Lin Ming-Hsien, Wu Shih-Yen, Wang Hsin-Ell, Liu Ren-Shyan, Chen Jyh-Cheng
Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, No. 155 Li-Nong Street, Section 2, Pei-Tou, Taipei 11221, Taiwan; Division of Nuclear Medicine, Taipei City Hospital Zhongxiao Branch, No.145, Zhengzhou Rd., Datong Dist., Taipei City 10341, Taiwan; Program in Molecular Medicine, National Yang-Ming University and Academia Sinica, No. 155 Li-Nong Street, Section 2, Pei-Tou, Taipei 11221, Taiwan.
Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, No. 155 Li-Nong Street, Section 2, Pei-Tou, Taipei 11221, Taiwan.
Appl Radiat Isot. 2016 Feb;108:1-7. doi: 10.1016/j.apradiso.2015.11.017. Epub 2015 Dec 1.
Apoptosis has been suggested as a cytocidal mechanism of the HSV1-tk-expressing cells when exposed to ganciclovir (GCV). This study evaluated the efficacy of (111)In-labeled Annexin V for monitoring tumor responses during prodrug activation gene therapy with HSV1-tk and GCV.
Annexin V was conjugated to DOTA using N-hydroxysulfosuccinimide (sulfo-NHS) and 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC), labeled with (111)In-InCl3 and purified using size exclusion chromatography to give (111)In-DOTA-Annexin V conjugate. The radiochemical yield and the radiochemical purity of (111)In-DOTA-Annexin V were 74±12% and 98±3%, respectively (n=10). (111)In-DOTA-BSA was prepared similarly. An in vitro study to demonstrate the apoptosis of NG4TL4-STK cells after GCV treatment has been performed. Mice bearing NG4TL4-STK and NG4TL4-WT tumors were treated with GCV (10 mg/kg daily) by i.p. injection for 7 consecutive days. Before and during the GCV treatment, biodistribution studies and scintigraphic imaging were performed at 2h post injection of the radiotracers.
The uptake of (111)In-DOTA-Annexin V in treated cells (13.41±1.30%) was 4.1 times higher than that in untreated cells (3.21±0.37%). The GCV-induced cell apoptosis in NG4TL4-STK tumor resulted in a significantly increasing accumulation of (111)In-DOTA-Annexin V (1.92±0.32%ID/g at day 0, 4.79±0.86%ID/g at day 2, 4.56±0.58%ID/g at day 4) was observed, but not for that of (111)In-DOTA-BSA. During consecutive GCV treatment, scintigraphic imaging with (111)In-DOTA-Annexin V revealed high uptake in NG4TL4-STK tumor compared with that in NG4TL4-WT tumor. However, no specific (111)In-DOTA-BSA accumulation in NG4TL4-STK and NG4TL4-WT tumors was observed throughout the course of GCV treatment.
This study demonstrated that (111)In-DOTA-Annexin V can be used for monitoring tumor cell apoptosis during prodrug activation gene therapy with HSV1-tk and GCV for cancer treatment.
有研究表明，当表达单纯疱疹病毒1型胸苷激酶（HSV1-tk）的细胞暴露于更昔洛韦（GCV）时，细胞凋亡是一种细胞杀伤机制。本研究评估了111铟标记的膜联蛋白V在HSV1-tk和GCV的前体药物激活基因治疗过程中监测肿瘤反应的疗效。
使用N-羟基琥珀酰亚胺（磺基-NHS）和1-乙基-3-[3-（二甲氨基）丙基]碳二亚胺（EDC）将膜联蛋白V与DOTA偶联，用111铟-氯化铟（111In-InCl3）标记，并通过尺寸排阻色谱法纯化，得到111铟-DOTA-膜联蛋白V偶联物。111铟-DOTA-膜联蛋白V的放射化学产率和放射化学纯度分别为74±12%和98±3%（n = 10）。同样制备了111铟-DOTA-牛血清白蛋白（111In-DOTA-BSA）。进行了一项体外研究，以证明GCV处理后NG4TL4-STK细胞的凋亡情况。对携带NG4TL4-STK和NG4TL4-WT肿瘤的小鼠腹腔注射GCV（每天10 mg/kg），连续注射7天。在GCV治疗前和治疗期间，在注射放射性示踪剂后2小时进行生物分布研究和闪烁成像。
经处理细胞对111铟-DOTA-膜联蛋白V的摄取（13.41±1.30%）比未处理细胞（3.21±0.37%）高4.1倍。GCV诱导的NG4TL4-STK肿瘤细胞凋亡导致111铟-DOTA-膜联蛋白V的蓄积显著增加（第0天为1.92±0.32%ID/g，第2天为4.79±0.86%ID/g，第4天为4.56±0.58%ID/g），但111铟-DOTA-BSA未出现这种情况。在连续GCV治疗期间，用111铟-DOTA-膜联蛋白V进行的闪烁成像显示，NG4TL4-STK肿瘤的摄取高于NG4TL4-WT肿瘤。然而，在整个GCV治疗过程中，未观察到NG4TL4-STK和NG4TL4-WT肿瘤中有特异性的111铟-DOTA-BSA蓄积。
本研究表明，111铟-DOTA-膜联蛋白V可用于在HSV1-tk和GCV的前体药物激活基因治疗癌症过程中监测肿瘤细胞凋亡。
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