基于纳米探针的KRAS基因突变检测及其在结直肠癌靶向治疗疗效预测中的应用

Detection of KRAS gene mutation in tissue specimens before therapy is clinically important for predicting the response to anti-epidermal growth factor receptor (antiEGFR) treatment in colorectal cancer. Additionally, recent studies have also found that, dynamic analysis of KRAS gene mutation in peripheral blood offer the potentials to real-time monitor the occurrence of acquired resistant during antiEGFR therapy. However, the commonly used methods for KRAS mutation detection always exhibit relatively low sensitivity, which has thereby reduced their effectiveness in detection of KRAS mutation in peripheral blood. This project is intended to develop a gold nanoprobe based detection system of KRAS gene mutations.Benefiting from the extraordinary optical and physicochemical properties of gold nanoparticles, the detection sensitivity and selectivity of the established system are intended to be significantly improved, which thereby can offer the potentials for real-time monitoring the KRAS mutaion status in peripheral blood. Following the above investigations, the establised system will be applied to predict the response to antiEGFR therapy in colorectal cancer, and then, by detecting the KRAS mutation in blood specimens, the established system will be further adopted to real-time monitor the response during the anti-EGFR therapy. The investigation of the project will not only provide a novel altinative strategy for response prediction of antiEGFR therapy in colorectal cancer, but also expand the applications of nanotechnology to the fields of molecular biomarker-guided individual treatment in cancer.

针对活检或手术获得组织标本中KRAS基因的突变检测已广泛应用于临床中，在结直肠癌抗表皮生长因子受体靶向治疗前实现治疗疗效的有效预测。此外，外周血中KRAS基因突变状态的实时动态监测，更有望为靶向治疗过程中的继发性耐药的产生提供依据，为结直肠癌个体化治疗提供更为有效的策略。然而，传统基因突变检测技术较低的灵敏度极大限制了其在血中微量KRAS基因突变的有效检测。本项目拟基于金纳米探针发展灵敏特异的KRAS基因突变检测体系，旨在基于金纳米探针优越的光学及物理化学性质，进一步结合已应用于生物医学研究的突变位点特异性识别及扩增技术，构建适宜于血中微量KRAS基因突变的检测策略。并以此为基础，研究其在结直肠癌靶向治疗疗效预测及实时动态监测领域的潜在应用价值，为基于纳米技术的分子标志物指导下的恶性肿瘤个体化治疗研究提供新的方法、思路和实验依据。

针对活检或手术获得组织标本实现恶性肿瘤相关分子标志物检测难以实现恶性肿瘤疾病状态及治疗疗效的连续动态监测，无法满足目前恶性肿瘤精准治疗要求。针对这一瓶颈问题，本项目基于前沿纳米技术，开展了一系列外周血“液体活检”策略在恶性肿瘤治疗疗效连续动态监测及耐药相关机制动态演变中的研究。研究首先基于水相合成聚乙二醇稳定CdTe量子点，制备高荧光效率及生物相容性纳米光学免疫探针，并成功应用于鼻咽癌患者外周血中抗Epstein-Barr病毒壳抗原的免疫球蛋白A抗体的特异性检测中。在此基础上，本项目研究进一步引入前沿“液体活检”策略—循环肿瘤细胞（Circulating Tumor Cells，CTCs），系统深入研究了CTCs分子分型与胃癌耐药相关动态演变机制，及耐药相关CTCs亚型在治疗疗效动态监测中的可行性。研究首次系统证实，在进展期胃癌中，外周血CTCs计数与其治疗疗效及患者预后明显相关，治疗过程中动态监测CTCs计数升高可预示治疗疗效降低，进一步确证CTCs不同核型及表型与胃癌原发及继发性耐药的发生密切相关，揭示胃癌治疗耐药机制的动态演变。项目相关研究证实前沿“液体活检”策略在恶性肿瘤疾病状态及治疗过程精准监测中应用的广阔前景，并为基于纳米技术的恶性肿瘤早期临床诊断及外周血中循环分子标志物监测提供新的途径。因此，本项目研究将为基于纳米技术的前沿“液体活检”检测策略的进一步构建，及研究恶性肿瘤耐药相关动态演变机制奠定坚实的基础。