PB0892 - C-3399 inhibitor of PDI isoforms with an attractive profile of antiplatelet action

Background: Inhibition of protein disulfide isomerases (PDIA) is becoming an attractive approach for the development of new antiplatelet drugs. Selective PDIA1 blockade by bepristat 2a was previously shown to block GPVI receptor-mediated platelet activation by inhibition of intraplatelet reactive oxygen species via tromboxane A2 (TXA2)-dependent pathway.

Aims: We investigated a newly in-house synthesized inhibitor, C-3399, blocking PDI isoforms PDIA1, PDIA3 and PDIA6.

Methods: To show antiplatelet effects of C-3399 compared to PDIA1 selective inhibitor, bepristat 2a, and reference PDIA1/3/6 inhibitor, PACMA-31, platelet aggregation and platelet surface activation markers in response to the GPVI receptor agonist convulxin versus TXA2-mimetic, U46619, were evaluated using light-transmission aggregometry and flow cytometry, respectively. TXA2 production in response to collagen/convulxin was measured by ELISA.

Results: C-3399 significantly inhibited in a concentration-dependent manner the activation of platelet αIIbβ3 integrin induced by convulxin, while it partially affected CD62P or CD63 surface expression in diluted platelet-rich plasma (PRP). The effects of PACMA-31 were similar to those observed for C-3399, whereas bepristat displayed the strongest inhibitory effect on αIIbβ3 integrin activation and nearly completely abolished surface expression of CD62P or CD63 compared to C-3399 and PACMA-31.
C-3399 and bepristat partially reduced platelet aggregation in PRP induced by high convulxin concentration, while PACMA-31 had no effect.
Interestingly, the blockade of PDIA1, PDIA3 and PDIA6 by C-3399 or PACMA-31 nearly completely inhibited U46619-induced activation of αIIbβ3 integrin, CD63 surface expression and platelet aggregation, while the selective blockade of PDIA1 by bepristat resulted in significantly weaker inhibitory effects on these platelet responses.

Conclusion(s): These results suggests that C-3399 shows differential inhibitory effects on GPVI-mediated platelet activation compared to bepristat and PACMA-1. In contrast to bepristat, C-3399 blocks not only collagen/convulxin-induced TxA2 synthesis but also TXA2-mediated activation of human platelets. However, further studies are required to explore mechanisms behind.