Abstract
Chemotherapy to date has not been effective in the treatment of cholangiocarcinoma. However, gemcitabine, a novel nucleoside analog, has shown remarkable biological activity against cholangiocarcinoma in some clinical studies. Combinations of gemcitabine with other agents have also shown promising results, with a tolerable toxicity profile. Nutlin-3 is a small-molecule inhibitor that acts to inhibit murine double minute-2 (MDM2) binding to p53 or p73 and subsequently activates p53- or p73-dependent apoptosis signaling pathways. To investigate their effects in combination, a p53-mutant cholangiocarcinoma line HuCCT1 was treated with Nutlin-3 and/or gemcitabine in the current study. Cell proliferation assay, apoptosis assay, Western blot, coimmunoprecipitation, and small interfering RNA (siRNA) experiments were analyzed in HuCCT1 cells. Antitumoral effects of Nutlin-3 and/or gemcitabine targeting the p73/MDM2 pathways were evaluated. Nutlin-3 and gemcitabine can both inhibit the growth of HuCCT1 cells. Nutlin-3 induces apoptosis and potentiates the cytotoxic effect of gemcitabine through disrupting the binding of p73 with MDM2. Nutlin-3 leads to activation of caspases, increase levels of puma and bax, and decrease the expression of bcl-2. Blocking p73 function with a siRNA attenuates the apoptosis induced by gemcitabine, Nutlin-3, and gemcitabine/Nutlin-3 combination. Our data provide evidence that p73 might compensate for p53 function in gemcitabine-induced apoptosis of HuCCT1 cells. Nutlin-3 acts through the inhibition of p73-MDM2 with subsequent activation of the apoptotic pathway signaling, which leads to the increase in chemosensitivity to gemcitabine. In summary, our findings suggest that Nutlin-3 will be active in the treatment of p53-mutant cholangiocarcinoma, especially when in combination with gemcitabine.
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Abbreviations
- (MDM2):
-
murine double minute-2
- (PBS):
-
phospho-buffered saline
- (siRNA):
-
small interfering RNA
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The authors thank Tohoku University for providing the HuCCT1 cell line and Dr. Nishant Bhatta for reviewing the manuscript.
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Tongsen Zheng and Jiabei Wang contributed equally to this work.
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Zheng, T., Wang, J., Chen, X. et al. Disruption of p73-MDM2 binding synergizes with gemcitabine to induce apoptosis in HuCCT1 cholangiocarcinoma cell line with p53 mutation. Tumor Biol. 31, 287–295 (2010). https://doi.org/10.1007/s13277-010-0035-7
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DOI: https://doi.org/10.1007/s13277-010-0035-7