The PKB inhibitor Akti-1/2 potentiates PAR-1-mediated platelet function independently of its ability to block PKB

Background: The role of protein kinase B (PKB) in platelet function remains unclear, primarily due to the absence of highly selective small-molecule inhibitors and limitations in genetic models. Recently, a selective, non-ATP-competitive PKB inhibitor known as Akti-1/2 was developed, but its effectiveness and specificity in inhibiting PKB activation in platelets have not been thoroughly investigated.

Objective and Methods: This study aimed to evaluate the impact of the PKB inhibitor Akti-1/2 on PKB activation and platelet function using Western blotting and aggregometry/flow cytometry techniques.

Results: Akti-1/2 effectively inhibited thrombin-induced PKB phosphorylation at Thr(308) and Ser(473), as well as the phosphorylation of its downstream target, GSK3beta, while having minimal impact on the phosphorylation of pleckstrin, p38, ERK, and JNK. Unexpectedly, Akti-1/2 significantly enhanced PAR-1-mediated platelet aggregation. This enhancement persisted even in the presence of PI3 kinase inhibitors, suggesting a PKB-independent mechanism. The increased aggregation was associated with elevated intracellular calcium levels, PKC activation, and degranulation, and it was suppressed by agents that antagonize this pathway.

Conclusions: The PKB inhibitor Akti-1/2 enhances PAR-1-mediated platelet responses through a PKB-independent mechanism that is dependent on calcium and PKC. This strong and rapid effect may have implications for the therapeutic use of Akti-1/2 and related compounds.