| Co-Investigator(Kenkyū-buntansha) |
MORISHIGE Kenichiro Osaka University Graduate School of Medicine, Assistant professor, 医学系研究科, 助手 (90283788)
SAKATA Msahiro Osaka University Graduate School of Medicine, Lecturer, 医学系研究科, 講師 (10260639)
KURACHI Hirohisa Osaka University Graduate School of Medicine, Associate professor, 医学系研究科, 助教授 (40153366)
YAMAMOTO Toshiya Osaka University Graduate School of Medicine, Assistant professor, 医学系研究科, 助手 (80283787)
池上 博雅 大阪大学, 医学部, 講師 (10184409)
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| Budget Amount *help |
¥11,600,000 (Direct Cost: ¥11,600,000)
Fiscal Year 2000: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1999: ¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 1998: ¥4,200,000 (Direct Cost: ¥4,200,000)
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| Research Abstract |
Although gonadotropin-releasing hormone agonists (GnRHa) have been used in the therapy of the endocrine-dependent cancers, their biological mechanism remained obscure. We have studied the roles of mitogen-activated protein kinase (MAPK) family in the antiproliferative effect of GnRHa on the cisplatin resistant Caov-3 human ovarian cancer cell line. Reverse-transcriptase PCR assays confirmed mRNA for GnRH receptor in Caov-3 cells. In the presence of 1 μM GnRHa, the proliferation of cells was significantly reduced to 76% of controls after 24 h and the effect was sustained up to 4 days. Although GnRHa had no effect on the activation of the Jun N-terminal kinase (JNK), treatment of Caov-3 cells with GnRHa activated extracellular signal-regulated protein kinase (ERK) and its effect was more than that induced by GnRH.Activation of ERK by GnRHa occurred within 5 min, with the maximum at 3 h and sustained until 24 h. To examine the role of ERK cascade in the antiproliferative effect of GnRHa,
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PD98059, an inhibitor of MEK, was used. This inhibitor canceled the antiproliferative effect of GnRHa and apparently reversed the GnRH-induced dephosphorylation of the retinoblastoma protein (pRb), whose hyperphosphorylation is a hallmark of G1-S transition in the cell cycle. These results provide evidence that GnRHa stimulation of ERK activity may play an important role in the antiproliferative effect of GnRHa in the Caov-3 human ovarian cancer cell line. Next, we have studied the roles of JNK and ERK cascade in both the cisplatin resistant Caov-3 and sensitive A2780 human ovarian cancer cell lines. Treatment of both Caov-3 and A2780 cells with cisplatin but not the transplatin isomer activates JNK and ERK.Activation of JNK by cisplatin occurred at 30 min, reached a plateau at 3 h, and declined thereafter, whereas activation of ERK by cisplatin showed a biphasic pattern (peaks at 30 min and 3 h), indicating the different time frame. Activation of JNK by cisplatin was maximal at 1000 μM, whereas activation of ERK was maximal at 100 μM and was less at higher concentrations, indicating the different dose-dependency. Exogenous expression of dominant negative c-Jun (dnJun) in both Caov-3 cells, which are highly resistant to cisplatin (IC50=380±25 μM), and A2780 cells, which are sensitive to cisplatin (IC50=84±4 μM), decreased viability following treatment with cisplatin : the IC50 for cisplatin was 7.6-and 4.2-fold less in dnJun expressing Caov-3 and A2780 cells, respectively. We further examined the role of ERK cascade on the viability following cisplatin treatment using PD98059. The treatment by this compound induced sensitivity to cisplatin, but not to transplatin, leading to a 9.7-and 1.8-fold less IC50 for cisplatin in Caov-3 and A2780 cells, respectively. Our findings suggest that cisplatin-induced DNA damage differentially activates JNK and ERK cascades and inhibition of either of these cascades sensitizes ovarian cancer cells to cisplatin. Less
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