The gene therapy specific to hepatoma was examined by 2-methacryloyloxyethyl phosphorylcholine (MPC) unit as bio-compatible biomaterials, which conjugated to the HBs antigen and sFlt-1 plasmid. <Materials and methods> 1. PMDN was used as a frame of gene transfer vector: PMDN; Polymerization of MPC unit as bio-compatible biomaterials, N,N-dimethylaminoethyl methacrylate (DMAEMA) which are cationic unit combinable with a gene, and p-nitrophenylcarbonyloxyethyl methacrylate (NPMA) which can obtain cell-specificity by ester combination to HBs antigen. A brief preparation of sFlt-1/PMDN with HBs complex is as follows. PBS containing a given amount of PMDN and HBs antigen was mixed softly. PMDN with HBs was purified by dialysis for molecular weight 30000, which removed p-nitrophenol separated by the compound. Moreover, PMDN with HBs and sFlt-1, which inhibit the angiogenic action of VEGF, was dissolved and mixed in PBS. 2. Tumor cells (HepG2; hepatoma, WiDr; adenocarcinoma) were seeded at 1 x 104 cells/well in 6-well plates 24 h prior to transfection. Transfection complex (sFlt-1/PMDN with HBs, PMDN with HBs, and PBS (control)) were diluted to a total amount of 40μg of plasmid. We obtained each supernates on day 1, 3, 5, 7, 10 after transfection and measured the concentration of sFlt-1 by ELISA method. 3. Tumor cells were injected s.c. in the backs of nude mice, which nodule containing 1.0 x 108 HepG2 cells and 1.0 x 106 WiDr cells respectively. From the time of tumor diameter about >0.5cm, we injected the same drugs as in vitro, a total amount of which were 200μg of plamid, administered by i.p. daily for a total 5 doses, and monitored tumor growth for 21 days. Statistical analysis was all calculated by Mann-Whitney’s U-test. The concentration of sFlt-1 on day 7 were 1603.7 +/− 57.7 pg/ml, 105.2 +/− 18.8 pg/ml, 202.8 +/− 38.8 pg/ml (mean +/− SD) in the group of HepG2 respectively transfected by sFlt-1/PMDN with HBs, PMDN with HBs, and PBS. The concentration of sFlt-1 in the group transfected by sFlt-1/PMDN with HBs was significantly higher than in the other group of HepG2 (p<0.05). On the other hand, the concentration of sFlt-1 was low in all groups of WiDr. And the s.c. tumor growth rate of HepG2 on day 12 were 1.65 +/− 0.28, 12.8 +/− 4.45, 9.1 +/− 2.33 (mean +/− SD), in mice respectively administered by sFlt-1/PMDN with HBs, PMDN with HBs, and PBS. These in mice administered by sFlt-1/PMDN with HBs were significantly smaller than the other drugs (p<0.05). There is almost no change of the tumor growth in the group of sFlt-1/PMDN with HBs until day 14 and the tumor size increased gradually after that. On the other hands, WiDr transfected by all drugs showed respectively identical growth rates in vivo. It was suggested that PMDN conjugated with HBs and sFlt-1 could inhibit hepatoma growth selectively and these were a hepatocyte-specific vector.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]