Defibrotide Blunts the Prothrombotic Effect of Thalidomide on Endothelial Cells

Abstract

Patients with multiple myeloma (MM) are at relatively high risk of developing thromboembolic events such deep venous thrombosis (DVT) where thalidomide therapy has been identified to increase this risk. Defibrotide (DF), a polydisperse oligonucleotide, showed previously to counteract the alterations in endothelial cells (ECs) induced by lipopolysaccharide. It prompts us to investigate the impact of thalidomide on ECs and whether DF modulates changes in fibrinolysis induced by thalidomide. In this in vitro study, MM by itself alters the profibrinolytic potential of ECs decreasing the tissue plasminogen activator (t-PA) and increasing the plasminogen activator inhibitor 1 (PAI-1) levels which is potentiated by thalidomide. Defibrotide was able to counteract these effects. Additionally, DF upregulated the t-PA and downregulated PAI-1 gene expression modulated by thalidomide. Defibrotide also protects ECs from thalidomide-mediated cell death without interfering with its antitumor effects. These findings support DF clinical use for the prevention of DVT induced by immunomodulatory drugs.

Defibrotide: Properties and clinical use of an old/new drug

“…The 1 year progression-free survival and 1-year overall survival rates were 34% and 90% respectively (Palumbo et al, 2010). These data were confirmed by an "in vitro" study in which DFT was able to protect endothelial cells by thalidomide-mediated cell death without interfering with its anti-tumor effect (Echart et al, 2012).…”

“…DFT has anti-thrombotic-thrombolytic activity in several different experimental models (Fumagalli et al, 1987;Fumagalli et al, 1989;Giedrojc and Breddin, 1991;Grodzinska et al, 1987;Niada et al, 1981Niada et al, , 1982Paul et al, 1993;Tettamanti et al, 1992). DFT increases TM (Zhou et al, 1994), protein C plasmin activity, t-PA (Echart et al, 2012;Klocking, 1992), TFPI (Cella et al, 2001), which is also endowed with anti-inflammatory properties effects (Bajaj et al, 2001), PGI2 (Berti et al, 1988(Berti et al, , 1990aHohlfeld et al, 1992;Lobel and Schror, 1985;Rossoni et al, 2006), PGI2 receptors on the surface of platelets and their affinity for PGI2 (Lobel and Schror, 1989), production of NO (Masini et al, 1995) and nitric oxide synthase (NOS) activity. DFT decreases PAF (Berti et al, 1990a), PAI (Klocking, 1992), TF (Falanga et al, 2003;Francischetti et al, 2012), which represents the link between inflammation and thrombosis (Penn and Topol, 2001) and TXA2 (Bracht and Schror, 1994;Hohlfeld et al, 1992).…”

Defibrotide: A Review of Its Use in Severe Hepatic Veno-Occlusive Disease Following Haematopoietic Stem Cell Transplantation

“…Increased t-PA antigen levels and t-PA activity in resting HMECs and HUVECs [21] Counteracted LPS-induced increases in PAI-1 antigen levels and reductions in t-PA activity in LPS-stimulated HMECs and HUVECs [21] Counteracted thalidomide-induced reductions in t-PA antigen levels in thalidomide-stimulated HMECs [26] Counteracted thalidomide-induced reduction in the ability of HMECs to degrade a fibrin clot [26] Increased plasma t-PA release [49], increased plasma t-PA activity [52] and decreased PAI-1 activity [49,53] in rats and/or mice Increased t-PA antigen levels and decreased PAI-1 antigen levels in primates [54] Stimulated TM expression and activity in HUVECs [55] Stimulated plasmin activity in a dose-dependent manner [56] Counteracted LPS-induced increases in TF antigen levels and TF procoagulant activity in LPS-stimulated HMECs [21] Inhibited PAF formation in a rabbit model of myocardial ischaemia [57] Inhibited thrombin-induced platelet aggregation [58] and protected mice against thrombin-induced thromboembolism [59] Inhibited platelet activation by cathepsin G derived from stimulated polymorphonuclear leukocytes [60] Attenuated increased reactivity (in response to sera from HSCT pts) of HMEC-and HUVEC-generated ECM to platelets [61] Attenuated increased reactivity (in response to ciclosporin) of HMEC-generated ECM to platelets [62] Weak/negligible anticoagulant activity in plasma and whole blood assays [22] Effects on prostanoids Induced PGE 2 release in a rabbit model of myocardial ischaemia [63] and increased urinary PGE 2 levels in rats and mice [52] Induced 6-keto-PGF 1a release in a rabbit model of myocardial ischaemia [57,63], in a rat model of myocardial ischaemia [64], in leukocyteperfused isolated rabbit hearts [23] and in human saphenous vein samples [65] Reduced plasma TXB 2 levels in a rat model of myocardial ischaemia [64] and inhibited thrombin-induced TXB 2 formation in platelets [58] Effects on inflammatory cytokines/mediators…”

“…inhibiting the formation of cysteinyl leukotrienes, which increase vascular permeability [23]) and improving vascular tone (Table 1). In addition, defibrotide protected endothelial cells against the cytotoxic effects of inflammatory cytokines such as tumour necrosis factor-a [24], as well as protecting HMECs against fludarabine-mediated apoptosis [25] and thalidomide-induced cell death [26] ( Table 1). Defibrotide also increased the tubular morphogenesis of HMECs, suggesting that it may promote hepatic revascularization in VOD [27] (Table 1).…”

“…Binds with high affinity to the heparin-binding protein bFGF and mobilizes bFGF from its low-affinity binding sites on ECM [27] Promoted HMEC cell mitogenesis and tubular morphogenesis and may promote endothelial repair and revascularization [27] Reduced the generation of oxygen free radicals by monocytes [70] Decreased heparanase expression and activity in multiple myeloma cells [66] Protected HMECs against fludarabine-mediated apoptosis [25] and thalidomide-induced cell death [26] bFGF basic fibroblast growth factor, ECM extracellular matrix, HMEC human microvascular endothelial cell, HSCT haematopoietic stem cell transplantation, HUVEC human umbilical vein endothelial cell (macrovascular endothelial cell), ICAM intercellular adhesion molecule, IL interleukin, 6-keto-PGF 1a 6 keto-prostaglandin-F 1a , LFA lymphocyte function associated antigen, L-NAME N G nitro-L-arginine methylester, LPS lipopolysaccharide, MHC major histocompatibility complex, NFjB nuclear factor jB, PAF platelet activating factor, PAI plasminogen activator inhibitor, PGE 2 prostaglandin E 2 , pts patients, TF tissue factor, TFPI tissue factor pathway inhibitor, TM thrombomodulin, TNF tumour necrosis factor, t-PA tissue plasminogen activator, TXB 2 thromboxane B 2 a In vitro studies, unless specified otherwise. Not all studies used the approved defibrotide preparation derived from porcine intestinal mucosal DNA (e.g.…”