Natural killer (NK) cell tumor infiltration is associated with good prognosis in patients with metastatic castration-resistant prostate cancer (mCRPC). NK cells recognize and kill targets by a process called natural cytotoxicity. We hypothesized that promoting an antigen-specific synapse with co-activation may enhance NK cell function in mCRPC. We describe a Tri-specific Killer Engager (TriKE) construct that engages with the activating receptor CD16 on NK cells, prostate-specific membrane antigen (PSMA) on mCRPC cells, and has an interleukin (IL)-15 moiety that is essential for NK cell survival, proliferation, and priming. We show that the PSMA TriKE specifically binds to PSMA-expressing cells and significantly enhances expansion, degranulation and cytokine production of NK cells derived from healthy donors or prostate cancer patients. Bystander killing of PSMA-negative was also achieved with PSMA TriKE treatment when co-cultured with PSMA-positive cells, suggesting potential PSMA TriKE benefit in controlling tumor antigen escape. When tested under physiologic conditions recapitulating the mCRPC tumor microenvironment (TME), NK cells treated with PSMA TriKE and prolonged exposure to hypoxia or MDSCs maintained their potent function while IL-15 treated NK cells showed greatly impaired cytotoxicity. Finally, in vivo testing of PSMA TriKE showed improved tumor control and survival of mice as compared to IL-15 and untreated control groups. In conclusion, PSMA TriKE demonstrates potential as a new therapy for advanced prostate cancer by providing additional signals to NK cells to maximize their anti-tumor potential in prostate cancer, especially in the setting of a hostile TME.