- 學位論文
Epirubicin HCl奈米粒子製備與其在癌細胞毒殺作用及膀胱壁穿透之研究
Preparation and Cytotoxicity Evaluation of Epirubicin-loaded Nanoparticles and its Penetration to Bladder
摘要
膀胱內化療藥物灌注常用於膀胱上皮細胞癌治療。很多文獻顯示膀胱內灌注Epirubicin(EPI)治療膀胱上皮細胞癌，效果不佳的原因在於藥物無法有效穿透膀胱組織。本實驗製備含有EPI的聚氰基丙烯酸乙基酯奈米粒子(poly ethylcyanoacrylate nanoparticles) 以改善膀胱內灌注藥物治療。 以乳化聚合法來製備EPI聚氰基丙烯酸乙基酯奈米粒子。並利用23複因子實驗設計法來探討界面活性劑的濃度、種類和聚合反應時媒液的pH值對於粒子大小及包埋率的影響。以兩組較佳處方 TW80 NP及F68 NP進行物化性質及生物活性分析。並利用螢光顯微鏡影像觀察EPI在膀胱壁的穿透情形；並利用冷凍切片及萃取組織切片的方式來定量EPI在膀胱壁的量。 從23複因子實驗設計法的統計結果得知界面活性劑的種類及聚合反應時媒液的pH值為影響粒徑大小的主要變因；而聚合反應時媒液的pH值是影響包埋率的主要變因。TW80 NP和F68 NP的平均粒徑大小為90 nm及230 nm；PI值分別為0.4及0.06；藥品的包埋率分別為63.19 %及57.28 %。以MTS assay來比較EPI水溶液、不含藥的奈米粒子及含藥奈米粒子對於T24及RT4膀胱癌細胞株的毒殺作用。相較之下，含藥的奈米粒子對兩種膀胱癌細胞株(T24及RT4)所產生的細胞毒性都比不含藥奈米粒子及EPI水溶液高。在穿透性的實驗，由螢光影像可清楚的看出，TW80 NP這個處方在尿路上皮上的螢光訊號較強；且在膀胱壁其EPI的穿透量也是最高。故本實驗的結果，EPI-PECA奈米粒子可被用於膀胱內灌注治療膀胱上皮癌值得被發展的劑型。
並列摘要
Intravesical administration of cytotoxic agents is commonly used for treatment of superficial bladder cancer. Many reports have shown that failure of intravesical therapy with Epirubicin(EPI) for the treatment of superficial urinary bladder cancer is due in part to the inability of the drug to penetrate the bladder tissue. The purpose of this study is to prepare EPI-loaded poly ethyl-2-cyanoacrylate (PECA) nanoparticles for intravesical improving the efficacy of intravesical chemotherapy. Epirubicin-loaded PECA nanoparticles were prepared by emulsion polymerization. 23 factorial designs was used to estimate the effects of the surfactant type, surfactant concentration and the pH value of the polymerization medium on the particle size and the entrapment efficiency. The physicochemical characterization and biological properties of the optimal formulation, 2% tween 80 PECA NP (TW80 NP) and 0.5% pluronic F68 PECA NP (F68 NP) were performed. Fluorescence microscopy imaging was employed to visualize the penetration of EPI into the bladder wall. Its distribution in the bladder wall was determined from sectioning of frozen tissue and extraction of tissue slices. By 23 factorial designs, the main effects of two factors are the surfactant type and the pH value of the polymerization medium on particle size; the pH value of the polymerization medium is the main effect on entrapment efficiency. The mean particle size of TW80 NP and F68 NP were 90 nm and 230 nm and the polydispersity index were 0.4 and 0.06, respectively. The entrapment efficiency of TW80 NP and F68 NP were 63.19 % and 57.28 %. The cytotoxic effects of EPI-SOL, blank nanoparticles or EPI-loaded nanoparticles on T24 and RT4 bladder cancer cell lines were determined by MTS assay. The EPI-PECA NP exhibited a higher cytotoxicity in both cell lines compared to the blank nanoparticles and EPI-SOL. In permeability study, the fluorescence imaging and tissue depth profiles revealed that TW80 NP had a strong penetration and localization of EPI within urothelium. In this study, a novel formulation of EPI-loaded nanoparticles was explored for intravesical instillation in the treatment of superficial blaader cancer.