The Aptamer ARC1779 Potently and Specifically Inhibits the Excessive von Willebrand Factor (vWF) Activity and vWF-Mediated Ex Vivo Platelet Function of Patients with Thrombotic Thrombocytopenic Purpura (TTP).

Abstract

Background: ARC1779 is an aptamer which blocks the binding of the vWF A1 domain to platelet GPIb receptors. In TTP there is an excess of ultra-large multimers of vWF which are especially avid for binding GPIb and give rise to disseminated platelet thrombi which are fibrin-poor and vWF-rich in composition. ARC1779 is being evaluated for use as front-line therapy of acute TTP in conjunction with plasma exchange. ARC1779 has already been demonstrated in healthy volunteers to inhibit vWF activity and vWF-dependent platelet function. ARC1779 has no anticoagulant effect and does not inhibit other pathways of platelet activation. ARC1779 is expected to normalize platelet dysfunction and prevent the thrombotic end-organ complications of TTP based upon the mechanism of action defined for ARC1779 and the mechanism of thrombosis defined for TTP. Methods: We first assessed vWF activity (vWF:RiCO) and platelet function in blood samples taken from TTP patients and age-matched, healthy controls. We then studied the ex vivo dose response curves for ARC1779 on vWF activity (free A1 domain sites) and on platelet function assessed by the Platelet Function Analyzer (PFA-100®), cone and plate analyzer (IMPACT®), and agonist-induced impedence platelet aggregometry (Multiplate®) of TTP patients (N=10, 2 in acute phase and 8 in remission) and healthy age-matched controls (N=23). Results: vWF:RiCO activity (p=0.002) and vWF-dependent platelet plug formation (p=0.001) were increased in TTP patients relative to healthy controls, but agonist-induced platelet aggregation (ADP, arachidonic acid, collagen, TRAP) was not. ARC1779 fully blocked platelet plug formation as measured by PFA-100® with an IC100 of ∼ 1 mcg/mL with citrate anticoagulation, and ∼ 3–4 mcg/mL with hirudin anticoagulation in both TTP patients and in healthy controls. ARC1779 fully blocked shear-dependent platelet adhesion measured by the IMPACT® analyzer with an IC100 of ∼ 1 mcg/mL with citrate anticoagulation in both TTP patients and in healthy controls. ARC1779 fully blocked vWF activity (free A1 domain sites) with an IC90 of ∼ 6 mcg/mL in TTP patients and ∼ 2 mcg/mL in young controls (p<0.001 between groups). ARC1779 did not inhibit platelet aggregation by ADP, collagen or arachidonic acid at concentrations (10mcg/mL) that fully inhibited vWF dependent platelet function. Conclusions: ARC1779 potently and specifically inhibits vWF activity and vWF dependent platelet function in the setting of TTP where vWF activity is increased. ARC1779 represents a novel therapeutic principle (vWF antagonism) and a novel therapeutic class (aptamers) with potential for the treatment of TTP. vWF Activity and Platelet Function in TTP Patients and Age-Matched Controls Population Healthy Controls TTP Patients values shown as mean +/− standard deviation Sample Size N=23 N=10 vWF:RiCO (%) 94.9 +/− 60.4 153.3 +/− 55.9 PFA-100® Closure Time in Citrate (sec) 90.9 +/− 16.0 66.8 +/− 12.7 PFA-100® Closure Time in Hirudin (sec) 84.0 +/− 12.9 64.6 +/− 11.9 ARC1779 IC100 PFA in Citrate (ARC1779 mcg/mL) 0.9 +/− 0.4 1.4 +/− 0.6 ARC1779 IC100 PFA in Hirudin (ARC1779 mcg/mL) 3.2 +/− 1.5 4.4 +/− 2.7 IC100 IMPACT in Citrate (ARC1779 mcg/mL) 0.8 +/− 1.2 0.8 +/− 0.8 IC90 vWF free A1 domain sites (ARC1779 mcg/mL) 1.8 +/− 0.8 6.2 +/− 2.7

Anti–von Willebrand factor aptamer ARC1779 for refractory thrombotic thrombocytopenic purpura

“…Results are reported in percent (%) of normal, relative to a calibrator that has been standardized against the third International Standard for Factor VIII and VWF in Plasma (91/666). VWF activity correlates well with VWF ristocetin cofactor activity under baseline, that is, untreated, conditions 5 …”

“…It specifically inhibits the VWF A1 domain and thus shear‐dependent PLT function as demonstrated by the PLT function analyzer (PFA‐100, Dade Behring, Marburg, Germany). In contrast, PLT aggregation by a variety of agonists is not inhibited 5 . The concentration effect relationship shows a 90% inhibition of VWF (EC 90 ) at mean plasma concentrations of 2‐3 µg/mL, 6 which can easily be achieved.…”

“…No treatments other than plasma exchange have been documented to be effective nor are approved for treatment of acute TTP, but the ability of rituximab to prevent relapsing TTP is currently tested 3,4 . The investigational anti–von Willebrand factor (VWF) aptamer ARC1779 effectively inhibits VWF activity in plasma samples of TTP patients and thus shear‐dependent platelet (PLT) function 5 . As this would offer a new treatment possibility for a critically ill patient, refractory to standard care, we explored the pharmacokinetics, pharmacodynamics, safety, and efficacy of ARC1779.…”