Pomegranate seed is a high-protein food by-product with demonstrated in vitro antioxidant activity. In this work, we evaluated the effect of pomegranate seed protein isolates (PIs) and hydrolysates, obtained with high-intensity focused ultrasounds (HIFU) and pressurized liquids (PLE), in several cancer-related processes using prostate cancer cells (PC-3) and immunodeficient mice. Specifically, cell viability, migration, clonogenicity, adhesion, cell cycle, metalloproteinase (MMP) activity, vascular endothelial growth factor (VEGF) expression, and physical tumors parameters were assessed. The extracts reduced tumor cell viability, migration, and proliferation. IC values for cell viability ranged from 3.4 ± 0.1 mg/mL (PLE PI) to 8.0 ± 0.7 mg/mL (HIFU PI). Tumor cell migration in the presence of PIs or hydrolysates was reduced by 35-51%, compared to the control (no extract or hydrolysate addition). Additionally, hydrolyzed HIFU PI increased cell adhesion by 43.2%. HIFU-treated samples induced cell cycle arrest in the S-phase, whereas PLE-treated samples led to apoptosis. In vivo experiments supported these findings, showing that hydrolysates significantly reduced tumor volume (by 42% for HIFU and 57% for PLE) and tumor weight (by 27% for HIFU and 35% for PLE). These changes were accompanied by significant decreases in VEGF levels and MMP activity. The antitumor effects of the extracts and hydrolysates may be attributed not only to intact proteins and peptides but also to co-extracted phenolic compounds, more abundant in the PLE-derived extract.