Effects of novel Btk and Syk inhibitors on platelet functions alone and in combination in vitro and in vivo

Abstract

Background: Inhibitors of tyrosine kinases downstream of the B-cell receptor, such as Bruton's tyrosine kinase (Btk) or Spleen tyrosine kinase (Syk), used alone or in combination are new therapeutic options in the treatment of B-cell malignancies. A challenge in the development of second-generation Btk inhibitors is to limit their side effects such as the increased bleeding risk. Considering the pivotal role of Syk in immunoreceptor tyrosine-based activation motif mediated platelet signaling, the impact of inhibiting this kinase on platelet functions is also worth analyzing. Objectives: We investigated the effect of a novel Btk inhibitor, tirabrutinib, and a Syk inhibitor, entospletinib, alone and in combination on platelet signaling and functions in vitro and ex vivo. Methods: Platelet aggregation, secretion, and signaling responses as well as thrombus growth under flow were analyzed in the presence of the inhibitors alone or in combination in vitro, at clinically relevant doses, and ex vivo in patients treated with these inhibitors in the context of a phase I trial. Results: Although tirabrutinib alone had modest effects on platelet activation in vitro and ex vivo, entospletinib alone efficiently inhibited washed platelet aggregation in response to collagen. However, entospletinib weakly affected platelet activation in platelet-rich plasma, in whole blood and ex vivo. Importantly, the combination of tirabrutinib and entospletinib induced a significant decrease in platelet response to collagen in vitro and ex vivo correlating with mild bleedings reported in some of the treated patients. Conclusion: These new results should contribute to improve the safety of these targeted therapies.

Vaccine-Induced Immune Thrombotic Thrombocytopenia (VITT): Targeting Pathomechanisms with Bruton Tyrosine Kinase Inhibitors

“…Similar observations have been made recently with the new Syk inhibitor entospletinib (45), at present in phase 3 clinical trials of AML.…”

“…However, the therapeutic levels of R406 are apparently too low to directly inhibit platelets ex vivo. 44 Similar observations have been made recently with the new Syk inhibitor entospletinib, 45 at present in phase 3 clinical trials of acute myeloid leukaemia (AML).…”

Bleeding by Bruton Tyrosine Kinase-Inhibitors: Dependency on Drug Type and Disease

“…Tirabrutinib was 7–10 times less potent than ibrutinib in inhibiting GPVI-mediated platelet aggregation in hirudin-anticoagulated blood [ 8 , 41 ]. Thrombus formation onto collagen under arterial shear rate was significantly reduced at 2 µM tirabrutinib pre- incubated for 60min with blood in one study [ 93 ], but not in another study with tirabrutinib 2 µM and 5 µM pre-incubated for 15 min with blood [ 8 ]. Tirabrutinib inhibited also ristocetin-induced platelet aggregation and FcγRIIa-stimulated platelet aggregation in whole blood [ 13 , 41 ], and increased at 5 µM (but not 2 µM) in vitro bleeding time as measured by the PFA [ 41 ] ( Table 2 ).…”

“…In patients with B-cell malignancies treated within a clinical trial with a low dose of tirabrutinib (80 mg/day) for 1 month, a tendency of decreased aggregation following collagen 3.3 μg/mL stimulation was observed compared to pretreatment values [ 93 ]. No inhibition of platelet aggregation induced by other platelet stimuli (ADP, TRAP) was found ex vivo similar to the results after tirabrutinib incubation of blood [ 8 ].…”

Fine-Tuning of Platelet Responses by Serine/Threonine Protein Kinases and Phosphatases—Just the Beginning

“…Clinically established cancer therapeutics targeting Y-PKs such as Syk or Btk have been intensively analyzed regarding their antiplatelet properties due to their bleeding risk. [86][87][88] Interestingly, current clinical trials for the approvement of MAPK inhibitors as anticancer drugs revealed that MEK1/2 (ERK1/2) inhibitors are associated with higher bleeding risk than p38 inhibitors. 21 Much less is known about platelet effects of clinically used anticancer drugs targeting the S/T-PP PP2A.…”