Characterization of a Novel 12(S)-HETE Receptor and its Role in Prostate Cancer Progression; Annual rept. 30 Dec 2006-29 Dec 2007

摘要

P06/0412(S)-hydrox yeicosatetraenoic 12(S)-HETE, a lipoxygenase metabolite of arachidonic acid, has been demonstrated to enhance metastatic capacity during tumor progression via its receptor. Using cloning and binding strategy, we have identified a G-protein coupled receptor, GPR31, as a 12(S)- HETE receptor. In this project, we aimed to characterize the biochemical properties of GPR31 and study the role of GPR31 in tumor survival and metastasis. We have constructed a secondary structure model for GPR31 and using the structural characteristics, we designed two peptides to produce antibodies that against both sides of this membrane protein. We have analyzed the expression of GPR31 in cell lines and human prostate specimens. The GPR31 protein was expressed in various cancer cells and stained in prostate cancer tissue. In order to further characterize this receptor, we designed a series experiments to determine its biochemical properties. Binding experiments with or without GPR31 expressed in Chinese hamster ovary cells demonstrated that this receptor has a Kd value of 4.8 for (3H)12(S)-HETE specific binding with Bmax value of 38.3 pmol/mg protein. We also purified the two antibodies from immunized rabbit antiserums. This project will provide significant insights into the process of prostate cancer progression and metastasis.