Natural killer (NK) cells display built-in activity against tumor cells, which can be enhanced and rendered antigen-specific through expression of chimeric antigen receptors (CARs). Recently, CAR-expressing NK cells have  been reported to confer promising safety and efficacy in patients with CD19-positive lymphoid neoplasms. However, CAR NK cell therapy has shown no clear benefit thus far against solid tumors. Aim of this project is the generation and preclinical evaluation of CAR NK cells for prostate cancer immunotherapy that recognize prostate-specific membrane antigen (PSMA). So far, we have developed robust and efficient protocols for the expansion and transduction of NK cells, as well as the functional evaluation of the resulting CAR NK cells. Primary NK cells were derived from healthy donors and expanded using feeder cells. Transduction with a gamma-retroviral vector encoding a second-generation PSMA-specific CAR, IL-15 and the dLNGFR marker was embedded into the expansion process. Upon transduction, some 60% of NK cells expressed the transgenes over a period of several weeks, during which the CAR NK cells maintained proliferative capacity. In order to assess anti-tumor activity in vitro, CAR-expressing NK cells were co-incubated with PSMA-positive or PSMA-negative tumor cells, and target cell killing was tracked over time using live cell imaging. Our results demonstrate that the CAR NK cells readily targeted tumor cells in a CAR-dependent and PSMA-specific manner, which was further enhanced by IL-15. Based on these promising results, in vivo evaluation of the PSMA-specific CAR NK cells in mouse tumor models is planned as the next step.