| Co-Investigator(Kenkyū-buntansha) |
KANAMORI Yasunobu Tottori Univ., University Hospital, Research Associate, 医学部附属病院, 講師 (70283984)
ITAMOCHI Hiroaki Tottori Univ., University Hospital, Research Aaaociates, 医学部附属病院, 助手 (20314601)
TERAKAWA Naoki Tottori Univ., Faculty of Medicine, Professor, 医学部, 教授 (90163906)
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| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2006: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2005: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2004: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Research Abstract |
Gene therapy has been tried however, its success is limited in patients who develop chemoresistance a distinctly different clinical behavior. Many mechanisms that may be involved in drug resistance have been proposed, including a decrease in the accumulation of the drug, an increase In detoxification of the drug within the cell, and an increase in DNA repair activity. Additionally, recent studies suggest that apoptosis also relates to chemoresistance.
Mutation of p53 have frequently been observed in ovarian cancer. p53, which is known to be a cell cycle checkpoint protein, plays a regulatory role in the control of cell proliferation and apoptosis. First, we conducted the present study to investigate whether and how chemosensitivity can be determined by means of genetic diagnosis using drug-resistance genes in patients with epithelial ovarian cancer. A total of 75 patients who had epithelial ovarian cancer with measurable lesions were entered into this study. Thirty-three patients receiv
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ed first-line chemotherapy, consisting of paclitaxel and carboplatin (TJ). Forty-two patients received second-line chemotherapy : 22 received EP therapy consisting etoposide and cisplatin (CDDP), and 20 received irinotechan (CPT-11) and CDDP (CPT-11/CDDP) therapy. Tumor samples were obtained before chemotherapy. mRNA expressions of the multidrug resistance gene-1 (MDR-1), multi-drug resistance-associated protein-1 (MRP-1), topoisomerase (topo)-I, and topoisomerase (topo)-IIα were measured by real-time RT-PCR. The cut-off values of each gene were determined by the receiver operating characteristic curve. MDR-1 expression was significantly higher in patients who did not respond to TJ therapy. Tile expression of Topo-IIα was significantly higher in patients who did respond to EP therapy. The expression of Topo-I was significantly higher in patients who did respond to CPT-11/CDDP. MRP-1 expression did not differ between responders and non-responders in all regimens. The cut-off value was 80 for MDR-1, 90 for topo-II, and 200 for topo-I. Next, to evaluate genetic diagnosis, 31 patients were newly added. The accuracy of this genetic diagnosis for chemosensitivity was 85.7 % for TJ, 77.8% for EP, and 100.0% for CPT-11/CDDP therapy. The present study suggests that genetic diagnosis may be useful to determine chemosensitivity in patients with epithelial ovarian cancer.
Next, resistance of ovarian clear cell carcinoma (CCC) to platinum-based chemotherapy is associated with poor prognosis and an effective treatment for advanced disease is urgently needed. HER2/neu is upregulated more often in CCC than in other histologic types of epithelial ovarian cancer. The purpose of this study was to assess possible treatment for ovarian CCC with the anti-HER2 antibody trastuzumab or human adenovirus type 5 E1A. We treated 10 CCC cell lines with trastuzumab or E1A and assessed cell viability, proliferation, and colony formation and the expression of HER2 and wild-type p53 proteins and molecules downstream of those signaling pathways. HER2 protein was detected at various levels in all 10 cell lines by western blotting and in five CCC cell lines by immunohistochemical staining ; HER2 gene amplification was detected (by fluorescence in situ hybridization) in only one cell line (RMG-I). Trastuzumab did not inhibit proliferation in any of the four CCC cell lines tested (RMG-I, SKOV-2, OVTOKO, and OVSAYO). However, transfection with E1A (as compared with control vectors) reduced colony formation in all 10 CCC cell lines regardless of HER2 expression level. Infection of RMG-I and SMOV-2 cells with an adenoviral vector encoding E1A led to significant (P<0.05) suppression of proliferation and enhancement of cell death ; this effect required stabilization of p53 (but not p73) protein and was associated with the upregulation of Bax and the cleavage of caspase 9. Other mechanisms, such as p53-independent apoptosis, may be also involved in EJA-mediated cell death in CCC. Finally, treatment with E1A prolonged survival in a CCC xenograft model (P<0.001). E1A gene therapy, because of its ability to stabilize wild-type p53, is worth exploring as a treatment modality for women with ovarian CCC, which typically expresses wild-type p53.
These studies suggest that biological behivior contributes drug-induced apoptosis and future gene threapy should be based on biological behivior in ovarian cancer. Less
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