Cancer Cell-Derived Extracellular Vesicles Are Associated with Coagulopathy Causing Ischemic Stroke via Tissue Factor-Independent Way: The OASIS-CANCER Study

Abstract

Background: Cancer and stroke, which are known to be associated with one another, are the most common causes of death in the elderly. However, the pathomechanisms that lead to stroke in cancer patients are not well known. Circulating extracellular vesicles (EVs) play a role in cancer-associated thrombosis and tumor progression. Therefore, we hypothesized that cancer cell-derived EVs cause cancer-related coagulopathy resulting in ischemic stroke.

Methods: Serum levels of D-dimer and EVs expressing markers for cancer cells (epithelial cell adhesion molecule [CD326]), tissue factor (TF [CD142]), endothelial cells (CD31+CD42b-), and platelets (CD62P) were measured using flow cytometry in (a) 155 patients with ischemic stroke and active cancer (116 - cancer-related, 39 - conventional stroke mechanisms), (b) 25 patients with ischemic stroke without cancer, (c) 32 cancer patients without stroke, and (d) 101 healthy subjects.

Results: The levels of cancer cell-derived EVs correlated with the levels of D-dimer and TF+ EVs. The levels of cancer cell-derived EVs (CD326+ and CD326+CD142+) were higher in cancer-related stroke than in other groups (P<0.05 in all the cases). Path analysis showed that cancer cell-derived EVs are related to stroke via coagulopathy as measured by D-dimer levels. Poor correlation was observed between TF+ EV and D-dimer, and path analysis demonstrated that cancer cell-derived EVs may cause cancer-related coagulopathy independent of the levels of TF+ EVs.

Conclusions: Our findings suggest that cancer cell-derived EVs mediate coagulopathy resulting in ischemic stroke via TF-independent mechanisms.

Fig 1. Flow cytometry and Western blot test results.

(A) Flow cytometry results using size beads showed that most circulating cancer-derived extracellular vesicles (EVs) were distributed with a size between 200nm-1,000 nm. (B) Most EVs were degradated after treatment with 0.1% triton-X100. (C) Western blot test showed that EVs expressed flotillin-1.

Fig 2

D-dimer levels (A) and cancer-cell derived EV levels (B–E) among the groups.

Fig 3. Results of Path analyses.

(A) Path analysis using cancer-related stroke as the outcome, cancer cell-derived EVs as a predictor, and cancer-related coagulopathy as measured by D-dimer levels as a mediator. (B) Path analysis for cancer cell-derived EVs and cancer-related coagulopathy with tissue factor-bearing EVs (cancer cell-derived EVs, platelet-derived EVs, and endothelial-derived EVs) as mediators. N/A, no association. Numbers are β-coefficients that were statistically significant.