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The present study investigated the potential of vascular endothelial growthfactor (VEGF) promoter to derive cytosine deaminase (CD) transfected bypolyamidoamine (G4-PAMAM) dendrimers to 4T1 murine breast cancer cell line asgene-directed enzyme/prodrug therapy. The VEGF promoter and cytosinedeaminase gene were cloned into the pEGFP-N1vector from the genomic DNA of 4T1and E. coli, respectively. The frequency of transfection forVEGF-CD-pEGFP-N1 and pEGFP-N1- CD treated groups was 35±3 and36±4, respectively. MTT assay was perform to evaluate the cytotoxiceffects of converted 5-flurocytosine on 4T1 cells. Also, the optimalconcentration of 5-FC in 4T1 cells transfected by VEGF-CD-pEGFP-N1 plasmid wasevaluated. The GFP expression of transfected 4T1 cells by VEGF-CD-pEGFP-N1wereobserved by fluorescent microscopy and flowcytometry. Results demonstrated thatthe suicide CD gene was successfully expressed in 4T1 cells determined by RT-PCRand GFP expression. A concentration of 200 μg/ml 5-FC wasidentified as optimal dose of prodrug. Furthermore, the CD/5-FCenzyme/prodrug system not only demonstrated toxicity on transformed 4T1cells but also exerted a ‘bystander effect’ determined by MTTassay. The results showed that by 35% transfection withVEGF-CD–pEGFP-N1and CD-pEGFP-N1 plasmids, 80% and 90%inhibition of the cells growth occurred, respectively.

drug research

Non-viral Suicide Gene Therapy: Cytosine Deaminase Gene Directed by VEGF Promoter and 5-fluorocytosine as a Gene Directed Enzyme/prodrug System in Breast Cancer Model