Introduction: Direct oral anticoagulants (DOACs) are the standard care for stroke prevention in
non-valvular atrial fibrillation and managing venous thromboembolism. DOACs can cause
major bleeding episodes in up to 5% of patients, making DOAC reversal strategies crucial
in acute trauma or emergent surgery. Currently, andexanet alfa is the only DOAC reversal
option for FXa dependent DOACs. However, major clinical limitations are known, including
elimination of physiological anticoagulant protection and triggering prothrombotic
adverse reactions. Additionally, its high costs makes provide a drawback. Here, we
studied the effects of alternative, DOAC-reversal nanobodies on coagulation under
flow. We aim to evaluate the capability of novel DOAC-reversing nanobodies to normalize
anticoagulant effects on the platelet clot formation under flow using a novel microfluidic
device.
Method: A novel microfluidic blood perfusion system was employed operating at wall shear
rate (1000 s-1) and at 37°C. Platelet-thrombus formation was evaluated from collagen/tissue factor
surfaces by automated capturing of brightfield and fluorescence images during clot
formation. Healthy donor blood was compared with DOAC-treated blood (300 nM edoxaban
or 600 nM rivaroxaban) in presence or absence of nanobodies against specific DOACs
(600 nM) or andexanet (600 nM).
Results: Blood treatment with either DOAC did not affect platelet deposition, but decreased
thrombus contraction by 20-27% (P<0.05). Simultaneously, both treatments delayed and
impaired fibrin formation, reducing fibrin deposition at 10 minutes by 70%-83%, depending
on the DOAC used (41 SAC% vs. 8.2-14.5 SAC%, P=0.012-0.049). Each nanobody against
the respective DOAC counteracted the inhibitory effects and essentially restored coagulant
function (8.2 SAC% vs 41.7-44.5 SAC% fibrin deposition, P=0.009-0.047). Andexanet
alfa caused a similar reversal capacity (P=0.016).
Conclusion: This in vitro analysis indicated that the developed nanobodies against rivaroxaban
and edoxaban restore the hemostatic process after DOAC treatment, and hence provide
a promising strategy to reverse DOAC-dependent bleeding complications.
