光动力脂质体凝胶协同曲妥珠单抗体外杀伤耐药性乳腺癌细胞的效果

Cytotoxic effect of photodynamic liposome gel combined with trastuzumab on drugresistant breast cancer cells in vitro

目的制备负载光敏剂二氢卟吩(Ce6)和肿瘤靶向药曲妥珠单抗(Tmab)的脂质体凝胶(Ce6-PC-Tmab@A-Gel),实现对耐药性HER2+乳腺癌的光动力治疗(PDT)和靶向治疗。方法采用旋蒸薄膜水化法制备Ce6-PC-Tmab脂质体,检测其纳米载体一般特性、包封率及近红外光响应性,同时采用冷冻干燥法及搅拌交联法,制备具有剪切响应性脂质体凝胶Ce6-PC-Tmab@A-Gel,通过扫描电镜(SEM)观察其微观结构,通过流变仪评价凝胶的剪切响应性。细胞毒性试验(MTT)检测Ce6-PC-Tmab@A-Gel联合近红外光对于SK-BR-3细胞株的抑制效果。结果制备的Ce6-PC-Tmab粒径为239.7±9.7 nm,电位为-2.03±0.09 mV,Ce6-PCTmab脂质体中Tmab的包封率为(40.22±0.73)%;形成脂质体凝胶后剪切响应性良好;具有优良的近红外光响应释放特性,随近红外光刺激强度增加和时间延长,Tmab释放均增多;Ce6-PC-Tmab@A-Gel在室温下性质稳定,在模拟肿瘤微环境中(pH 6.25)仍然具有稳定的结构;细胞毒性试验中,对耐药性HER2+人源乳腺癌细胞SK-BR-3的增殖有明显抑制作用,Ce6-PC-Tmab@AGel联合近红外光治疗组的SK-BR-3细胞生存率为(31.37±1.73)%,与未治疗组及其他实验组比较差异有统计学意义(P<0.01),同时具备较高的活性氧(ROS)产生效率,Ce6-PC-Tmab@A-Gel治疗组ROS释放量在光照2 min后达到(22.36±0.11)%;对于耐药性乳腺癌细胞的杀伤具有显著效果(P<0.01)。结论本研究制备的Ce6-PC-Tmab@A-Gel粒径小,均一性好,有良好的近红外光响应释放特性,保证了Ce6-PC-Tmab@A-Gel中Tmab的高效靶向治疗性,可注射凝胶体系为肿瘤局部的长时释药提供了可能。

Objective To evaluate the cytotoxic effect of photodynamic therapy(PDT)combined with targeted therapy using cross-linked liposomes and gels(Ce6-PC-Tmab@A-Gel)loaded with photosensitizer Chlorin(Ce6)and the tumor-targeting drug Trastuzumab(Tmab)in drug-resistant HER2+breast cancer cells.Methods Ce6-PC-Tmab liposomes were prepared using the thin-film hydration method.The general properties,encapsulation efficiency and near-infrared responsivity of the nanoparticles were evaluated.Ce6-PC-Tmab@A-Gel with a shear response was prepared by freeze drying and stirring crosslinking,and its microstructure was observed with scanning electron microscopy(SEM)and the shear response evaluated using a rheometer.The inhibitory effect of Ce6-PC-Tmab@A-Gel in drug-resistant HER2+breast cancer SK-BR-3 cells was assessed with cytotoxicity assay(MTT assay)combined with near-infrared light.Results The particle size of Ce6-PC-Tmab was 239.7±9.7 nm and the potential was-2.03±0.09 mV.The entrapment efficiency of Tmab by Ce6-PC-Tmab liposomes was(40.22±0.73)%.The prepared Ce6-PC-Tmab@A-Gel had a good shear response with excellent drug release characteristics under nearinfrared light,and increased intensity and duration of near-infrared light exposure enhanced Tmab release from the gel.Ce6-PC-Tmab@A-Gel was stable at room temperature and in a simulated tumor microenvironment(pH 6.25).Cytotoxicity assay(MTT)showed that Ce6-PC-Tmab@A-Gel combined with near-infrared light resulted in a survival rate of(31.37±1.73)%in SKBR-3 cells,much lower than that in the control group and other treatment groups(P<0.01);the combined treatment also had a high efficiency of ROS production,and ROS release reached(22.36±0.11)%after 2 min of near-infrared light exposure.Conclusion The prepared Ce6-PC-Tmab@A-Gel has good near-infrared light response release characteristics to ensure effective targeted therapy with Tmab.The injectable gel system potentially allows long-term local drug release in the tumor to improve the treatment efficacy against drug-resistant breast cancer.