1,25D3 Enhances antitumor activity of gemcitabine and cisplatin in human bladder cancer models

Abstract

BACKGROUND: 1,25 dihydroxyvitamin D₃ (1,25D₃) 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 1,25D₃ 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 1,25D₃ 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: 1,25D₃ pretreatment enhanced gemcitabine and cisplatin‐induced apoptosis and the activities of caspases 8, 9, and 3 in T24 and UMUC3 cells. 1,25D₃ synergistically reduced gemcitabine and cisplatin‐suppressed surviving fraction in T24 cells. 1,25D₃, gemcitabine, or cisplatin induced p73 accumulation, which was enhanced by gemcitabine and cisplatin or 1,25D₃ 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 1,25D₃, gemcitabine and cisplatin, or 1,25D₃ and gemcitabine and cisplatin. 1,25D₃ and gemcitabine and cisplatin combination enhanced tumor regression compared with 1,25D₃ or gemcitabine and cisplatin alone. CONCLUSIONS: 1,25D₃ 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. © 2010 American Cancer Society.