[Synergetic cytotoxicity of cisplatin and etoposide to leukemia cell line K562 and its mechanism].
OBJECTIVE
Etoposide (VP-16) is one of the most common chemotherapy drugs, but its usage is limited by drug resistance. Some researches on solid tumors show that cisplatin (DDP) have synergetic effect with VP-16. This study was to evaluate synergetic cytotoxicity of VP-16 and DDP to leukemia cell line K562, and explore the mechanism.


METHODS
K562 cells were treated with VP-16 (0 or 5 microg/ml) and DDP (0, 0.3, 3, 15, or 30 microg/ml) in different combination patterns. Inhibitory effect of VP-16 and DDP on survival of K562 cells was measured by MTT assay. Cell apoptosis was evaluated by AO/EB double fluorescent labeling. The expression of topoisomerase (TOPO) II alpha and II beta, and transcription factor Sp1 and Sp3 were measured by semi-quantitive reverse transcription-polymerase chain reaction (RT-PCR) and Western blot.


RESULTS
MTT assay showed significant synergetic cytotoxicity of VP-16 and DDP. VP-16 in combination with DDP obviously enhanced cell apoptosis. RT-PCR showed that DDP significantly up-regulated the expression of TOPO II and Sp1 in concentration-dependent manners (TOPO II alpha, II beta, and Sp1 were up-regulated by 36%, 25%, and 75% of control, respectively, when treated with 30 microg/ml of DDP), and down-regulated Sp3 by 49% of control; VP-16 (5 microg/ml) down-regulated TOPO II alpha by 71.9%, and up-regulated Sp3 by 14%; VP-16 (5 microg/ml) in combination with DDP (30 microg/ml) up-regulated TOPO II alpha by 83%, II beta by 11%, and Sp1 by 58% when compared with using VP-16 alone (but the levels were lower than using DDP alone), and down-regulated Sp3 by 61.3% when compared with using DDP alone. Western blot showed similar results to RT-PCR.


CONCLUSIONS
Through up-regulating TOPO II, DDP could enhance the chemotherapeutic effect of VP-16 on K562 cells by provide more target enzyme to act on.
Ma
Wei-Dong
WD
Cell Biology Division, Institute of Basic Medicine, Hebei Medical University, Shijiazhuang, Hebei, P. R. China.
Yin
Mei-Yun
MY
Jiang
Chang-Wen
CW
Xu
Shi-Rong
SR
Zhai
Li-Dong
LD
Zheng
Li-Fen
LF
Wang
Yan-Ling
YL
Yan
Yun-Li
YL
D000951:Antigens, Neoplasm / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D002945:Cisplatin / Q000008:administration & dosage / Q000494:pharmacology*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D005047:Etoposide / Q000494:pharmacology*; D006801:Humans; D020014:K562 Cells; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051705:Sp3 Transcription Factor / Q000378:metabolism
D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D004268:DNA-Binding Proteins; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D005047:Etoposide; D004250:DNA Topoisomerases, Type II; D002945:Cisplatin