Continuous exposure to low-dose cisplatin and apoptosis.
A high concentration of cisplatin induces apoptosis in many tumor cell lines. Whether cisplatin induces apoptosis even in a controlled release formulation has not been determined. We therefore studied the relationship between the dosing regimen of cisplatin and the induction of apoptosis in rat hepatoma AH-109A cells. A colorimetric assay was used to quantify cell proliferation and viability, and caspase activity was determined using an exogenous fluorogenic peptide substrate. When delivered as a single dose, cisplatin caused a dose-dependent inhibition of AH-109A growth and enhancement of caspase-3 activity. Also, DNA laddering was detected in cells that had elevated caspase-3 activity. However, caspase-3 activity was low and DNA laddering and a sub-G1 population were not detected when cells were treated with a combination of cisplatin and the caspase inhibitor Z-VAD-FMK. These results suggest that cisplatin is cytotoxic in AH-109A cells because it induces apoptosis. We next examined intermittent exposure to cisplatin to estimate the effects of continuous exposure by a controlled release formulation. Cisplatin was divided into equal parts and was added intermittently into the medium resulting in the same final concentration as the single dose. The individual additions alone were not cytotoxic, but all of the doses together had a similar cytotoxic effect as a single exposure of cisplatin. The intermittent exposure resulted caspase-3 activity even higher than a single dose. These findings indicate that cisplatin induces apoptosis in AH-109A cells when delivered continuously even at the concentration that alone have no activity.
Kishimoto
Shuichi
S
Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Kobe Gakuin University; Kobe 651-2180, Japan. skisimot@pharm.kobegakuin.ac.jp
Kawazoe
Yuji
Y
Ikeno
Mako
M
Fukushima
Shoji
S
Takeuchi
Yoshikazu
Y
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000008:administration & dosage / Q000494:pharmacology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D016193:G1 Phase; D008114:Liver Neoplasms, Experimental / Q000201:enzymology / Q000473:pathology; D051381:Rats
D000970:Antineoplastic Agents; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D002945:Cisplatin