Caspase-3 activation during apoptosis caused by glutathione-doxorubicin conjugate.
Glutathione-doxorubicin (GSH-DXR) effectively induced apoptosis in rat hepatoma cells (AH66) at a lower concentration than DXR. After 24 h of drug treatment, DNA fragmentation of the cells was observed at the concentration of 1.0 microM DXR or 0.01 microM GSH-DXR. Increase in caspase-3 activity and DNA fragmentation were observed within 12 h and 15 h after treatment with either drug. Intracellular caspase-3 activity was increased in a dose-dependent manner after treatment with DXR or GSH-DXR, and caspase-3 activity correlated well with the ability to induce DNA fragmentation. When the cells were treated with either DXR or GSH-DXR for only 6 h, apoptotic DNA degradation and caspase-3 activation occurred 24 h after treatment. DNA fragmentation caused by these drugs was prevented completely by simultaneous treatment with the caspase-3 inhibitor, acetyl-Asp-Glu-Val-Asp-aldehyde (DEVD-CHO), at 10 microM. By contrast, DNA fragmentation was not prevented by the caspase-1 inhibitor, acetyl-Tyr-Val-Ala-Asp-aldehyde (YVAD-CHO), at the same concentration as DEVD-CHO, and caspase-1 was not activated at all by the treatment of AH66 cells with both DXR and GSH-DXR. These results demonstrate that DXR and GSH-DXR induce apoptotic DNA fragmentation via caspase-3 activation, but not via caspase-1 activation, and that GSH-DXR enhances the activation of caspase-3 approximately 100-fold more than DXR. Moreover, the findings suggested that an upstream apoptotic signal that can activate caspase-3 is induced within 6 h by treating AH66 cells with the drug.
Asakura
T
T
Department of Biochemistry (I), Jikei University School of Medicine, Tokyo, Japan.
Sawai
T
T
Hashidume
Y
Y
Ohkawa
Y
Y
Yokoyama
S
S
Ohkawa
K
K
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002417:Cattle; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects / Q000378:metabolism; D004317:Doxorubicin / Q000737:chemistry / Q000494:pharmacology*; D004337:Drug Carriers; D004789:Enzyme Activation / Q000187:drug effects; D005978:Glutathione / Q000737:chemistry / Q000494:pharmacology*; D008114:Liver Neoplasms, Experimental / Q000188:drug therapy* / Q000201:enzymology* / Q000473:pathology; D046911:Macromolecular Substances; D009842:Oligopeptides / Q000494:pharmacology; D051381:Rats; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D004273:DNA, Neoplasm; D004337:Drug Carriers; D046911:Macromolecular Substances; D009842:Oligopeptides; C108637:aspartyl-glutamyl-valyl-aspartal; C096429:L 709049; D004317:Doxorubicin; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D005978:Glutathione