构建级联双重补氧纳米载药系统以提高肿瘤光动力学治疗疗效

Hypoxia of tumor tissue is the key factor that influence photodynamic therapy of solid tumor. However, the single method of oxygenating can’t effectively reduce tumor environment of hypoxia. Hence, this project of application aims to make red blood cell-derived vesicle (RDV) and catalase into combination organically and realize cascaded double oxygenating to tumor tissue. Due to the tumor is rich in hydrogen peroxide, firstly we innovatively combine hydrophilic catalase with hydrophobic polycaprolactone to form nanoprotein micelles that as a photosensitizer carrier can also hydrolyze hydrogen peroxide to release oxygen. And then micelle is wrapped by RDV. Not only hemoglobin in RDV can firstly carry the oxygen which supplies PDT, but also it can produce reactive oxygen to damage cytomembrane and enter tumor tissue. Furthermore, RDV release micelle to hydrolyze hydrogen peroxide, and achieve cascaded double oxygenation by further oxygen supplement. At the same time, we enhance the tissue penetration of PDT by combining the up-conversion nanoparticles which our research group pre-construct with photosensitizer. After optimizing the preparation of the nanoparticle drug carrier system, our project makes detailed investigations on the ability of improving hypoxia in vivo or in vitro and the ability of treating tumors, in order to explore a new method to improve the efficacy of tumor photodynamic therapy by reducing the hypoxia environment.

肿瘤组织乏氧是影响实体肿瘤光动力学治疗的关键因素，而单一的补氧方法并不能有效改善肿瘤乏氧环境。所以本申请课题拟将过氧化氢水解酶Catalase与红细胞囊泡RDV进行有机结合，实现对肿瘤组织的级联双重补氧。由于肿瘤中富含过氧化氢，首先我们创新性地将亲水的Catalase与疏水的聚己内酯结合形成纳米蛋白胶束，使其作为光敏剂载体的同时还可以水解过氧化氢释放氧气。然后利用RDV将上述胶束进行包裹，RDV中血红蛋白携带的氧气率先供给PDT，产生的活性氧可以破坏RDV膜结构进入肿瘤组织，同时释放出蛋白胶束水解过氧化氢，进一步补充氧气达到级联双重补氧的目的。同时我们将课题组前期构建的上转换纳米粒子与光敏剂相结合，进而增强PDT组织穿透深度。本课题拟在优化制备上述纳米载药系统后，对其在体内外乏氧改善能力及肿瘤治疗能力等方面进行细致考察，以期探究出一套通过改善肿瘤乏氧环境提高肿瘤光动力学治疗疗效的新方法。

研究背景及目的:光动力疗法（Photodynamic Therapy，PDT）是利用光敏剂在特定波长光下产生活性氧杀死肿瘤细胞，是一种具有高时空选择性的微创光触发疗法。然而PDT治疗效果会受到肿瘤乏氧微环境的限制，因此我们构建了红细胞囊泡包裹过氧化氢酶蛋白胶束的纳米载药系统 Red Blood Cell-derived vesicles@Catalase-PCL@Ce6( RDV@CP@Ce6）将体外递氧与原位产氧有机结合起来，进行级联双重补氧，增强 PDT治疗效果，同时改善PDT后乏氧，调节肿瘤免疫微环境。.研究内容：1，优化合成及表征纳米载药系统 RDV@CP@Ce6；2，体外探究RDV@CP@Ce6的携氧能力、活性氧产生增强水平等 PDT 增强性能；体内验证RDV@CP@Ce6的肿瘤组织乏氧改善能力，免疫逃逸能力，体内分布，体内循环代谢情况等；3，体内考察RDV@CP@Ce6激活全身免疫的能力，包括对相关免疫细胞及细胞因子的评估等；4.体内评估RDV@CP@C6肿瘤治疗能力、体内毒副作用等；体内评估RDV@CP@Ce6联合免疫检查点抑制剂抗PD-1抗体治疗抑制肿瘤转移的效果。.结果：我们成功制备了仿生级联补氧纳米载药系统RDV@CP@Ce6，该体系粒径均一，体内外稳定性良好，体外实验表明，RDV@CP@Ce6具有优异的体外补氧能力及光动力效应，且能在PDT作用后充分补氧。体内实验结果表明，RDV@CP@Ce6能高效在肿瘤部位蓄积，有效进行双重级联补氧， PDT后补氧可有效激活全身免疫；级联双重补氧纳米载药系统增强PDT治疗后明显抑制肿瘤生长和转移，改善肿瘤免疫微环境。.结论：RDV@CP@Ce6具有良好的体内稳定性及良好的生物相容性，该体系通过RDV携带的氧气进行一级补氧有效增强PDT疗效，利用过氧化氢酶蛋白胶束分解瘤内过氧化氢产生的氧气进行PDT后的二次补氧，进一步缓解PDT后肿瘤微环境的乏氧，最终重塑瘤内免疫抑制微环境，激活全身免疫，抑制肿瘤生长和转移。