Nonviral cytokine gene therapy on an orthotopic bladder cancer model

Purpose: The purpose is to assess cytokine gene transfection in tumor cells and its therapeutic efficacy in an orthotopic mouse bladder cancer model after liposome-mediated gene transfer. Experimental Design: A total of 1 x 10(5) MB49 cells was instilled into the bladder of C57BL/6 mice after electrocautery to establish the tumor model. The plasmids were constructed by inserting the coding sequences for murine IFN-alpha1 and granulocyte macrophage colony-stimulating factor into a plasmid vector pBudCE4.1. Transient transfection was performed using a cationic lipid N-[1-(2,3-dioleoyloxyl)propyl]-N,N,N-trimethylammoniummethyl sulfate and methyl-beta-cyclodextrin-solubilized cholesterol. The in vitro expression of cytokines was checked by ELISA. The expression of the transgene in situ was confirmed by immunohistochemistry and 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside staining. Mice bearing orthotopic tumors were treated with plasmid DNA/liposome complex by intravesical instillation twice a week for 3 weeks. Results: Superficial bladder tumors were established by intravesical instillation of MB49 into cauterized bladders. The expression level of cytokines in transfected cell lines was increased significantly. In situ gene transfer to bladder tumors was accomplished via intravesical instillation of plasmid DNA/N-[1-(2,3-dioleoyloxyl)p ropyl]-N,N,N-trimethylammoniummethyl sulfate/methyl-beta-cyclodextrin-solubilized cholesterol after a single 2 h in situ transfection. The tumor incidence in the treatment groups was dramatically decreased from 76.9% in the control group to 15.4-30.8% in the treatment groups. Conclusions: We demonstrated in the orthotopic mouse bladder cancer model that successful inhibition of tumor cell growth could be obtained with cytokine gene therapy. The results suggest that our liposome transfection system appears to be a promising method for gene therapy of bladder cancer in vivo.