volasertib and Bile-Duct-Neoplasms

To examine the influence on apoptotic mechanisms following inhibition of polo-like kinases as therapeutically approach for cholangiocellular cancer treatment.. As most cholangiocarcinomas are chemotherapy-resistant due to mechanisms preventing tumor cell death, we investigated the effect of Cisplatin on cholangiocellular carcinoma (CCA) cell lines KMCH-1 and Mz-Ch-1. Polo-like kinases (PLK) are important regulators of the cell cycle and their inhibition is discussed as a potential therapy while PLK inhibition can regulate apoptotic mediators. Here, cells were treated with PLK inhibitor BI6727 (Volasertib), Cisplatin, and in combination of both compounds. Cell viability was assessed by MTT; apoptosis was measured by DAPI staining and caspase-3/-7 assay. Western blot and qRT-PCR were used to measure expression levels of apoptosis-related molecules Bax and Bcl-2.. The cell viability in the CCA cell lines KMCH-1 and Mz-Ch-1 was reduced in all treatment conditions compared to vehicle-treated cells. Co-treatment with BI6727 and cisplatin could even enhance the cytotoxic effect of cisplatin single treatment. Thus, co-treatment of cisplatin with BI6727 could slightly enhance the cytotoxic effect of the cisplatin in both cell lines whereas there was evidence of increased apoptosis induction solely in Mz-Ch-1 as compared to KMCH-1. Moreover, PLK inhibition decreases protein levels of Bcl-2; an effect that can be reversed by the proteasomal degradation inhibitor MG-132. In contrast, protein levels of Bax were not found to be altered by PLK inhibition. These findings indicate that cytotoxic effects of Cisplatin in Mz-Ch-1 cells can be enhanced by cotreatment with BI6727.. In conclusion, BI6727 treatment can sensitize CCA cells to cisplatin-induced apoptosis with proteasomal Bcl-2 degradation as an additional pro-apoptotic effect.

Topics: Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Bile Duct Neoplasms; Cell Cycle Proteins; Cell Line, Tumor; Cell Survival; Cholangiocarcinoma; Cisplatin; Humans; Polo-Like Kinase 1; Proteasome Endopeptidase Complex; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proteolysis; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Pteridines; Signal Transduction

Cholangiocarcinoma (CCA) cells paradoxically express the death ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and thus rely on potent survival signals to circumvent cell death by TRAIL. Hedgehog (Hh) signaling is an important survival pathway in CCA. Herein, we further examine the mechanisms whereby Hh signaling mediates apoptosis resistance in CCA, revealing a pivotal role for the cell division regulating serine/threonine kinase polo-like kinase 2 (PLK2). We employed 50 human CCA samples (25 intrahepatic and 25 extrahepatic CCA) as well as human KMCH-1, Mz-CHA-1, and HUCCT-1 CCA cells for these studies. In vivo experiments were conducted using a syngeneic rat orthotopic CCA model. In human samples, polo-like kinase (PLK)1/2/3-immunoreactive cancer cells were present in the preponderance of intra- and extrahepatic CCA specimens. Inhibition of Hh signaling by cyclopamine reduced PLK2, but not PLK1 or PLK3, messenger RNA and protein expression in vehicle-treated and sonic Hh-treated CCA cells, confirming our previous microarray study. PLK2 regulation by Hh signaling appears to be direct, because the Hh transcription factors, glioma-associated oncogene 1 and 2, bind to the PLK2 promotor. Moreover, inhibition of PLK2 by the PLK inhibitor, BI 6727 (volasertib), or PLK2 knockdown was proapoptotic in CCA cells. BI 6727 administration or PLK2 knockdown decreased cellular protein levels of antiapoptotic myeloid cell leukemia 1 (Mcl-1), an effect reversed by the proteasome inhibitor, MG-132. Finally, BI 6727 administration reduced Mcl-1 protein expression in CCA cells, resulting in CCA cell apoptosis and tumor suppression in vivo.. PLK2 appears to be an important mediator of Hh survival signaling. These results suggest PLK inhibitors to be of therapeutic value for treatment of human CCA.

Topics: Animals; Apoptosis; Bile Duct Neoplasms; Bile Ducts, Intrahepatic; Cell Line, Tumor; Cell Survival; Cholangiocarcinoma; Disease Models, Animal; Enzyme Inhibitors; Hedgehog Proteins; Heterografts; Humans; Male; Protein Serine-Threonine Kinases; Pteridines; Rats; Rats, Inbred F344; Signal Transduction; TNF-Related Apoptosis-Inducing Ligand