The development of immunotherapy for cancer, such as synthetic peptide-based vaccines, relies heavily on the identification of appropriate epitopes capable of eliciting antitumor T-cell responses. We have used a combination of computer-based algorithms to predict peptide sequences from prostate-specific membrane antigen (PSMA) capable of stimulating in vitro CTLs restricted by the HLA-A2 MHC molecule. Four of the five peptides that were predicted by these algorithms were capable of inducing antigen-specific CTLs that killed target cells that were pulsed exogenously with the corresponding peptides. However, only one of the four peptides, PSMA(27), induced CTLs that were effective at recognizing prostate tumor cells expressing the HLA-A2 and PSMA molecules. These results underline the importance of demonstrating antitumor reactivity of peptide-induced CTLs for the selection of epitopes destined to become immunotherapeutic for prostate cancer.