Ibrutinib, but not zanubrutinib, induces platelet receptor shedding of GPIb-IX-V complex and integrin αIIbβ3 in mice and humans

The Bruton’s tyrosine kinase (BTK) inhibitor, ibrutinib, has proven to be efficacious in the treatment of B-CLL and related diseases. However, a major adverse side effect of ibrutinib is bleeding including major haemorrhages. The bleeding associated with ibrutinib use is thought to be due to a combination of on-target irreversible Btk inhibition as well as off targeted inhibition of other kinases including EGFR, ITK, JAK3 and Tec kinase. In this study, we investigated the effects of ibrutinib versus zanubrutinib (a more selective Btk inhibitor) on platelet activation, glycoprotein expression and thrombus formation. Ibrutinib, but not zanubrutinib, induced a time and dose dependent shedding of GPIb-IX complex and integrin αIIbβ3 but not GPVI and GPV from the platelet surface. The shedding of GPIbα and GPIX was blocked by GM6001 and TAPI-2, an ADAM17 inhibitor but not ADAM10 inhibitor. Ibrutinib but not zanubrutinib treatment of human platelets increased ADAM17 activation. Pretreatment of C57BL/6 mice with ibrutinib (10 mg/kg) but not zanubrutinib (10 mg/kg) inhibited ex vivo and in vivo thrombus growth over time. Platelets from ibrutinib-treated CLL patients showed reduced GPIb-IX complex and integrin αIIbβ3 surface expression and reduced ex vivo thrombus formation under arterial flow, which was not observed in zanubrutinib-treated patients. Ibrutinib but not zanubrutinib treated mice led to increased soluble GPIbα and soluble αIIb levels in plasma. These data demonstrate that ibrutinib induces shedding of GPIbα and GPIX by an ADAM17-dependent mechanism and integrin αIIbβ3 by an unknown sheddase, and this process occurs in vivo to regulate thrombus formation.