南方医科大学学报

南方医科大学学报 ›› 2022, Vol. 42 ›› Issue (1): 26-35.doi: 10.12122/j.issn.1673-4254.2022.01.03

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偶联 CD133 核酸适体的载紫杉醇 PLGA-PEG 纳米载体靶向清除CD133阳性肺癌干细胞

庞丽莹，黄小龙，朱玲玲，肖韩艳，李梦雨，关会林，高 洁，金 红   

  1. 牡丹江医学院第一临床医学院，黑龙江 牡丹江 157011；牡丹江医学院附属红旗医院检验科，黑龙江 牡丹江 157011；湖北省武汉市第一医院呼吸与危重症医学科，湖北 武汉 430022；上海大学转化医学研究院，上海 200444；牡丹江医学院附属红旗医院血液科，黑龙江 牡丹江 157011；牡丹江医学院附属二院妇产科，黑龙江 牡丹江 157011；牡丹江医学院科研处，黑龙江 牡丹江 157011
  • 出版日期:2022-01-20 发布日期:2022-03-02

Targeted killing of CD133+ lung cancer stem cells using paclitaxel-loaded PLGA-PEG nanoparticles with CD133 aptamers

PANG Liying, HUANG Xiaolong, ZHU Lingling, XIAO Hanyan, LI Mengyu, GUAN Huilin, GAO Jie, JIN Hong   

  1. First School of Clinical Medicine, Mudanjiang Medical University; Clinical Laboratory, Department of Hematology, Affiliated Hongqi Hospital, Mudanjiang Medical University; Department of Respiratory and Critical Care Medicine, Wuhan First Hospital; Institute of Translational Medicine, Shanghai University; Department of Obstetrics and Gynecology, Second Affiliated Hospital of Mudanjiang Medical University; Research Department, Mudanjiang Medical University
  • Online:2022-01-20 Published:2022-03-02

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摘要： 目的 构建偶联CD133核酸适体载紫杉醇的聚乳酸-乙醇酸-聚乙二醇（PLGA-PEG）纳米载体（N-Pac-CD133）拟清除肺癌干细胞。方法 采用乳液/溶剂蒸发的方法制备 N-Pac-CD133，同时对 N-Pac-CD133 进行表征，利用磁珠分离法分离出CD133+ 肺癌干细胞后并对该群体的肺癌干细胞特异性进行检测，同时对肺癌细胞的靶向性和杀伤活性进行检测。小鼠体内接种A549肿瘤后，肿瘤治疗分组：生理盐水，空纳米载体链接CD133核酸适体（N-CD133），紫杉醇，负载紫杉醇的纳米载体（N-Pac）和N-Pac-CD133，8只/组，5 mg/kg紫杉醇，分别于第10、15和20天进行注射。在第40天时，处死小鼠后，对肿瘤进行摘除并称重，同时测量小鼠的体质量。结果 N-Pac-CD133的粒径为100 nm左右，包封率>80%，载药量>8%，在48 h内都显示出持续的药物释放。肺癌细胞的CD133+细胞群体表现出肺癌干细胞的特征：更快的肺癌生长速度（30 d，P=0.001）和更高的肿瘤干细胞基因表达：OV6（P<0.001）、CD133（P=0.001）、OCT3/4（P=0.002）、EpCAM（P=0.04）、NANOG（P=0.005）和 CD44（P=0.02）。与非靶向 N-Pac 和紫杉醇相比，N-Pac-CD133 对肺癌干细胞的靶向性（P<0.001）和细胞毒性作用显著增强。另外，N-Pac-CD133可显著减少肿瘤球的形成（P<0.001）。在治疗终点时，取小鼠肿瘤并对肿瘤进行称量，N-Pac-CD133治疗组和其他治疗组相比，肿瘤体质量显著减小（P<0.001）。结论 CD133核酸适体可以促进紫杉醇纳米载体靶向递送至CD133+肺癌干细胞并杀伤肺癌干细胞。N-Pac-CD133可能是一种有效的靶向肺癌干细胞的治疗手段。

关键词: 纳米载体；肺癌；紫杉醇；肿瘤干细胞；CD133

Abstract: Objective To construct a polylactic acid-glycolic acid-polyethylene glycol (PLGA-PEG) nanocarrier (N-Pac-CD133) coupled with a CD133 nucleic acid aptamer carrying paclitaxel for eliminating lung cancer stem cells (CSCs). Methods Paclitaxel-loaded N-Pac-CD133 was prepared using the emulsion/solvent evaporation method and characterized. CD133+ lung CSCs were separated by magnetic bead separation and identified for their biological behaviors and gene expression profile. The efficiency of paclitaxel-loaded N-Pac-CD133 for targeted killing of lung cancer cells was assessed in vitro. SCID mice were inoculated with A549 cells and received injections of normal saline, empty nanocarrier linked with CD133 aptamer (N-CD133),paclitaxel, paclitaxel-loaded nanocarrier (N-Pac) or paclitaxel-loaded N-Pac-CD133 (n=8, 5 mg/kg paclitaxel) on days 10, 15 and 20, and the tumor weight and body weight of the mice were measured on day 40. Results Paclitaxel-loaded N-Pac-CD133 showed a particle size of about 100 nm with a high encapsulation efficiency (>80%) and drug loading rate (>8%), and was capable of sustained drug release within 48 h. The CD133+ cell population in lung cancer cells showed the characteristic features of lung CSCs, including faster growth rate (30 days,P=0.001) and high expressions of tumor stem cell markers OV6(P< 0.001), CD133 (P=0.001), OCT3/4 (P=0.002), EpCAM (P=0.04), NANOG (P=0.005) and CD44 (P=0.02). Compared with N-Pac and free paclitaxel, paclitaxel-loaded N-Pac-CD133 showed significantly enhanced targeting ability and cytotoxicity against lung CSCs in vitro (P<0.001) and significantly reduced the formation of tumor spheres (P<0.001). In the tumor-bearing mice, paclitaxel-loaded N-Pac-CD133showed the strongest effects in reducing the tumor mass among all the treatments (P<0.001). Conclusion CD133 aptamer can promote targeted delivery of paclitaxel to allow targeted killing of CD133+ lung CSCs. N-Pac-CD133 loaded with paclitaxel may provide an effective treatment for lung cancer by targeting the lung cancer stem cells.

Key words: nanoparticles; lung cancer; paclitaxel; cancer stem cells; CD133