Stable thrombus formation on irradiated microvascular endothelial cells under pulsatile flow: Pre-testing annexin V-thrombin conjugate for treatment of brain arteriovenous malformations

Abstract

The annexin V-thrombin conjugate induced rapid thrombosis (fibrin deposition) on irradiated endothelial cells under shear stress in the parallel-plate flow device. Unconjugated, non-targeting thrombin did not induce fibrin deposition. A synergistic interaction between radiation and conjugate dose was observed. Thrombosis was greatest at the highest combined doses of radiation (25 Gy) and conjugate (2.5 μg/mL). The parallel-plate flow system provides a rapid method to pre-test pro-thrombotic vascular targeting agents. These findings validate the translation of the annexin V-thrombin conjugate to pre-clinical studies.

The Evolution of Safe and Effective Coaguligands for Vascular Targeting and Precision Thrombosis of Solid Tumors and Vascular Malformations

“…While PS targeting in tumors has been more avidly pursued in the context of cancer immunotherapy [35], it has been investigated as a radiation-stimulated target for vascular infarction in brain AVMs [36][37][38][39][40]. Thorpe and colleagues demonstrated that translocation of this phospholipid to the surface of tumor vessels could be enhanced in response to oxidative stress, via stimulation by growth factors, hydrogen peroxide, and ionizing radiation [28,41,42].…”

“…Subsequent studies then used a ligand-directed approach to improve targeted occlusion of the larger AVM vessels. Using a PS-targeting annexin V protein conjugated to thrombin as an effector, it was first established in vitro that rapid and selective thrombosis could be induced on an irradiated endothelial cell layer using whole human blood circulating in a parallel-plate flow system [40]. In vivo studies subsequently demonstrated that significant occlusion of the rat model AVM vessels could be achieved with this coaguligand [46].…”

Modeling blood–brain barrier pathology in cerebrovascular disease in vitro: current and future paradigms

“…Addition of flow and physiological shear stress (~ 5 dyne/cm 2 ) to BEC monolayers in a parallel plate flow chamber (PPFC) is a step forward for building 3-D BBB models. The PPFC platform consists of a polycarbonate chamber which mimic the lumen of blood vessels and BECs plated on the glass coverslip [124,125]. The culture media flow from one side to other in a flow chamber.…”

“…The culture media flow from one side to other in a flow chamber. The PPFC platform is widely used in vascular biology to study leukocyte-endothelial cell interaction, tumor-endothelial cell interaction, cellular chemotaxis, endothelial cells and drug resorption [125]. The major limitation is an inability to form perivascular cell-BEC interaction and reproduce BBB complexity.…”

“…There are still no studies regarding ischemic injury in the μBBB model, although the μBBB is currently utilized to investigate the hemodynamic effect of thrombosis and microvascular occlusion in hematological diseases (i.e. Sickle cell disease) [125].…”