The PTEN gene, located on chromosome 10, is a phosphatase in the phosphatidylinositol 3'-kinase (PI3'K)-mediated signal transduction pathway. PTEN inhibits the activation of Akt, a serine-threonine kinase involved in proliferative metabolic and anti-apoptotic pathways, and has tumor suppressor properties. We used a PTEN adenoviral vector, Ad-MMAC, to assess the role of PTEN in the treatment of prostate cancer. Infection of Ad-MMAC in PC-3 and LNCaP prostate cancer cells (PTEN deleted, up-regulation of phosphorylated Akt) resulted in PTEN expression and significantly decreased growth compared with Ad-CTR or mock infected cells. Infection of Ad-MMAC did not inhibit the growth of DU-145 cells (wild-type PTEN). Combination therapy with Ad-MMAC and doxorubicin improved the efficacy of PTEN gene therapy in PC-3 and DU-145 cells. These data demonstrate that PTEN gene therapy can effectively treat some prostate cancers that have genomic alterations in PTEN. In others, PTEN gene therapy combined with chemotherapy is more effective.