- 學位論文
放射性碘標誌胸腺嘧啶鹼基化合物[123I]IaraU作為基因探針之研究
Evaluation of 123I-labeled 2'-hydroxy-2'-deoxy-1-β-D-arabino-furanosyl-5-iodouracil ([123I]IaraU) as a gene probe
摘要
本實驗的目的在於發展新的基因探針。在研究中測試放射性標誌嘧啶核苷的類似物(123I-labeled 2’-hydroxy-2’-deoxy-1-β-D- arabino-furanosyl-5-iodouracil ([123I]IaraU)，作為觀察HSV1-sr39tk基因表現的造影劑之可能性。實驗方法：利用老鼠的攝護腺癌細胞（Tramp-C1），同時轉殖突變型自殺基因（HSV1-sr39tk）及細胞間素-3（IL-3），在活體外試驗，以經轉殖的細胞Tramp-C1/IL3-tk為實驗組，野生型Tramp-C1為對照組，測量細胞吸收[123I]IaraU的情形。在活體內試驗，以Tramp-C1/IL3-tk及Tramp-C1細胞分別種植於近交系小鼠的皮下，待形成腫瘤後，將[123I]IaraU注入體內，ㄧ則以犧牲動物方式求取體內器官組織的分布，另ㄧ則藉由動物用微型單光子/電腦斷層融合影像儀（microSpect/CT）觀察[123I]IaraU在老鼠體內的分布情形。實驗結果：以Tramp-C1/IL3-tk及Tramp-C1細胞測試細胞毒殺濃度（CC50），發現[123I]IaraU對於tk基因轉殖之細胞不具有專一性，Tramp-C1/IL3-tk及Tramp-C1的CC50之值相近，分別為363.9μM及 309.7μM。注入8小時後的細胞吸收測試，得到Tramp-C1/IL3-tk細胞吸收稍略大於Tramp-C1細胞，並得知[123I]IaraU停留在細胞質而未進入細胞核。[123I]IaraU注入植有腫瘤之老鼠所得到生物分布結果在6小時胃腸道有大量吸收，24小時後從體內代謝，所得到之microSpect/CT影像，於注射後一小時略有滯留tk轉殖腫瘤的影像，同時在胃腸道有大量吸收的情形。結論：由本實驗得知，[123I]IaraU需進一步詳細測試以確認作為觀察HSV1-sr39tk基因系統表現的基因探針之可能性。
並列摘要
Evaluating a new probe for monitoring gene expression for the gene therapy efficacy was dealt with in the study. The radiolabeled nucleoside analogue, 5-123I-iodo-2’-hydroxy-2’- deoxy-1-β-D- arabinofuranosyl-uracil ([123I]IaraU), was proposed to be a substrate for imag-ing HSV1-tk expression. Methods: The rat prostate cancer cell line Tramp-C1 was trans-duced with HSV1-sr39tk and IL-3. The transduced Tramp referred as Tramp-C1/IL3-tk to-gether with the Tramp-C1 as a control was employed for the test of in vitro uptake of [123I]IaraU. Tramp-C1/IL3-tk cells as well as the Tramp-C1 cells were subcutaneously inocu-lated to produce the xenografts in C57BL/6J mice. Biodistribution study was carried out for the animals after being [123I]IaraU injected for 6h and 24h. The animals were microSpect/CT imaged after 1h postinjection. Results: The cytotoxicity of IaraU was surveyed on Tramp-C1/IL3-tk and Tramp-C1. There was no significant difference between the tk trans-duced and the wild-type cells. The CC50 of IaraU was 309.7μM for Tramp-IL3-tk and 363.9μM for Tramp-C1. The cell uptake for Tramp-C1/IL3-tk by [123I]IaraU was slightly greater than Tramp-C1 after incubation for 8 h. The cell internalization test showed that Tramp-C1/IL3-tk cells trapped [123I]IaraU in the cytoplasm. For in vivo trial, Tramp-C1/IL3-tk and Tramp-C1 were inoculated subcutaneously into the left and right flank of C57BL/6J mice separately. After the xenograft was generated, [123I]IaraU was injected via tail vein for biodistribution study and microSpect/CT imaging. The biodistribution results showed that GI tract retention at 6h, and metabolized from the body at 24h. The imaging showed early (1h) ambiguous localization of [123I]IaraU in Tramp-C1/IL3-tk xenograft, and no accumulation in the Tramp-C1 xenograft. Simultaneously, the image showed significant and apparent retention of [123I]IaraU in GI tract. Conclusion: [123I]IaraU has not sufficiently evidenced as an efficacious gene probe for monitoring the gene expression in the HSV1-sr39tk system from the study.
參考文獻
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