Ca2+-dependent caspase activation by gallic acid derivatives

Gallic acid (GA) derivatives, 3,1-methylenedioxyphenyl 3,4,5-trihydroxybenz oate (GD-1) and S-(3,4-methyl-enedioxyphenyl)3,4,5-trihydroxythiobenzoate ( GD-3), were previously reported to induce apoptosis in tu mor cells with IC (50)s of 14.5 muM and 3.9 muM, respectively. To elucidate the mechanism by which these gallic acid derivatives (GDs) induce apoptosis, we studied whet her GD-I and GD-3 can activate caspases, When promyelocytic leukemia HL-60R G cells were treated with GD-1 and GD-3, poly(ADP-ribose)polymerase (PARP), a substrate of caspase-3, was cleaved into 85kDa of degradative product wi th increasing incubation time. GA also activated PARP cleavage, which was i nhibited by catalase, N-acetyl-L-cysteine (NAC). and intracellular Ca2+ che lator 1,2-bis(2-aminophenoxyethane)-N,N,N,N'- tetraacetic acid tetrakis (ac etoxymethyl ester) (BAPTA-AM), in addition to a caspase inhibitor, Z-VAD-FM K. Its inhibitory pattern was identical with that of hypoxanthine/xanthine oxidase, On the other hand, GD-1- and GD-3-induced PARP cleavage was not su ppressed by catalase or NAG, but by BAPTA-AM. This suggested that the GD-el icited signaling pathway is different from GA's, Taken together, GDs activa ted caspase-3 following intracellular Ca2+ elevation independent of reactiv e oxygen species. Thus, it became evident that the signaling pathway leadin g to apoptosis was regulated by GDs in a different manner from GA.