特异性分子探针11C-AZD9291 PET-CT检测肺癌EGFR蛋白突变

The presence of EGFR(epidermal growth factor receptor) gene-sensitive mutations is the basis of clinical use of EGFR-TKI(tyrosine kinase inhibitor) targeted therapy for advanced lung cancer. The main method of EGFR mutation detection is DNA analysis based on tissue samples of lung cancer, which is limited by tissue sampling and heterogeneity of tumor space-time. Furthermore, the EGFR mutation of ctDNA in peripheral blood is more specific and less sensitive, but it can not solve the problem in spatial heterogeneity. If the highly specific molecular probe of EGFR mutation is successfully synthesized, PET-CT imaging can be used to detect the mutation status of EGFR in vivo, and solve the problem in obtaining tissue and spatial and temporal heterogeneity. AZD9291 is the first commercially available third-generation EGFR-TKI, irreversibly and specifically binds to mutant EGFR, which is 40 times the binding force of wild-type EGFR, resulting in a higher tumor-to-background ratio, which has laid a material foundation for 11C labeled AZD9291 to be a PET-CT specific molecular probe for detection of EGFR mutations in lung cancer. In previous experiment, two kinds of 11C-AZD9291 precursor were designed and synthesized successfully. N-desmethyl AZD9291 was screened in the previous experiment and the synthesis of 11C-AZD9291 was accomplished by reacting it with 11C-Triflate-CH3. The aim of this study was using 11C-AZD9291 as the probe of PET-CT to explore the feasibility of detecting EGFR mutation and to find a method for detecting EGFR mutation at protein level by using different non-small cell lung cancer cell lines and their corresponding xenografts, recruiting 10 patients with lung cancer for the clinical research, the control group is using non-specific molecular probe 11C-erlotinib as a probe.

存在EGFR基因敏感突变是临床使用EGFR-TKI靶向治疗晚期肺癌的依据。目前EGFR突变检测的标准方法为基于肺癌组织的DNA分析，受到取材、肿瘤时空异质性的局限。AZD9291能与突变EGFR不可逆特异性结合，是野生型EGFR结合力的40倍，可形成高靶/本比，为11C-AZD9291作为PET-CT特异性分子探针检测肺癌EGFR突变奠定了物质基础。课题组自主设计并合成了2种11C-AZD9291前体药物，前期实验已筛选出N-去甲基AZD9291，并采用标记技术，得到11C-AZD9291。本研究以11C-AZD9291为分子探针，与非特异探针对照，不同EGFR状态肺癌细胞株、荷瘤裸鼠模型为材料，采用标准品阻断，行MicroPET显像及相关研究，并比对其与EGFR基因突变状态的一致性。另招募10例肺癌患者，探索PET-CT检测EGFR突变的可行性，为蛋白质水平检测EGFR突变提供新探针。

目的：本课题以非小细胞肺癌（NSCLC）中EGFR为靶点，设计并合成了11C-AZD9291探针，评价其作为靶向肺癌EGFR显像剂的可行性，评估其用于筛选EGFR-TKIs敏感个体的潜力。 方法：在自动化合成模块中利用11C标记N-去甲基-AZD9291合成11C-AZD9291。体外实验中，测定了11C-AZD9291探针的脂水分配系数，并评价其稳定性。检测NSCLC细胞系A549、HCC827和H1975对AZD9291抑制的敏感性，研究11C-AZD9291在这三种细胞系中的摄取和内化率的差异。体内实验中，进行了11C-AZD9291探针在正常小鼠内的急性毒性、药物动力学及体内生物分布的研究。进行了11C-AZD9291探针在具有不同EGFR基因突变及蛋白表达类型的A549、HCC827及H1975荷瘤鼠模型中的Micro PET/CT显像，并对比了18F-FDG与11C-AZD9291探针对荷瘤鼠内肿瘤的显像效果。最后对具有不同EGFR基因突变患者进行了11C-AZD9291探针PET/CT显像并与常规的18F-FDG PET/CT显像进行比对。结果：1. 11C-AZD9291探针的自动化合成时间为15分钟，放射性化学纯度大于99%。2.对AZD9291抑制敏感的HCC827细胞及H1975细胞对11C-AZD9291的摄取值均大于不敏感细胞A549。3. 11C-AZD9291在荷H1975瘤鼠的各脏器摄取值比正常小鼠摄取值更高，在5、15、30及60分钟时肿瘤的摄取值分别为1.17±0.17 %ID/g、1.66±0.49 %ID/g、1.40±0.34 %ID/g及1.46±0.47 %ID/g。4. 11C-AZD9291在荷HCC827瘤鼠及荷H1975瘤鼠内的肿瘤组织的SUVmax均高于荷A549瘤鼠肿瘤，其中荷H1975瘤鼠肿瘤的SUVmax值最高（P <0.05）。与18F-FDG对比，11C-AZD9291在靶向显像具有EGFR T790M与 L858R双突变的荷H1975及具有19del 突变的HCC827肿瘤具有优势性。结论：11C-AZD9291可在不同EGFR基因突变患者肺癌组织聚集，间接识别突变的EGFR蛋白，可运用于临床筛选具有EGFR敏感突变的肺癌，但11C-AZD9291在正常组织中的分布会弱化PET显像的图像质量。

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