姜黄素肿瘤靶向上转换发光固体脂质纳米粒的制备及其光动力治疗的机理研究

Photodynamic therapy has become an important method for cancer treatment due to low-toxicity, rapid efficacy, strong targeting ability etc. It is a key point to find a suitable photosensitizer for this therapy. Curcumin (Cur), the composition of the traditional Chinese medicine, is one of the hot photosensitizers on research at present. Our previous studies, including the preparation of Cur-loaded solid lipid nanoparticles (Cur-SLN), its characterization, and its mechanism of the photo-toxicity to tumor cells, indicated that Cur-SLN is an excellent photosensitizer. But it was only excitated by blue light (λ= 450 nm) with a poor tissue penetration. Besides, it lacks the tumor targeting. In order to overcome these shortcomings, we plan to incorporate upconversion luminescent nanoparticles (UCNPs) into Cur-SLN, as well as graft the tumor targeting molecules such as folic acid to the surface of nanoparticles, so that constitute a new PDT drug nano-carrier system with a powerful tissue penetration (λex= 980 nm) and a high tumor targeted property. Furthermore, we will evaluate its security and anti-tumor effects in vitro and in vivo. Additionally, it is essential to discuss the characteristics and efficiency of its upconversion, as well as the mechanism of its inhibition on the proliferation of tumor cells. All together, these experiments would provide theoretical basis for looking for new PDT photosensitizer candidates and its formulations. Meanwhile, we would obtain a new kind of efficient and safe formulations of Cur that can be applied for the photodynamic therapy of cancer. Besides, we also would establish a new platform that can be used to screen and research PDT drug carrier system.

光动力疗法（PDT）以其毒性小、疗效快、靶向性强等优点成为肿瘤治疗的重要方法，如何发现合适的光敏剂是该疗法的关键。传统中药成分姜黄素（Cur）是目前研究的热点光敏剂之一，课题组的前期工作研究了姜黄素固体脂质纳米粒（Cur-SLN）的制备、表征和光动力机理，发现Cur-SLN是一种非常优秀的光敏剂。但该体系只能在组织穿透力差的450nm蓝光激发。为了克服这些缺点，本课题组拟将上转换发光纳米材料引入Cur-SLN内部，同时将叶酸等肿瘤靶向分子连接到纳米颗粒表面，构成一个能被穿透力强的980nm红外光高效激发，且具有肿瘤靶向性PDT纳米载药的新剂型。在细胞和动物水平上评价新剂型的安全性和抗肿瘤效果，探讨上转换的特性、效率与抗肿瘤细胞增殖的机制，为寻找具有新的PDT候选光敏剂及剂型提供理论依据，获得一种高效、安全用于肿瘤光动力治疗的姜黄素新剂型，并建立一套用于PDT载药体系筛选和研究的平台。

传统中药成分姜黄素（Cur）是目前研究的热点光敏剂之一，课题组的前期工作制备了姜黄素固体脂质纳米粒（Cur-SLN），该体系在450 nm蓝光激发下发挥PDT效果。但450 nm波长激发光组织穿透力较差，为了克服该缺点本课题制备了以上转化纳米颗粒为基础的纳米光敏剂新剂型。.运用连续逐层包被法制备980 nm波长近红外光激发的上转换纳米颗粒（UCNPs），再通过分子相互作用成功制备核/壳结构的Cur-UCNPs-PEI。通过TEM、紫外可见吸收光谱、荧光发射光谱等技术手段对所制备的Cur-UCNPs-PEI进行表征，确定了该纳米光敏剂新剂型的理化特性；验证了该纳米复合物的光学特性，存在上转换发光和能量共振转移现象。同时，从细胞和动物水平初步考察了Cur-UCNPs-PEI对神经胶质瘤肿瘤细胞U87的PDT效果，利用CCK8法和H&E染色法对该纳米复合物进行体内外安全性评价。. 合成的UCNPs粒径约为30 nm，具有均一的尺寸和形貌，并具有高度晶化的六方相结构。最终获得Cur-UCNPs-PEI，该纳米载药体系呈圆球状，粒径约为100 nm。经计算Cur-UCNPs-PEI的载药量约为21%，载药量较高，适合于用来进行肿瘤的光动力治疗。荧光发色光谱结果显示制备的UCNPs发射波长在450-500 nm处，发蓝光；搭载Cur制备得Cur-UCNPs-PEI，其发射峰与UCNPs的发射峰相比较，Cur-UCNPs-PEI在450-500 nm之间的发射峰消失了，是被姜黄素吸收了的结果。CCK8结果表明，UCNPs和980 nm近红外光均对U87细胞没有毒性；Cur-UCNPs-PEI在黑暗条件下对U87细胞没有明显毒性；经980 nm近红外光辐照之后，因PDT效应对U87细胞具有明显抑制作用。此外，动物实验结果进一步表明相比较于其它处理组，Cur-UCNPs-PEI 980 nm光照组可以更有效的抑制肿瘤在荷瘤小鼠体内生长，抑瘤率达到90%。同时，在治疗过程中各组实验裸鼠体重没有明显变化；对比各实验组的心、肝、脾、肺、肾组织H&E染色结果，同样没有明显差异。这些结果表明Cur-UCNPs-PEI 980 nm光照治疗没有组织毒性，具有良好的生物安全性。.研究结果表明，Cur-UCNPs-PEI作为一种新的纳米光敏剂新剂型对恶性胶质瘤的光动力治疗具有较好的应用前景。