PI3K/p110alpha inhibition selectively interferes with arterial thrombosis and neointima formation, but not re-endothelialization: a new concept in drug-eluting stent design

Abstract

Background: Impaired re-endothelialization and stent thrombosis are a safety concern associated with drug-eluting stents (DES) as mTOR inhibitors reduce restenosis rates, but impair endothelialization and facilitate thrombus formation. PI3K/p110α controls cellular wound healing pathways, thereby representing an emerging drug target to modulate vascular homeostasis after injury. However, its effect on arterial thrombosis and neointima formation remains unknown. Methods: PI3K/p110α was inhibited by treatment with the small molecule inhibitor PIK75 or a specific siRNA. Arterial thrombosis, neointima formation and re-endothelialization studies were performed in a murine carotid artery injury model. Proliferation and migration of VSMC and EC were assessed by cell number and Boyden chamber, respectively. Endothelial senescence was evaluated by β-galactosidase assay, endothelial dysfunction by organ chambers for isometric tension. Tissue factor (TF), plasminogen activator inhibitor type 1 (PAI-1), and eNOS expression was determined by Western blot and real-time PCR analysis. Results: Male C57Bl6 mice were either treated with PIK75 (10 mg/kg/d for 7 days) or vehicle. Arterial thrombus formation was delayed in mice treated with PIK75 as compared to controls (n=8; p<0.005). PIK 75 impaired arterial expression and activity of TF and PAI-1 as well as NFκB activity (n=8; p<0.05); in contrast, plasma clotting and tail bleeding times did not differ (n=8; p=NS). In human vascular smooth muscle and endothelial cells, PIK75 inhibited expression and activity of TF and PAI-1 (n=4; p<0.01). These effects occurred at the transcriptional level via the RhoA signaling cascade and the transcription factor NFkB. Furthermore, inhibition of PI3K/p110α with PIK75 or a specific siRNA selectively impaired proliferation and migration of VSMC while sparing EC completely. In line with this observation, treatment with PIK75 selectively inhibited neointima formation without affecting re-endothelialization following vascular injury. In contrast to rapamycin and paclitaxel, treatment with PIK75 did not induce endothelial senescence nor inhibit eNOS expression or endothelium-dependent relaxations. Conclusions: Following vascular injury, PI3K/p110α inhibition selectively interferes with arterial thrombosis and neointima formation, but not reendothelialization. Hence, PI3K/p110α represents an attractive new target in DES design.