202. Using RNA To Inhibit Intimal Hperplasia by Blocking E2F Function

Abstract

Top of pageAbstract The six members of the E2F family of transcription factors are key players in the Rb tumor suppressor pathway leading to DNA replication and cell cycle control. The family members have both distinct and overlapping functions, and gaining an understanding of the roles each factor plays in the cell cycle will lead to a more precise understanding of the Rb pathway. This, in turn, will lead to ways to control the pathway in proliferative disorders such as restenosis after angioplasty and cancer. Using a large combinatorial library of RNA molecules, called aptamers, and an interative in vitro selection technique, the few aptamers that can bind to and inhibit a protein's function can be isolated. This technique was previously employed to isolate RNA aptamers that bind to E2F1 with Kd's of 2nM. These aptamers were able to block cell cycle progression when microinjected into cell nuclei. The same technique has been used here to isolate modified, nuclease resistant RNA aptamers that bind to E2F3 with Kd's of < 1 nM, and that are able to block DNA binding to E2F. One family of aptamers has been shown to bind preferentially to E2F3, while another family has been shown to bind E2F1-5 equally, yielding two very different tools with which to study E2F function. Another method of using RNA to inhibit E2F function is to use siRNA to individual E2F members. Both the aptamers and the siRNA have been shown to inhibit function of overexpressed E2Fs in NIH/3T3 cells, and are currently being tested for their ability to inhibit intimal hyperplasia in mouse vein graft models using a variety of delivery methods including naked nucleic acid, a pleuronic gel, and viral vectors.