双功能融合膜纳米粒子水凝胶的制备及其抗肿瘤疗效评价

Preparation and anti-tumor efficacy evaluation of a hydrogel loaded with bifunctional fusion membrane nanoparticles

目的制备双功能融合膜纳米粒子海藻酸盐水凝胶(fusion membrane nanoparticles/alginate,FM-NPs/ALG),在H22小鼠肝癌皮下瘤术后复发模型中评价其预防肿瘤复发的疗效并初步探究其免疫机制。方法分别提取H22肿瘤细胞膜(tumor cell membrane,TM)和草分枝杆菌细菌膜(Mycobacterium phlei membrane,MM),在脂质体挤出器上制备成粒径约为100 nm的融合膜,并将其负载于海藻酸盐(alginate,ALG)水凝胶中构成FM-NPs/ALG,在H22小鼠肝癌皮下瘤术后复发模型中以生理盐水作为对照组,观察FM-NPs/ALG延缓肿瘤复发的疗效,并使用流式细胞术等探究可能的抗肿瘤免疫机制。结果成功制备了TM和MM的融合膜纳米粒子(fusion membrane nanoparticles,FM-NPs),平均粒径为87.4 nm。体外实验表明,与对照组相比,FM-NPs使树突状细胞(dendritic cells,DCs)活化的比例提高了约3.4倍。动物实验证实FM-NPs/ALG治疗能显著延缓肿瘤复发,在30 d内小鼠的平均肿瘤体积仅为对照组的17.3%;流式细胞术显示,FM-NPs/ALG能显著促进肿瘤引流淋巴结中DCs的活化,从而提高肿瘤浸润性T淋巴细胞(tumor-infiltrating T lymphocytes,TILs)的比例。治疗期间,小鼠的体重、重要器官脏器和肝肾功能等各项指标均在正常范围内。结论FM-NPs/ALG可以有效促进TILs的比例,延缓H22肝癌皮下瘤切除术后的复发并延长荷瘤小鼠的生存期,且该治疗方式具有良好的安全性。

Objective To prepare an alginate(ALG)hydrogel loaded with bifunctional fusion membrane nanoparticles(FM-NPs/ALG),and to evaluate its efficacy in preventing tumor recurrence and preliminarily explore its immune mechanism in H22 postoperative recurrence model of subcutaneous tumor of liver cancer.Method H22 tumor cell membrane(TM)and Mycobacterium phlei membrane(MM)were extracted and prepared on a 100 nm extruder.The fusion membranes were loaded into ALG hydrogel to form FM-NPs/ALG.The efficacy of FM-NPs/ALG in delaying tumor recurrence was observed in H22 subcutaneous tumor recurrence model,and the possible anti-tumor immune mechanism was explored by flow cytometry.Result Fusion membrane nanoparticles(FM-NPs)with an average particle size of 87.4 nm were prepared successfully.In vitro experiments showed that FM-NPs increased the proportion of activated dendritic cells(DCs)by about 3.4 times compared with the control group.Animal experiments showed that FM-NPs/ALG treatment could significantly delay tumor recurrence,and the average tumor volume in mice within 30 days was only 17.3%of that in control group.Flow cytometry showed that FM-NPs/ALG significantly promoted the activation of DCs in tumor draining lymph nodes,thus increasing the proportion of tumor-infiltrating T lymphocytes(TILs).During the treatment,the body weight,vital organs,liver and kidney function and other indicators of mice were within the normal range.Conclusion FM-NPs/ALG can effectively promote the proportion of TILs,delay the recurrence of H22 hepatocellular subcutaneous tumor after resection and prolong the survival of tumor-bearing mice,and this treatment has good safety.