Etoposide inhibits DNA synthesis by forming a complex with topoisomerase II and DNA (IC50: 60.3 μM).
Etoposide inhibits proliferation of a variety of adenocarcinoma cells (IC50s: 0.005-12,200 μM) and HUVEC cells (IC50: 0.249 μM) . Etoposide is capable of causing cytotoxicity on pancreatic β-cells by inducing apoptosis through the JNK/ERK-mediated GSK-3 downstream-triggered mitochondria-dependent signaling pathway in RIN-m5F cells .
Etoposide (25 mg/kg) reduces tumor growth in a Ma human embryonal carcinoma mouse xenograft model . Etoposide (10 mg/kg/day, i.v.) with ifosfamide and carboplatin, reduces the tumor volume in the hepatoblastoma cell injected NMRI nude mice .
Nuclear extracts are prepared, and nuclei are isolated. The activity of topoisomerase II is calculated from the percentage of decatenation obtained. Tritiated kinoplast DNA (KDNA 0.22 μg) is used as a substrate. Etoposide and topoisomerase II are incubated for 30 min at 37 ℃ and are stopped with 1% sodium dodecyl sulfate (SDS) and proteinase K (100 μg/mL). The percentages of decatenation and inhibition of topoisomerase II by Etoposide are obtained .
After the Etoposide treatment, cells are removed from the dish with phosphate-buffered saline (PBS) containing 0.03% trypsin and 0.27 mM ethylenediaminetetraacetic acid (EDTA) and are diluted into culture dishes in appropriate numbers to yield between 20 and 200 colonies. After 12 days, cultures are fixed with methanol-acetic acid, stained with crystal violet, and scored for colonies containing more than 50 cells .
The in vivo model for nude mice HB (NMHB) has been established. Only HB cells with embryonal components are grafted and reproduced successfully in this model. Each NMHB subsequently is transplanted into 50 mice for treatment groups. Treatment is initiated when the majority of the tumors reach a volume of 50-100 mm3. The mice are stratified according to their tumor volume and randomly assigned to groups of ten animals each. The animals injected with tumor are given ifosfamide, cisplatin, doxorubicin, etoposide (10 mg/kg/day, i.v.), and carboplatin as single agents in two blocks. One group of ten animals for each original xenograft served as a control group. After initiation of treatment, the tumor growth is recorded at 5-day intervals for 25-30 days and the relative tumor volumes are calculated. Twenty-four hours before the animals are sacrificed, bromodeoxyuridine (BrdU) is injected intraperitoneally for the semiquantitative determination of proliferation activity of the tumor cells (50 μg of BrdU/g body weight) .