新型PET-MR双模式小分子细胞凋亡显像剂的研制及其监测肺癌化疗疗效的实验研究

Induction of apoptosis is one of the primary mechanisms through which most chemotherapies cause tumor cell to death. Cell apoptosis/death is porven to be a very useful tool to assist in monitoring disease course and anti-tumor therapeutic effect earlier. However, in vivo cell apoptosis/necrosis imaging is still a challenging issue. Until now, traditional imaging methods(MRI and 18F-FDG PET) can not monitor the anti-tumor therapeutic effect early, which restrict the individual therapy. Although radiolabeled annexin V derivatives remain under extensive investigation, their current in vivo usefulness is found to be limited for their low target-to-background ratios and long biological half-life with high uptake in liver and kidney. Low-molecular weight probes are more amenable to structural optimization and display more favorable pharmacokinetic properties compared to protein-based probes like annexin V. It is discovered that rationally designed bis(zinc(II)-dipicolylamine) (Zn2+-DPA) coordination complexes can mimic the apoptosis sensing function of annexin V. Two meta-oriented Zn2+-DPA units are responsible for PS recognition and binding..In our previous study, we have successfully synthesized and radiolabeled Zn2+-DPA with 18F-SFB for the first time. And the chelate compound of 18F-FB-DPA with zinc is a new PET tracer for apoptosis imaging. PET imaging has demonstrated that 18F-FB-Zn2+-DPA is a sensitive and selective marker of apoptosis in vivo with potential utility in tumor. Based on our previous research on 18F-FB-Zn2+-DPA as a new low-molecular tracer for apoptosis imaging, the aim of this study is to design and synthesize dual-targeted 18F-FB-Zn2+-DPA labeled with 18F-SFB and superparamagnetic Gd3＋ for the first time, and to study its apoptosis imaging mechanism in vivo and in vitro through lung cancer cell and apoptotic tumor model mice, its pharmacokinetics, biodistribution and PET imaging characteristics. With the utility of high sensitivity of PET and high spatial resolution ability of MRI, the study is to evaluate the tumor cell apoptosis/necrosis early through radioactive uptake and signal intensity, and to analyze its characteristics of tumor cell apoptosis imaging. It is expected to construct the dual-targeted PET-MR apoptosis imaging model in vivo, and to provide the evidence for the further clinical application of low-molecular cell apoptosis imaging.

目前监测肿瘤化疗疗效常用的影像学方法MRI和18F-FDG PET显像均不能早期监测肿瘤疗效，制约了肿瘤的个体化治疗。大量研究证实细胞凋亡可用于肿瘤化疗早期反应的评估。目前，凋亡显像多通过各种报告要素标记的Annexin V与凋亡细胞外在的磷脂酰丝氨酸（PS）结合并成像，但Annexin V属于大分子物质，各种因素制约了其临床应用。N,N-二甲基吡啶类（DPA）是一种能够螯合Zn2+的小分子化学基团，能够靶向结合PS。本研究在前期针对细胞凋亡DPA类显像剂研究的基础上，首次构建放射性核素氟-18和顺磁性物质钆离子标记的双靶点小分子探针DPA(18F-FB-Zn2+-DPA/di-DPA-Gd-DTPA)，利用PET高敏感性和MRI高空间分辨率，从放射性摄取和信号强度两方面，通过肺癌细胞及肺癌动物模型，研究其与肿瘤细胞凋亡的作用机制、药代动力学和代谢特征，期望实现PET-MR双模式活体显像。

诊断治疗学是一个快速发展的研究领域，其内容涉及新型靶向探针、试剂和在体内特定的分子通路的显像方法，其中用于治疗和诊断用的高选择性药物的发展尤其重要。目前监测肿瘤化疗疗效常用的影像学方法MRI和18F-FDG PET显像均不能早期监测肿瘤疗效，制约了肿瘤的个体化治疗。因此，研发具有更加特异性的分子靶向探针具有十分重要的意义。本课题组成功研制成功了氨基酸类分子探针S-11C-甲基-L-半胱氨酸（11C-MCYS)，细胞凋亡类分子探针18F-FB-DPAZn2，以及神经降压素受体分子靶向探针，并分别进行了分子生物学的评价。此外还研究了恶性肿瘤转录翻译RNA结合蛋白（TARBP1）的表达与肿瘤进展分期的相关性。我们的研究结果：（1）11C-MCYS在体内吸收迅速。与正常组织比较，肿瘤组织放射性摄取（(0. .05) %ID/g）明显增高。肿瘤组织摄取11C-MCYS较高,SUVmax为4. .65,而炎性反应组织摄取11C-MCYS较低,SUVmax为1.18,肿瘤/肌肉及炎性反应/肌肉的放射性比值分别为7.27和1.62,差异有统计学意义(t=2.906, P<0.01)。所以，11C-MCYS是一种有前景的新型氨基酸类PET显像剂。 2）细胞凋亡类分子探针18F-FB-DPAZn2在恶性肿瘤化疗组的摄取较非化疗组的摄取明显增高。18F-FB-DPAZn2能够应用于细胞凋亡显像，与FITC-Annexin V相类似，是一种有前景的细胞凋亡PET分子探针。3）自行合成了放射性核素64Cu-和发光染料Cy5.5双标记的双模态靶向探针64Cu-DOTA-NT-Cy5.5。小动物PET图像定量分析显示在1小时和4小时时间点HT-29肿瘤对64Cu-DOTA-NT-Cy5.5摄取剂量分别为1.91±0.22, 1.33±0.14 %ID/g。对放射性和荧光染料摄取的强度，肿瘤与肌肉的比值具有很好的相关性。64Cu-DOTA-NT-Cy5.5是一种非常有前途的双模态探针，它可以进行NTR阳性表达的PET/fluorescence显像。4）与正常组织比较，TARBP1 mRNA在恶性肿瘤组织中具有明显的高表达。统计学分析显示TARBP1的高表达与肿瘤的AJCC分期及临床分期具有明显的相关性。多因素方差分析显示TARBP1可以作为一个独立性的恶性肿瘤生存率预测指标。