Evaluation of gene transfer efficiency by viral vectors to murine bladder epithelium

Purpose: In pre-clinical gene therapy studies of bladder cancer there is tr emendous variation in the ability of viral vectors to deliver genetic mater ial to bladder epithelium. Possible explanations for this variability may i nvolve the physical parameters of delivering vectors in these experimental models. We examined the effects of intravesical volume and pressure during instillation as well as chemical modification of the bladder epithelium on subsequent gene expression in the bladder in mice. Materials and Methods: Female C57B1/6 mice underwent intravesical instillat ion of the replication restricted canarypox virus (ALVAC) recombinant for t he reporter genes luciferase or p-galactosidase. Similar viral titers were instilled at different volumes and a pressure transducer measured intravesi cal pressure when the vector was instilled. Also, various agents, including 0.6 N hydrochloric acid, 0.4% oxychlorosene, poly-l-lysine and 0.25 M. amm onium chloride, were used to modify the bladder surface before vector insti llation and then assayed for transgene expression. Results: As expected, maximum intravesical pressure measured during instill ation was significantly greater in mice instilled with a higher volume (33. 1 versus 9.8 mm. Hg). Significantly more gene expression was detected in bl adders instilled with a higher volume of viral vectors (p <0.05). Likewise, higher instillation pressures resulted in higher transgene expression in d istant organs. Modification of the bladder epithelium with agents such as o xychlorosene and poly-L-lysine resulted in elevated gene expression with on ly minimal increases in systemic activity. Conclusions: Significant differences in gene expression are achieved by var ying physical parameters during intravesical instillation. Increased gene e xpression associated with larger volume instillation may be responsible for some reported variability of gene transfer to the bladder. Alternate manip ulations, such as modifying the bladder surface, may be done to enhance gen e transfer to the urothelium without increasing systemic distribution.