Peptide-Mediated Disruption of Calmodulin–Cyclin E Interactions Inhibits Proliferation of Vascular Smooth Muscle Cells and Neointima Formation
Rationale: Cell cycle progression in vascular smooth muscle cells (VSMCs) is a therapeutic target for restenosis.
Objective: Having discovered that calmodulin (CaM)-dependent cyclin E/CDK2 activity underlies Ca2+-sensitive G1-to-S phase transitions in VSMCs, we sought to explore the physiological importance of the CaM–cyclin E interaction.
Methods and Results: A peptide based on the CaM binding sequence (CBS) of cyclin E was designed to interfere with CaM–cyclin E binding. Compared with control peptides, CBS blocked activating Thr160 phosphorylation of CDK2, decreased basal cyclin E/CDK2 activity, and eliminated Ca2+-sensitive cyclin E/CDK2 activity in nuclear extracts from mouse VSMCs. Nucleofection with CBS, or treatment with CBS conjugated to the HIV-1 TAT protein transduction domain to improve bioavailability, inhibited G1-to-S cell cycle progression in a dose-dependent manner. These effects were not observed with control peptides. TAT-CBS inhibited 3H-thymidine incorporation in primary human aortic SMCs (HA-SMCs) in vitro, manifested greater transduction into HA-SMCs compared with endothelial cells in vitro, and limited decreased SM22α expression, neointima formation, and medial thickening without affecting collagen deposition or reendothelialization in a mouse model of carotid artery injury in vivo. The antiproliferative effects of CBS remained evident in mouse embryonic fibroblasts derived from wild-type mice but not cyclin E1/E2 double knockout mice.
Conclusions: A synthetic peptide designed to disrupt CaM–cyclin E binding inhibits Ca2+/CaM-dependent CDK2 activity and cell cycle progression and proliferation in VSMCs and limits arterial remodeling following injury. Importantly, this effect appears to be cyclin E–dependent and may form the basis of a potentially novel therapeutic approach for restenosis.
- Received December 20, 2010.
- Revision received February 17, 2011.
- Accepted February 23, 2011.
- © 2011 American Heart Association, Inc.