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The heated products of peptic hydrolysates of half-fin anchovy (Setipinna taty) (HAHp-H) were fractioned using Bio-Gel P-4 and Sephadex G 25 chromatography, and RP-HPLC to obtain peptides with antiproliferative activities on human prostate cancer PC-3 cells. One peptide was identified with the sequence of YALPAH. The synthetic YALPAH induced PC-3 cell apoptosis at the concentration of 4.47 μM. Three analogous peptides, i.e., YALRAH, YALPAR and YALPAG, were also synthesized. All these synthetic peptides displayed concentration-dependent manners in inhibiting the growth of PC-3 cells. Furthermore, peptide YALRAH with the highest positive charge exhibited the strongest antiproliferation, with IC50 value of 8.1 mg/ml (or 11.1 μM). While as with the same net charges, the lower molecular weight of YALPAH (670.77 Da) was proposed to be responsible for its higher cytotoxicity compared with YALPAR (689.81 Da). This is the first report of an antiproliferative peptide on PC-3 cells derived from half-fin anchovy.