1,25D3 Enhances Antitumor Activity of Gemcitabine and Cisplatin in Human Bladder Cancer Models

BACKGROUND: 1,25 dihydroxyvitamin D-3 (1,25D(3)) potentiates the cytotoxic effects of several common chemotherapeutic agents. The combination of gemcitabine and cisplatin is a current standard chemotherapy regimen for bladder cancer. The authors investigated whether 1,25D(3) could enhance the antitumor activity of gemcitabine and cisplatin in bladder cancer model systems. METHODS: Human bladder cancer T24 and UMUC3 cells were pretreated with 1,25D(3) followed by gemcitabine and cisplatin. Apoptosis was assessed by annexin V staining. Caspase activation was examined by immunoblot analysis and substrate-based caspase activity assay. The cytotoxic effects were examined by using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and in vitro clonogenic assay. p73 protein levels were assessed by immunoblot analysis. Knockdown of p73 was achieved by siRNA. The in vivo antitumor activity was assessed by in vivo excision clonogenic assay and tumor regrowth delay in the T24 xenograft model. RESULTS: 1,25D(3) pretreatment enhanced gemcitabine and cisplatin-induced apoptosis and the activities of caspases 8, 9, and 3 in T24 and UMUC3 cells. 1,25D(3) synergistically reduced gemcitabine and cisplatin-suppressed surviving fraction in T24 cells. 1,25D(3), gemcitabine, or cisplatin induced p73 accumulation, which was enhanced by gemcitabine and cisplatin or 1,25D(3) and gemcitabine and cisplatin. p73 expression was lower in human primary bladder tumor tissue compared with adjacent normal tissue. Knockdown of p73 increased clonogenic capacity of T24 cells treated with 1,25D(3), gemcitabine and cisplatin, or 1,25D(3) and gemcitabine and cisplatin. 1,25D(3) and gemcitabine and cisplatin combination enhanced tumor regression compared with 1,25D(3) or gemcitabine and cisplatin alone. CONCLUSIONS: 1,25D(3) potentiates gemcitabine and cisplatin-mediated growth inhibition in human bladder cancer models in vitro and in vivo, which involves p73 induction and apoptosis. Cancer 2010;116:3294-303. (C) 2010 American Cancer Society