三重态-三重态湮灭上转换发光探针的设计合成及在体成像检测肿瘤研究

Fluorescence imaging in vivo is of great significance to uncover the role of molecules in biology, medicine, and the treatment of diseases. The materials based on triplet-triplet annihilation up-conversion luminescence show large absorption coefficient, high quantum yield, low excitation energy, readily adjustable excitation and emission wavelengths, etc. To detect tumor by imaging in vivo, a series of up-conversion luminescent probes will be reasonably designed and synthesized. The incorperation of organic chromophores will alter the energy levels to match that of metal centers, increasing the absorption of visible region and the life time of triplet excited states to obtain higher quantum yields of the up-conversion. A series of metal sensitizers will be prepared based on this strategy. By introducing iodine having heavy metal effects to the system, a series of halogenated organic fluorophores as organic sensitizers and single molecular up-conversion systems with higher quantum yield, conversion efficiency and atom economy will be synthesized. By coating the sensitizer and annihilation agent or the single molecular up-conversion system with carboxyl or amino derivatives of F-127 and successively organic silane hydrolysis, the up-conversion luminescence nanoparticles will be synthesized. By coupling nanoparticles with antibodies, a number of probes will be prepared to detect tumors by imaging in vivo.

荧光在体成像对于阐明分子的生物、医学作用及相关疾病治疗具有重要指导意义。三重态-三重态湮灭上转换发光具有吸收系数较大、量子产率较高、激发所需能量很低、激发和发射波长易于调节等优点。本课题拟制备系列上转换发光探针应用于在体成像检测肿瘤。向配体中引入有机发色团，能够促进配体和金属的能级匹配，增强可见光吸收和延长三重态寿命，从而提高上转换的量子产率。据此合成金属配合物敏化剂。通过引入碘原子产生类似重金属效应，合成卤代有机荧光分子敏化剂和量子产率、上转换效率、原子经济性都较高的单分子上转换发光体系。将敏化剂、湮灭剂，或上转换发光单分子体系，以羧基或氨基衍生的F-127包覆，有机硅烷试剂水解，制备上转换发光纳米颗粒；再将其与抗体偶联，制备上转换发光探针并应用于在体成像靶向检测肿瘤。

荧光成像对于阐明分子的生物、医学作用及相关疾病治疗具有重要指导意义。三重态-三重态湮灭上转换发光具有吸收系数较大、量子产率较高、激发所需能量很低、激发和发射波长易于调节等优点，正在得到迅速发展。本项目首先以F127包覆水解法或正相微乳液法设计和制备羧基或氨基修饰的上转换发光纳米颗粒并进行表征，再将其与靶向基团如表皮生长因子、环状RGD肽、埃罗替尼等偶联制备成像探针，最后将探针应用于成像检测肿瘤细胞。结果如下：（1）以金属配合物或卤代荧光素为敏化剂，通过F127包覆水解法或正相微乳液法制备了系列上转换发光硅纳米颗粒并进行了透射电镜、动态光散射、Zeta电位、红外光谱和上转换发光等表征。（2）制备了系列上转换发光成像探针并用于检测肿瘤细胞。（i）制备了以表皮生长因子为靶向的上转换发光成像探针，该探针对肺癌细胞A549（实验组）和人乳腺癌细胞MCF-7（对照组）的成像结果表明：以表皮生长因子为靶向的成像探针对A549细胞有良好的靶向检测作用。作为对比，未接表皮生长因子靶向基团的上转换发光纳米颗粒则不具有靶向检测效果。（ii）制备了以环状RGD肽c(RGDfK)为靶向的上转换发光成像探针，该探针对A549细胞和MCF-7细胞成像效果没有明显差异，需要进一步选择整合素αvβ3更低表达的或不表达的肿瘤细胞作对照组。（iii）埃罗替尼对肿瘤细胞有很好的靶向和抑制作用。制备了以埃罗替尼为靶向的香豆素荧光探针，初步实验表明埃罗替尼是检测A-549细胞的一个新颖的靶向点，将有望用于上转换发光探针的靶向基团。上转换发光靶向成像探针有很高的信噪比，只需很低的激发能量即能满足成像要求，能够以非常高的灵敏度、非常低的检测限特异性检测肿瘤细胞，在肿瘤的早期成像检测方面具有重要的应用前景。