Egr1/HSP70 Promoter-Driven Activation of Gene Expression for Synergistic Anti-Hepatoma Using PEI-MZF Nanoparticles and Radiation

Abstract

Jia Zhang,1,2 Min Dong,1 Yan Feng,1 Dongsheng Zhang,2 Mei Lin,2 Chenyan Yuan,2 Hongbo Li,2 Ling Wang,2 Hao Zhang,2 Chen Liang2 1Nanjing Medical University Affiliated Cancer Hospital & Jiangsu Cancer Hospital & Jiangsu Institute of Cancer Research, Nanjing, Jiangsu, People’s Republic of China; 2Imaging and Nuclear Medicine, Southeast University, Nanjing, Jiangsu, People’s Republic of ChinaCorrespondence: Yan FengNanjing Medical University Affiliated Cancer Hospital, No. 42 Baiziting Road, Nanjing, 210009, Jiangsu, People’s Republic of ChinaTel [phone]Email [email protected] ZhangImaging and Nuclear Medicine, Southeast University, No. 87 Dingjiaqiao Road, Nanjing, 210000, Jiangsu, People’s Republic of ChinaTel [phone]Email [email protected]: Spatially restricted gene expression circumvents the gene expression and gene vector problem by enabling localized amplification. The objective of this study is to construct a spatially restricted gene expression for liver cancer therapy based upon the MFH-absorbing properties of PEI- Mn0.5zn0.5Fe2o4, gene therapy and radiation.Methods: Mn0.5zn0.5Fe2O4 (MZF) magnetic nanoparticles were prepared by an improved chemical co-precipitation method, modified by polyethylene imine (PEI), and then the structure, modification characters, biocompatibility, temperature rise and control ability and binding efficiency of the plasmid were characterized. Then, the dual-promoter plasmid PCDNA3.1-EGR1-HSP70-HSVTK was constructed. The recombinant vectors were identified by enzyme digestion analysis and DNA sequencing. The TK gene expression level was detected by realtime-PCR assay in HEK293 cells. Also, the HSV-TK gene expression was detected in SMMC7721 cells with the help of PEI-Mn0.5Zn0.5Fe2O4. In vitro anti-tumor experiment, MTT assay and flow cytometry were used to evaluate the therapeutic effects of the cultured SMMC7721 cells treated by different ways. In vivo anti-tumor experiment, the xenografted mice were treated by different ways for three times to detect the antitumor effect.Results: The Mn0.5Zn0.5Fe2O4 magnetic nanoparticles could be successfully prepared through improved co-precipitation process and showed good biocompatibility. And PEI had been coated on MZF complex. The modified PEI-MZF presented favorable dispensability, responsibility to magnetism, good loading capability and transfect capability. Also, pCDNA3.1-Egr1-Hsp70-HSVTK plasmid had been constructed successfully and could be induced by heat and irradiation. It would be used for further target gene therapy research. The antitumor results in vitro showed: The therapeutic effects of nanosized PEI-MZF-HSV-TK complex could significantly inhibit the proliferation of cultured liver cancer cells (SMC7721), induce cell apoptosis and had a prominent cell cycle disturbance in the S phase in vitro. The results in vivo showed: The combined therapy induced by PEI-MZF-HSV-TK could inhabit the growth of hepatocellular carcinoma xenografts by killing and inhabiting the proliferation of the tumor cells.Conclusion: The novel site-directed heat/radiation-inducible expression system based upon the hyperthermia (by MFH) and radiation possessed superior antitumor effect in vivo and in vitro.Keywords: radiation, magnet-induced heating, dual sensitive, magnetic nano vector

Keywords

• radiation
• magnet-induced heating
• dual sensitive
• magnetic nano vector