PTEN dephosphorylates the 3-position phosphate of phosphatidylinositol 3,4,5 triphosphate (PIP3), thereby inhibiting Akt activation.The PTEN gene is frequently mutated in a variety of human cancers and compromising PTEN function promotes tumorigenesis.Consistent with this concept, PTEN is a dominant tumor suppressor preventing prostate cancer formation.Loss of PTEN is sufficient to initiate and promote prostate tumorigenesis.However, mechanisms governing PTEN inactivation during prostate tumorigenesis remain unclear.To search for PTEN negative regulators (P10NRGLs), we developed a novel LNCaP cell line-based screen system.Ectopic PTEN potently inhibited LNCaP cell proliferation, which was prevented by co-transduction of a human prostate eDNA library.By using this system, we identified three candidates of P10NRGLs, P 10NRGL 1, 2 and 3.P10NRGL 3 sustained LNCaP cell proliferation when PTEN was overexpressed, and activated Akt in PTEN-positive DU145 cells.