Significant advances in gene therapy have been made as a result of the improvement of gene delivery systems, discovery of new therapeutic genes, better understanding of mechanisms of disease progression, exploration and improvement of tissue-specific gene regulatory sequences, and development of better prodrug/enzyme systems. This review discusses adenoviral-based and prostate-specific cancer gene therapy--emphasizing tissue-specific promoter choices to increase gene therapy safety and specificity--and the development of prostate-targeted vectors, with a focus on the two-step transactivation system for amplifying gene expression, specifically in prostate cancer cells. Several examples will be discussed for the scientific basis and therapeutic applications. In addition, prostate cancer gene therapy clinical trials and future directions in this field will also be described briefly.