血管內皮生長因子(VEGF)為調控血管新生(angiogenesis)機制的一個重要因子,通常在各類的腫瘤中都有過量的表現。在本研究中,將聚乙二醇微脂體(pegylated liposome)之表面修飾上予以耦合一人類抗血管內皮生長因子(anti-VEGF)單株抗體,bevacizumab(商品名:Avastin)進而建立一免疫微脂體(immunoliposome)脂藥物傳遞系統。此免疫微脂體為一奈米粒子大小約為100 nm,在許多腫瘤中可表現出通透性增強及停滯效應(enhanced permeability and retention effect, EPR effect),而修飾在其表面之bevacizumab對腫瘤細胞所分泌的VEGF有特異性結合進而提供靶向性之效應。此外, 免疫微脂體能透過螯合劑DTPA將放射性核種銦-111(indium-111)包埋在微脂體中心親水層，為一值得嚐試之造影劑,期進而達到診斷方面之需求與策略。本次實驗使用有過量表現VEGF之細胞株之人類大腸結腸癌細胞,LS174T 作為此實驗特異性免疫微脂體之標靶細胞。而特異性免疫微脂體(Bev-111In-liposome)與非特異性微脂體(111In-liposome)則植有LS174T腫瘤的小鼠做生物分佈與影像之比較。
方法:本實驗製備具有靶向性之免疫微脂體並包埋銦-111放射性核種與製備非靶向性之微脂體包埋銦-111放射性核種。將此二試劑由尾靜脈注入植有LS174T腫瘤的小鼠,分別比較時間點在1, 4, 8,24, 48, 72, 96小時的生物分佈與時間點在1, 4, 24, 48, 72, 96小時的影像。靶向性免疫微脂體之藥物動力研究進行時間點為0.25,0.5, 1, 4, 6, 24, 48, 72, 96,與168小時。
結果: 活體外的穩定性測驗顯示靶向性免疫微脂體無論在生理食鹽水或在老鼠血清中在長時間中皆有高穩定性。而在藥物動力研究方面, 靶向性免疫微脂體的t1/2α(起始再分佈相之半衰期)與t1/2β(排除相之半衰期)之值分別為0.666 and 13.975 小時。腫瘤在24小時有最高的藥物吸收量, 靶向性免疫微脂體與飛靶向性微脂體吸收值分別為14.24±3.59 %ID/g 與15.3±3.66 %ID/g。mircoSPECT 影像顯示靶向性免疫微脂體能夠在給藥後1小時偵測到腫瘤位置早於非靶向性微脂體的24小時。
結論: 實驗顯示,在診斷植有LS174T腫瘤隻小鼠,靶向性免疫微脂體與非靶向性微脂體皆有潛力的診斷劑。而靶向性免疫微脂體,Bev-111In-liposome在偵測一定程度上大之LS174T腫瘤上擁有以早於非靶向性微脂體之診斷潛力。

Vascular endothelial growth factor (VEGF) is one of important key factors of angiogenesis which is often expressed by a variety of tumors. In this study, we have developed an immunoliposome (IL) system that a liposome was pegylated and conjugated with bevacizumab, a humanized anti-VEGF monoclonal antibody. In this drug delivery system, the fabricated immunoliposome was of a nanoparticle that could passive retention in a variety of tumors through enhanced permeability and retention effect (EPR effect), with a size around 100 nm where bevacizumab could act a function targeting to VEGF for cancer cell. Furthermore, the immunoliposome was encapsulated with the radionuclide, indium-111 via DTPA as the complexing agent for imaging purpose. Human colorectal cancer cell, LS174T with overexpression of VEGF was adopted as the targeting cell in this work. An animal model bearing the tumor was used for the in vivo biodistribution and imaging studies.
Methods: The 111In entrapped immunoliposome, bevacizumab-111In- liposome (abbreviated as Bev-111In-liposome) and 111In entrapped liposome, 111In-liposome were prepared. The Bev-111In-liposome and 111In-liposome were subjected to administrate respectively into the LS174T tumor bearing mice via the tail vein. Biodistribution and microSPECT/CT imaging were carried out at the following postinjection times, 1, 4, 8, 24, 48, 72 and 96 h. Pharmacokinetics was simultaneously studied along with the postinjection times, 0.25, 0.5, 1, 4, 6, 24, 48, 72, 96, and 168 h.
Results: The in vitro stability study indicated that Bev-111In-liposome was quite stable either in both normal saline and rat plasma for a significant long time, i.e., at least 120 h. For pharmacokinetics study, the the t1/2α (initial redistribution phase with a short half-life) and t1/2β (elimination phase with a longer half-life) of Bev-111In-liposome were 0.666 and 13.975 h, respectively. The tumor uptake could reach maximum at 24 h postinjection, at 15.3±3.66 %ID/g for 111In-liposome and at 14.24±3.59 %ID/g for Bev-111In- liposomes, respectively. The results of microSPECT/CT imaging showed that the immunoactive liposome could clearly target the tumor at l h, i.e., much earlier than the passive liposome at 24 h.
Conclusions: Both of 111In-liposome and Bev-111In-liposome showed the potentiality as a diagnostic agent in the study using the VEGF overexpressive tumor, LS174T bearing mouse model. However, Bev-111In-Liposome could detect the localization of the tumor as early as at 1 h while much later for the 111In-Liposome.

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