放射性標靶微脂體E[c(RGDyK)]2-188Re-Liposome之研發與植有C26腫瘤小鼠之療效評估

血管新生(Angiogenesis)在腫瘤細胞增生與實體腫瘤形成上扮演著不可或缺的角色。在腫瘤血管新生之過程中，內皮細胞形成新的微血管時，會表現相當量的αvβ3整合素，由於αvβ3整合素與具Arg-Gly-Asp (RGD)序列之胜具有高度的專一結合性，因此，選擇αvβ3整合素作為腫瘤的生物性標記，開發標靶αvβ3整合素之治療劑乃是治療高血管新生腫瘤一新的策略。本研究旨在開發一包埋放射性藥物之標靶微脂體。此放射性標靶微脂體之建構係於微脂體表面耦合特定胺基酸序列Arg-Gly-Asp (RGD)，並同時於微脂體核內包埋Re-188放射性核種作為放射性治療藥物，以期導引正確腫瘤位置，以達到精確治療之目的。方法︰首先將RGD胜E[c(RGDyK)]2耦合於微脂體表面，同時將Re-188放射性核種以188Re-BMEDA形態包埋至微脂體，得到E[c(RGDyK)]2-188Re-liposome。另一方面，亦製備未具RGD胜耦合之188Re-liposome。以具有大量αvβ3整合素表現之人類臍帶靜脈內皮細胞(Human umbilical vein endothelial cells)進行細胞攝取試驗，並以螢光顯微鏡觀察。進而以植有C26腫瘤小鼠，進行造影、生物分佈及藥物療效評估等試驗。結果︰在細胞攝取試驗，靶向性微脂體E[c(RGDyK)]2-188Re-liposome於人類臍帶靜脈內皮細胞之攝取比非靶向性微脂體高出四倍之多，於螢光顯微鏡觀察下，靶向性微脂體之螢光強度亦明顯優於非靶向性微脂體。在活體外藥物穩定試驗顯示此靶向性微脂體在大鼠血清中，於72小時後仍具80%以上之穩定度。microSPECT/CT影像與生物分佈試驗顯示E[c(RGDyK)]2-188Re-liposome在植有C26腫瘤小鼠中，具有較高累積於網狀內皮系統(Reticuloendothelial system)，進而導致腫瘤部位累積量少於188Re-Liposome。但在療效評估試驗，E[c(RGDyK)]2-188Re-liposome與188Re-liposome對於抑制腫瘤生長並無顯著差異。結論︰本研究證實靶向性微脂體E[c(RGDyK)]2-188Re-liposome於活體外細胞實驗上具有高親和力與高穩定度。在動物實驗部份，本研究發現此靶向性微脂體於動物活體內易被網狀內皮系統所辨識，因而降低腫瘤部位之累積。未來需進一步針對E[c(RGDyK)]2-188Re-liposome表面之修飾進行探討，以減少網狀內皮系統吸收，改善於腫瘤位置之累積，以期達到腫瘤抑制功效。

關鍵字

血管新生； RGD ；微脂體；錸-188 ；放射治療

並列摘要

Angiogenesis is of a general property and is critical for tumor growth, where αvβ3 integrin is overexpressed on angiogenic endothelium in and around tumor tissue. αvβ3 integrin may therefore represent a possible target for drug delivery. In this study, a drug delivery system by a modified liposome specifically aiming at angiogenic tumor endothelial cells was developed. E[c(RGDyK)]2 peptide with affinity to this integrin was adopted to conjugate on the liposome surface and the 188Re radionuclide was simultaneously encapsulated in the liposome core. The resulted liposome, referred to E[c(RGDyK)]2-188Re-liposome, was investigated for its application potential in radionuclide therapy and diagnostic imaging for tumors. Methods: In this study, an angiogenesis-targeting liposome system was constructed to have dimeric RGD peptide E[c(RGDyK)]2 conjugated on the liposome surface and a rhenium-188 radionuclide complex encapsulated inside the liposome core. The in vitro cellular uptake by radiotracing and fluorescence microscope imaging was studied using human umbilical vein endothelial cells with overexpression of αvβ3 integrin receptor. For animal trial, the main works comprised microSPECT/CT imaging and biodistribution study and investigation of antitumor efficacy of E[c(RGDyK)]2-188Re-liposomes in comparison with non E[c(RGDyK)]2 conjugated liposome using C26 murine colon tumor-bearing animal model. Results: The in vitro stability study showed that E[c(RGDyK)]2-188Re-liposome was quite stable in rat plasma significantly for a long time, i.e., at least 72 h. The in vitro cell binding study demonstrated that the active targeting liposome surpassed non-targeted liposome in about four-fold higher of the cellular as observed from cellular uptake and cell staining. For in vivo studies, both of microSPECT/CT images and biodistribution studies indicated that the accumulation in the reticuloendothelial system for E[c(RGDyK)]2-188Re-liposome was apparently higher than the non-targeted liposome. The tumor uptake of E[c(RGDyK)]2-188Re-liposome was about six-fold lower than the non-targeted liposomes. Nevertheless, E[c(RGDyK)]2-188Re-liposome did inhibit the tumor growth as compared to untreated controls but no significant difference was observed in comparison with the non-targeted liposome. Conclusion: This study demonstrated the high cell binding affinity and stability of the active targeting liposome in the cell experiments. It was revealed from this study that the decrease of 188Re radioactivity accumulation in the colon tumor was relevant to the RES system uptake of E[c(RGDyK)]2-188Re-liposome. Modification on the E[c(RGDyK)]2-188Re-liposome surface to enhance the tumor uptake would be the future work to be investigated.

並列關鍵字

Angiogenesis ； RGD ； Liposome ； Rhenium-188 ； Radiotherapy

參考文獻

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放射性標靶微脂體E[c(RGDyK)]2-188Re-Liposome之研發與植有C26腫瘤小鼠之療效評估

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