A complex of human
alpha-lactalbumin and
oleic acid (HAMLET) was originally isolated from human milk as a potent
anticancer agent. It kills a wide range of transformed cells of various origins while leaving nontransformed healthy cells largely unaffected both in vitro and in vivo. Importantly, purified alpha-
lactalbumins from other mammals form complexes with
oleic acid that show biological activities similar to that of HAMLET. The mechanism by which these
protein-
lipid complexes kill
tumor cells is, however, largely unknown. Here, we show that complex of bovine
alpha-lactalbumin and
oleic acid (BAMLET), the bovine counterpart of HAMLET, kills
tumor cells via a mechanism involving lysosomal membrane permeabilization. BAMLET shows potent cytotoxic activity against eight
cancer cell lines tested, whereas nontransformed NIH-3T3 murine embryonic fibroblasts are relatively resistant. BAMLET accumulates rapidly and specifically in the endolysosomal compartment of
tumor cells and induces an early leakage of lysosomal
cathepsins into the cytosol followed by the activation of the proapoptotic
protein Bax. Ectopic expression of three
proteins known to stabilize the lysosomal compartment, i.e.
heat shock protein 70 (Hsp70), Hsp70-2, and
lens epithelium-derived growth factor, confer significant protection against BAMLET-induced cell death, whereas the antiapoptotic
protein Bcl-2,
caspase inhibition, and autophagy inhibition fail to do so. These data indicate that BAMLET triggers lysosomal cell death pathway in
cancer cells, thereby clarifying the ability of
alpha-lactalbumin:
oleate complexes to kill highly apoptosis-resistant
tumor cells.