Effects of the Btk-Inhibitors Remibrutinib (LOU064) and Rilzabrutinib (PRN1008) With Varying Btk Selectivity Over Tec on Platelet Aggregation and in vitro Bleeding Time.

Rundan Duan,Wolfgang Siess,Luise Goldmann,Philipp Von Hundelshausen,Michael Spannagl,Christian Weber,Richard Brandl

doi:10.3389/fcvm.2021.749022

Abstract

Background: Bruton tyrosine kinase inhibitors (BTKi) are used in B-cell malignancies and in development against various autoimmune diseases. Since Btk is also involved in specific pathways of platelet activation, BTKi might be considered to target platelet GPVI/GPIb-mediated atherothrombosis and platelet FcγRIIA-dependent immune disorders. However, BTKi treatment of patients with B-cell malignancies is frequently associated with mild bleeding events caused possibly by off-target inhibition of Tec. Here, we compared the platelet effects of two novel BTKi that exhibit a high (remibrutinib) or low (rilzabrutinib) selectivity for Btk over Tec.Methods and Results: Remibrutinib and rilzabrutinib were pre-incubated with anticoagulated blood. Platelet aggregation and in vitro bleeding time (closure time) were studied by multiple electrode aggregometry (MEA) and platelet-function analyzer-200 (PFA-200), respectively. Both BTKi inhibited atherosclerotic plaque-stimulated GPVI-mediated platelet aggregation, remibrutinib being more potent (IC50 = 0.03 μM) than rilzabrutinib (IC50 = 0.16 μM). Concentrations of remibrutinib (0.1 μM) and rilzabrutinib (0.5 μM), >80% inhibitory for plaque-induced aggregation, also significantly suppressed (>90%) the Btk-dependent pathways of platelet aggregation upon GPVI, von Willebrand factor/GPIb and FcγRIIA activation stimulated by low collagen concentrations, ristocetin and antibody cross-linking, respectively. Both BTKi did not inhibit aggregation stimulated by ADP, TRAP-6 or arachidonic acid. Remibrutinib (0.1 μM) only slightly prolonged closure time and significantly less than rilzabrutinib (0.5 μM).Conclusion: Remibrutinib and rilzabrutinib inhibit Btk-dependent pathways of platelet aggregation upon GPVI, VWF/GPIb, and FcγRIIA activation. Remibrutinib being more potent and showing a better profile of inhibition of Btk-dependent platelet activation vs. hemostatic impairment than rilzabrutinib may be considered for further development as an antiplatelet drug.

Highlights

Since the first description of a patient with recurrent infections and deficiency of immunoglobulins termed “Agammaglobulinemia” by Ogden Bruton in 1952 [1], it took more than 40 years of research, until Bruton tyrosine kinase (Btk) was identified in 1993 as the responsible protein that is deficient in patients with X-linked agammaglobulinemia [2, 3]
Btk belongs to the Tec family of non-receptor cytoplasmic tyrosine kinase, and contains five different protein interaction domains: an amino terminal pleckstrin homology (PH) domain, a proline-rich Tec homology (TH) domain, the SRC kinase homology (SH) domains SH2 and SH3, and a kinase domain [4]
Our results show that the remibrutinib concentration to fully inhibit Btk-dependent pathways of platelet aggregation (0.1 μM) is lower than the reported maximal plasma level (0.46 μM) in a phase I study after intake of 100 mg q.d. for 12 days [38]

Summary

Introduction

Since the first description of a patient with recurrent infections and deficiency of immunoglobulins termed “Agammaglobulinemia” by Ogden Bruton in 1952 [1], it took more than 40 years of research, until Bruton tyrosine kinase (Btk) was identified in 1993 as the responsible protein that is deficient in patients with X-linked agammaglobulinemia [2, 3]. Since the approval of ibrutinib, the covalent irreversible first in class Btk inhibitor (BTKi) in 2013 for treatment of certain B-cell malignancies, many more reversible and irreversible BTKi have evolved and the spectrum of diseases that are targeted extends from specific forms of B-cell malignancies to various autoimmune disorders [5]. Bruton tyrosine kinase inhibitors (BTKi) are used in B-cell malignancies and in development against various autoimmune diseases. Since Btk is involved in specific pathways of platelet activation, BTKi might be considered to target platelet GPVI/GPIb-mediated atherothrombosis and platelet FcγRIIA-dependent immune disorders. BTKi treatment of patients with B-cell malignancies is frequently associated with mild bleeding events caused possibly by off-target inhibition of Tec. Here, we compared the platelet effects of two novel BTKi that exhibit a high (remibrutinib) or low (rilzabrutinib) selectivity for Btk over Tec

Methods

Results

Discussion

Conclusion