In vitro evaluation of calphostin C as a novel agent for photodynamic therapy of bladder cancer.

摘要

OBJECTIVES: Calphostin C, a highly specific protein kinase C inhibitor, induces apoptosis in the presence of visible light. We report the photoactivatable cytotoxicity of calphostin C in a series of well-characterized human bladder cancer cell lines: RT4, UM-UC-3, and 5637. METHODS: The human bladder cancer cell lines RT4, UM-UC-3, and 5637 were chosen on the basis of their p53, pRb and 9p21 deletion status. Using standard tissue culture techniques, the cytotoxicity of 10 to 100 nM calphostin C in combination with increasing exposures of visible light was examined. Controls consisted of cells treated with calphostin C without visible light and cells exposed to visible light without calphostin C treatment. Cell viability was determined by MTT assay. The induction of apoptosis by activated calphostin C was determined by 4,6-diamidino-2-phenylindole (DAPI) staining/fluorescence microscopy of nuclei. RESULTS: In the absence of light, calphostin C did not demonstrate a cytotoxic effect on any of the cell lines tested. Increasing the duration of light exposure resulted in a concomitant decrease in cell viability. Significant cell death was seen with calphostin C concentrations as low as 10 nM. These studies also demonstrated that calphostin C induced apoptosis by a mechanism independent of p53 and pRb status and the presence or absence of 9p21 deletions. CONCLUSIONS: We demonstrated the ability of activated calphostin C to induce apoptosis in a light-dependent and concentration-dependent fashion in a bladder cancer model system. Activated calphostin C cytotoxicity is independent of tumor genetic background and the status of p53 and pRb. Further development of calphostin C as a photosensitizer for photodynamic therapy of superficial bladder cancer may be warranted.