Rice bran, an economical, underutilized coproduct of rough rice milling, was used to produce peptide hydrolysates, which were investigated for anticancer activity. Protein hydrolysates prepared by Alcalase hydrolysis under optimized conditions were treated further to obtain gastrointestinal (GI)-resistant peptide hydrolysates. They were fractionated into >50, 10-50, 5-10, and <5 kDa sizes and evaluated for inhibitory activity on proliferation of human colon (Caco-2) and liver (HepG2) cancer cell lines by Trypan blue dye exclusion assay. GI-resistant <5 and 5-10 kDa sized peptide fractions inhibited growth of Caco-2 cells by 80%, and the <5 kDa fraction inhibited growth of HepG2 cells by approximately 50% compared to controls and nonresistant fractions. An MTS cell titer assay confirmed antiproliferative effects of the peptide fractions. The results demonstrated that 5-10 and <5 kDa sized GI-resistant fractions promoted significant (p < 0.05) inhibitory activities on both cancer cell lines compared to controls. More investigations are needed to show such value-added effects on the technofunctional and sensorial properties of the food protein and peptide matrices.