Procoagulant Microparticles Derived from Myeloma Plasma Cells Have a Determinant Role in the Hypercoagulable State Associated with Multiple Myeloma. a Modelization in Vitro Study

Background: Multiple myeloma (MM) is a plasma cell malignancy characterized by bone marrow infiltration leading to multiple lytic bone lesions, renal failure, anemia and increased rate of thrombotic events. The cross-talk between cancer cells, plasma clotting mechanism, platelets and endothelial cells enhances hypercoagulability. The mechanisms by which myeloma plasma cells (MPCs) interfere with their microenvironment during the blood coagulation process have been poorly investigated. Several studies have established a link between levels of cancer cell derived procoagulant microparticles, cancer aggressiveness, response to anticancer treatments well as with the risk of cancer associated thrombosis.^, The mechanisms by which myeloma plasma cells interfere to blood coagulation process in their microenvironment have been poorly investigated.

Aim: Set up an experimental model that allows the identification of the procoagulant fingerprint of MPCs and myeloma cell-derived microparticles (MC-dMPs) as well as their impact on thrombin generation of normal plasma. In addition, we evaluated the relative impact of TF-pathway and the intrinsic clotting pathway on thrombin generation triggered by myeloma plasma cells and MC-dMPs. Finally, we performed a simulation of thrombin generation process in the presence of myeloma plasma cells and MC-dMPs as well as in the presence of varying concentrations of TF and procoagulant phopsholipids, mimicking some of the conditions that potentially exist in MPCs.

Methods. TF and annexin V expression by MPCs (MPC-dMPs) were analyzed by flow cytometry, TF activity (TFa) and TF gene expression was also determined. Thrombin Generation (TG) in the presence of MPCs or MPC-dMPs was assessed with the Calibrated Automated Thrombogram assay (CAT®) in normal human PPP. TG was also assessed in plasma spiked with MPCs and MPC-dMPs or variable concentrations of TF and procoagulant phospholipids.

Results: MPC-dMPs expressed about 8-fold higher levels of TF as compared to MPCs. TFa expressed by MPC- dMPs was significantly higher as compared to that expressed by MPC cells. MPCs and PC- dMPs enhanced thrombin generation of human plasma. Thrombin generation was significantly higher with MPC-dMPs compared to MPCs. MPCs and MPC-dMPs, when concomitantly present in normal PPP, significantly amplified thrombin generation as compared to that observed in the presence of MPCs alone. Presence of TF and procoagulant phospholipids in the microenvironment resulted in significant amplification of thrombin generation induced by MPCs.

Conclusion: This study support the concept that hypercoagulability induced by myeloma plasma cells is the result of the combination of the procoagulant properties of myeloma cells with the procoagulant elements of the plasma microenvironment which consist of procoagulant phospholipids and low TF concentration. Following this concept we assume that MPC-dMPs released by myeloma plasma cells into the microenvironment enhance the weak procoagulant activity of myeloma plasma cells. Whereas this combined effect of MPCs and MPC-dMPs on thrombin generation has an impact on the formation of local fibrin network their critical role into the hypercoagulablity, procoagulant MC-dMP could be a potential target for the evaluation of the aggressivity of MPCs and of their response to anti-myeloma treatment.