TFPI, D-Dimers, P-selectin, heparanase and thrombin generation (Calibrated Automated Thrombogram® and PPP-Reagent®) were measured. Results: At inclusion no patient had received any anti-myeloma treatment or thromboprophylaxis. Median age was 66.0±11.6 (36-86) years and 53% of the population was male. Regarding ISS disease stage: 32% were ISS-I, 23% ISS-II and 45% ISS-III. High risk cytogenetics [defined as presence of any of t(4;14), t(14;16) or del17p by FISH] were detected in 27% of the patients. In 32% of patients the treatment was immunomodulatory drug (IMiD) (90% lenalidomide and 10% thalidomide) and in 64% proteasome inhibitor (PI) based whereas 4% of patients received other regimens. Before any treatment administration - patients showed significantly increased levels of TFa, FVIIa, D-Dimers and FM and significantly shorter Procoag-PPL® as compared to the group of healthy individuals. Levels of P-selectin and TM were significantly lower in patients as compared to healthy individuals. Overall, thrombin generation was attenuated in patients compared to healthy individuals. Lag-time and ttPeak were significantly increased and Peak, MRI and ETP were significantly lower as compared to the group of healthy individuals At 3 months from treatment initiation, 23% (n=33) of the patients showed poor response or resistance to the anti-myeloma treatment. Among them, 7.6% (n=11) had progressive disease, 9.7% (n=14) had stable disease and 5.7% (n=8) had minor response. Among responders (n=111) 42.3 % (n=61) achieved PR and 34.7% (n=50) had VGPR. Multivariate logistic regression analysis for demonstrated that Procoag-PPL®≥41.7 versus