正电子核素标记异喹啉类细胞凋亡显像剂的研究

Molecular imaging in vivo have important implication for early measurement of response in cancer patient with irradiation or chemotherapy. Caspase-3 was known to play essential and distinct roles in apoptosis. Caspase-3 was an attractive monitor target for evaluation of treatment tumor. We are interested in design, radiolabled with 18F or 11C, and biological evaluation of isoquinline-1,3,4-trione derivatives (11C/18F-iQLT) as potent apoptosis imaging agent, to get intrinsic properties such as good selectivity, high whole-cell activity, and stability in vivo. To check 11C/18F-iQLT uptake in tumor tissue as measured by autoradiography would be linked to apoptosis, which was quantified by TUNEL and caspase 3 staining of adjacent tissue sections at A549 tumor apoptosis of mice models.The tumor/muscle ratios of 11C/18F-iQLT from Micro-PET images in tumor-bearing mice was correlated with the degree of apoptosis or with block as measured by caspase-3 activity in the tumor homogenate .Biostability of 18F-iQLT was studied in mice. Molecular imaging of apoptosis in vivo may serve for other diseases such as brain and myocardial ischemic reperfusion injury, detection of allograft rejection. We aimed to get a novel,high-selectivity and specificity, suitable for use in vivo with minimal metabolism caspase-3 imaging agent for clinical practice.

分子影像用于肿瘤放化疗、疗效早期评价有较大的价值。Caspase3是凋亡关键酶和执行者，因此Caspase3活性显像检测细胞凋亡可以达到早期评价疗效目的。本研究拟采用正电子核素（F-18，C-11）标记Caspase3特异性小分子阻断剂：异喹啉1，3，4-三酮衍生物，用于细胞凋亡显像,以克服多肽类显像剂体内不稳定和靛红类显像剂靶本比低的不足。以肺癌A549化疗后细胞凋亡为模型，通过放射身显影、Caspas3免疫染色、TUNEL染色研究放射性摄取与凋亡的相关性，通过体内分布、MicroPET显像/阻断等研究靶/本比和体内特异性，研究放射性摄取的特异性；通过代谢研究显像剂体内分布和稳定性等。Caspase3活性显像可用于其它如心肌和脑的再灌注损伤、器官移植排异等影像评价。本研究希望为临床提供一类新的特异性、高靶本比的Caspase3活性显像剂，用于在体细胞凋亡显像。

分子影像用于肿瘤放化疗、疗效早期评价有很大的价值。Caspase3 是凋亡关键酶和凋亡的执行者，因此 Caspase3 活性显像检测细胞凋亡可以达到早期评价疗效目的。本研究设计了基于 Caspase3 特异性小分子阻断剂： 1，3，4-三羰基异喹啉衍生物结构的前体，采用正电子核素（F-18，C-11）进行放射性标记，通过A549细胞摄取实验、阻断实验、生物分布和PET显像等实验，深入研究了Caspase 3特异的异喹啉类的凋亡探针在体内外的生物学特性,主要合成并分析了三种探针11C-iQLT、18F-iQLT和11C-HIQPM。其中，11C-HIQPM合成简便、稳定性较高、并展现出良好的生物学分布性能。研究利用11CH3-Triflate与前体一步反应，经HPLC分离纯化得到2位N甲基化的产品11C-HIQPM，放射化学纯度大于98 %，比活度为1.13±0.27 Ci/μmol（n = 5），体外稳定性高；在体外，凋亡的A549细胞摄取11C-HIQPM最高的时间是20min，之后反而被细胞排出；A549细胞对11C-HIQPM的摄取率与细胞凋亡率相关（y = 0.0192 x + 0.1310，R2 = 0.9257）；Caspase 3特异阻断剂Ac-DEVD-CHO抑制凋亡细胞摄取11C-HIQPM的IC50为113.4 nM，表明11C-HIQPM能与Caspase 3特异结合，12C-HIQPM对Caspase 3的IC50为36 nM；11C-HIQPM在大鼠体内主要经肾脏代谢，其余器官存在少量非特异性摄取，肝细胞凋亡模型组大鼠肝脏摄取值为正常大鼠肝脏摄取值的1.7倍；荷A549瘤裸鼠经化疗后24h和48h肿瘤的放射性摄取值比化疗前分别增加了40%和68%（P＜0.05），化疗前、化疗后24h、化疗后48h，荷瘤鼠的肿瘤/肌肉比分别为1.3、2.0和1.8。利用PET/CT显像，荷A549瘤裸鼠化疗前，肿瘤/肌肉摄取值比（T/M）为1.9，化疗后，肿瘤/肌肉SUV比值为5.8，化疗后肿瘤摄取值是化疗前肿瘤摄取值的3.6倍，凋亡的肿瘤组织在PET图像中呈现“高亮”区域，与周围组织对比度高。通过正电子核素标记得到新型异喹啉类凋亡显像剂，体外细胞摄取实验和体内生物分布实验均表明该类探针能够特异结合活化的Caspase 3，并有潜力应用于在体凋亡细胞检测。