Extracellular vesicles, tissue factor, cancer and thrombosis – discussion themes of the ISEV 2014 Educational Day

Figures & data

Fig. 1.  The diverse roles of vesiculation in cancer.

Fig. 2.  Tissue factor initiates coagulation by binding factor VIIa to form membrane bound complex which activates factor X. This forms a complex with factor Va which activates prothrombin to thrombin. Thrombin activates factor XI which creates an amplification loop through the factor IXa/VIIIa activation of factor X. The resulting burst of thrombin generation causes platelet activation and conversion of insoluble fibrinogen to an insoluble thrombin clot.

Fig. 3.  (a) TF-bearing EVs in glioma and meningioma patients and in healthy controls. (b) Origin of TF-bearing EVs in glioma patients and controls.

Fig. 4.  (a) The phosphorylation of Ser253 within the cytoplasmic domain acts to initiate the incorporation and release of TF within EVs. (b) The interaction between TF and filamin-A is required for the active incorporation of TF into EVs.

Fig. 5.  Model for the release of TF-exposing vesicles. TF-exposing vesicles originating from different membrane compartments.

Fig. 6.  TF-dependent coagulation activation is of relevance for metastasis, tumour growth, and angiogenesis through several distinct mechanisms, including the activation of thrombin further resulting in the generation of a fibrin network, and platelet activation. These end stages of coagulation can contribute to tumour progression and metastasis by, for example, protection of circulating cancer cells from immune cell attack. In addition, TF and associated coagulation proteases up-stream of thrombin can cleave and activate protease-activated receptors (PARs) to induce pro-migratory and survival signalling in cancer cells. Endothelial cells, however, are normally devoid in TF. At hypoxic conditions, cancer cells may release TF/FVIIa-bearing EVs that in a paracrine manner trigger an angiogenic response through activation of PAR-2 in endothelial cells. Further, it may be hypothesized that systemic release of TF-EVs from hypoxic tumour regions contributes to the hypercoagulable state of cancer patients.

Table I. Issues with measurement of MV-associated TF.

Pre-analytical variables – cell-free plasma, anticoagulant, g-values for centrifugation, discard tube in blood draw, freeze thawing
TF activity can be increased by post-translational modification and decreased by its inhibitor tissue factor pathway inhibitor
Higher concentrations of Factor VIIa can activate FX in a TF-independent manner in the presence of phospholipids
Binding of Factor VII/VIIa to TF blocks access of some anti-TF antibodies
The contact pathway may activate coagulation in some clotting-based assays with low TF as initiator. This pathway can be inhibited
Presence of antibody microaggregates causing false positive signals in flow cytometry studies
Detection limit of most flow cytometers above the size of small TF-positive vesicles
Few studies simultaneously compare more than one assay
No accepted international standard
No gold standard assay – the MP TF activity assay is the best available assay
Positive (plasma from whole blood treated with LPS) and Negative Controls (use of inhibitory antibodies are important to confirm that the procoagulant activity measured is due to TF)