臺灣博碩士論文加值系統

中文摘要
P-glycoprotein是多重抗藥性(MDR)基因的protein產物，且為一
efflux transporter，會降低藥物在細胞內的蓄積，因此被視為化學治療上的障礙。之前實驗已證實一結構與已知為Pgp抑制劑terfenadine相似的藥物hydroxyzine可增加etoposide在老鼠小腸中的吸收，但由於老鼠小腸中尚有其他transporters的存在，所以本實驗想更進一步探討hydroxyzine在Pgp上所扮演的角色。
本研究首先是建立一具有Pgp表現的HEK293(human embryonic kidney cell line)細胞株，再以不同的Pgp受質(vincristine, epirubicin, etoposide)進行細胞毒性試驗，得知在有Pgp表現的HEK293細胞株中EC50會明顯的增加。同時，藉由已被證實為Pgp受質的rhodamine123這個螢光性物質，來確認Pgp在HEK293細胞株的功能及表現，結果在有Pgp表現的HEK293細胞株中，rhodamine123的蓄積會降低；接著，再以rhodamine123當受質，以不同濃度的hydroxyzine當抑制劑，結果卻顯示在有MDR-transfected的HEK293 cells中 hydroxyzine並不會增加rhodamine123的蓄積。但由於不同的受質作用於Pgp上的位置並不完全相似，所以我們進一步再利用作用於另一個結合位置(H site)的受質colchicine來探討其與hydroxyzine之間的交互作用，結果顯示，hydroxyzine同樣也不會增加colchicine在MDR-transfected HEK293細胞中的蓄積，由以上結果可證實hydroxyzine並不會作用於Pgp來增加藥物在細胞中的蓄積。

Abstract
P-glycoprotein functions as an ATP-dependent conjugate efflux pump to decrease drug accumulation in a variety of systems. Therefore, it was regarded as a major obstacle in chemotherapy. Previous study has demonstrated that hydroxyzine could increase the uptake of etoposide in the rat intestine. Since there are other transporters existing in the rat intestine we further want to investigate the interaction between hydroxyzine and Pgp.
We have established the MDR-transfected HEK293 cell. The permanent expression of human MDR in stable transfected cell lines enabled us to study on cytotoxicity using Pgp substrates (vincristine, epirubicin, etoposide). Increased EC50 was observed in MDR-transfected HEK293 cells. Moreover, the function and expression of Pgp in HEK293 cells was confirmed using Pgp substrate R123. The accumulation of R123 was significantly decreased in MDR-transfected HEK293 cells. Furthermore, R123 was chosen as a substrate and different concentrations of hydroxyzine were regard used as inhibitors to assess the role of Pgp in drug interaction.
The result showed that hydroxyzine did not increase the accumulation of R123 in MDR-transfected HEK293 cells. Besides of R site, there is still the other binding site (H site) on Pgp. For this reason, colchicine, the substrate of H site, was used to investigate the interaction with hydroxyzine on Pgp. Similar to R123, hydroxyzine did not increase the accumulation of colchicine in MDR-transfected HEK293 cells. According to above results, we cannot prove that hydroxyzine is able to increase the accumulation of Pgp substrates.

目錄
誌謝 1
中文摘要 2
英文摘要 3
縮寫表 4
目錄 5
表目錄 7
圖目錄 8
第一章. 緒論 10
第一節. 藥物簡介 10
第二節. 多重抗藥性 11
第三節. 多重抗藥性的種類及Pgp的表現 11
第四節. Pgp的結構、功能及其分佈 12
第五節. Pgp在藥物交互作用的角色 13
第六節. Pgp的受質、作用機轉及其結合位置 14
第七節. Pgp的抑制劑及其調控機轉 15
第二章. 研究目的 26
第三章. 實驗材料及方法 27
第一節. 材料 27
第二節. 儀器 36
第三節. 實驗方法 37
第四章. 實驗結果 49
第一節. Construction of pcDNA3-hMDR1 49
第二節. Pgp在MDR-transfected HEK細胞中的表現 49
第三節. 細胞毒性試驗 50
第四節. Rhodamine123在HEK及HEK-MDR細胞株中的uptake的情形 51
第五節. [3H]-Colchicine在HEK及HEK-MDR細胞株中的uptake的情形 53
第五章. 討論 70
參考文獻 73

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