目的：建立p[5HRE]AFPp-p53/PEI-Fe3O4磁性纳米颗粒靶向基因治疗和磁流体热疗系统用于肝癌的联合治疗，以提高治疗的安全性和有效性。方法：亚克隆基因重组法构建靶向肝癌的治疗基因p[5HRE]AFPp-p53，并用限制性内切酶凝胶电泳法检测重组质粒是否构建成功；用共沉淀法制备磁性纳米颗粒PEI-Fe3O4，利用透射电镜、粒径仪、傅里叶转换红外光谱仪等对PEI-Fe3O4进行表征检测。MTT法检测PEI-Fe3O4转染p[HRE]AFPp-p53至不同细胞系后的细胞增殖及磁流体热疗和基因治疗联合作用对肝癌细胞HepG2的增值抑制作用。结果：成功制备载有靶向治疗基因的p[5HRE]AFPp-p53/PEI-Fe3O4磁性纳米颗粒，由其介导的基因治疗组与阴性及纳米颗粒对照组相比，明显抑制肝癌HepG2（AFP阳性）细胞 [(0592±0041) vs (1.052±0.031)、(1.012±0.021)，P<0.01]和SMMC7721（AFP阴性）细胞（0.813±0.042） vs（1.073±0.032）、（1.182±0.052），P<0.01]的增殖活性，但对非肝癌细胞（L929和Lovo) 增殖抑制作用无明显影响（P>0.05）。基因治疗和磁流体热疗联合组与单独使用热疗组及基因治疗组相比显著增加HepG2细胞的增殖抑制率（76.11% vs 35.22%、4292%,均P<001）。结论：p[5HRE]AFPp-p53 PEI-Fe3O4 磁性纳米颗粒对肝癌细胞具有特异性杀伤作用，并且能与磁流体热疗产生协同效应，是一种选择性高、治疗效果好的肿瘤治疗方法。

Objective:To establish a combined therapy of p[5HRE]AFPp-p53/PEI-Fe3O4 magnetic nanoparticle and Magnetic Fluid Hyperthermia (MFH) to treat liver carcinoma and improve the safety and efficacy of the treatment. Methods:Sub-clone was used to construct hepatoma targeted gene p[5HRE]AFPp-p53, which was then confirmed by enzyme digestion and gel electrophoresis. The PEI-Fe3O4 magnetic nanoparticles were prepared by coprecipitation method, and its surface characteristics were examined by transmission electron microscope, particle size analyzer and Fourier transform infrared spectrometer etc. The proliferation of cell lines transfected with p[HRE]AFPp-p53/ PEI-Fe3O4, and the inhibition effect of targeted gene therapy combined with MFH on HepG2 cell proliferation were detected by MTT method. Results: p[5HRE]AFPp-p53/PEI-Fe3O4 magnetic nanoparticles were successfully constructed. Comparing with negative control group and nanoparticles control group, the proliferation activity of HepG2 cells ([0.592±0.041] vs [1.052±0031]，[1.012±0.021], P<0.01) and SMMC772 cells ([0.813±0.042] vs [1.073±0.032], [1.182±0.052], P<0.01) was significantly inhibited in p[5HRE]AFPp-p53/PEI-Fe3O4 mediated gene treatment group, however the proliferation of nonhepatoma cells (L929 and LOVO) was not significantly inhibited (P>0.05). Comparing with MFH group and gene therapy group, the inhibition rate of HegG2 cell proliferation was significantly increased in gene+MFH group (3522%, 42.92% vs 76.11%, P<0.01).Conclusion: p[5HRE]AFPp-p53/PEI-Fe3O4 magnetic nanoparticle has specific killing effect on liver carcinoma cells; it will have synergistic effect when combined with MFH; the combined therapy is an anti-tumor treatment with high selection and good therapeutic effect.

国家自然科学基金资助项目（No.81271636）