naphthoquinones and Cholangiocarcinoma

Pyruvate kinase M2 (PKM2) is an enzyme that is predominantly overexpressed in various types of cancer. The role of PKM2 in liver fluke-associated cholangiocarcinoma (CCA) remains unclear. This study aimed to investigate the antitumor activity of shikonin, a PKM2 inhibitor, in CCA cells.. Immunohistochemistry and immunoblotting were used to determine PKM2 expression in CCA tissues and cells. Antiproliferative effects of shikonin were evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, colony-formation and trypan blue exclusion assays. The anti-metastatic activity of shikonin was determined using the Boyden chamber assay. Mechanisms by which shikonin inhibited CCA progression were determined.. PKM2 was overexpressed in CCA compared to normal bile duct epithelial cells. Shikonin significantly inhibited growth, and migration of CCA cells while inducing their death. A mechanistic study revealed that antitumor effects of shikonin in CCA cells depended on increased production of reactive oxygen species.. Shikonin may be a novel therapeutic agent for patients with CCA.

Topics: Antineoplastic Agents; Apoptosis; Bile Duct Neoplasms; Carrier Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cholangiocarcinoma; Humans; Membrane Proteins; Naphthoquinones; Reactive Oxygen Species; Thyroid Hormone-Binding Proteins; Thyroid Hormones

Cholangiocarcinoma is a malignant tumor with high metastatic and mortality rates. We investigated the effects\ of rhinacanthin-C on cell proliferation, migration, invasion and the expression of proteins regulating cancer cell\ invasion-regulated proteins in a cholangiocarcinoma (KKU-M156) cell line. Cytotoxicity of rhinacanthin-C was\ determined by the SRB assay. Using wound-migration, chamber-migration and chamber-invasion assays, we assessed\ the effects of rhinacanthin-C against KKU-M156 cells. The activities of matrix metalloproteinases 2 and 9 (MMP-2,\ MMP-9) and urokinase-type plasminogen activator (uPA) were determined using gelatinase and uPA zymography\ assays. The expression of invasion-regulated proteins was investigated using western-blot analysis. After treatment\ with rhinacanthin-C, KKU-M156 cells exhibited antiproliferative effects in a dose-dependent manner with greater\ efficacy than in Vero cells: IC50 values were 1.50 and 2.37 μM, respectively. Rhinacanthin-C significantly inhibited cell\ migration and invasion of KKU-M156 cells in a dose-dependent manner. Consistent with this observation, treatment\ with rhinacanthin-C was associated with a decrease in the expression levels of FAK, p-FAK, MMP-2, and a decrease in\ the levels of p38-, JNK1/2- and ERK1/2-MAPK pathways as well as inhibiting NF-κB/p65 expression and translocation\ of NF-κB/p65 to the nucleus. We have shown for the first time that the anti-metastatic effects of rhinacanthin-C on\ KKU-M156 cells are mediated via inhibition of the expression of invasion-regulated proteins. Rhinacanthin-C may\ deserve consideration as a potential agent for the treatment of cholangiocarcinoma.

Topics: Acanthaceae; Animals; Antineoplastic Agents; Cell Line; Cell Line, Tumor; Cell Movement; Cell Proliferation; Chlorocebus aethiops; Cholangiocarcinoma; Focal Adhesion Kinase 1; Humans; MAP Kinase Signaling System; Matrix Metalloproteinase 2; Mitogen-Activated Protein Kinases; Naphthoquinones; Neoplasm Invasiveness; NF-kappa B; Plant Extracts; Signal Transduction; Urokinase-Type Plasminogen Activator; Vero Cells

Cholangiocarcinoma (CCA), arising from varying locations within the biliary tree, is the second most common primary liver malignancy worldwide. Shikonin, an active compound extracted from the Chinese herb Zicao, holds anti-bacterial, anti-inflammatory, and anti-tumor activities. However, the effect of shikonin on human cholangiocarcinoma and detailed mechanisms of TRAIL enhancement remains to be elucidated. The purpose of the study was to investigate the protective functions of TRAIL enhancement for shikonin induced apoptosis in cholangiocarcinoma cells.. We use MTT assay, apoptosis assay, caspase activity assay, flow cytometry assay, real time PCR and Western blot to observe the effects of TRAIL on shikonin induced cholangiocarcinoma cells apoptosis and its mechanism.. Shikonin inhibited cell viability and induced apoptosis of CCA cells, effects enhanced by TRAIL treatment via activation of caspase-3, -8, -9. Furhermore, TRAIL enhanced anti-proliferation of shikonin and shikonin induced apoptosis through induction of ROS mediated JNK activation, while AKT activation had an effect on shikonin anti-proliferation activity, but not in the TRAIL enhanced counterparts. Finally, shikonin upregulated DR5 expression, an effect essential for TRAIL-enhanced activities of shikonin in RBE cells.. Our results revealed that shikonin could inhibit cells viability and induce apoptosis of CCA cells, effects enhanced by TRAIL treatment via ROS mediated JNK signalling pathways, involving up-regulation of DR5 expression. Our results provide further insight into the mechanism underlying the anti-tumor effects of shikonin by TRAIL enhanced in CCA and a new therapeutic strategy to CCA treatment.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Bile Duct Neoplasms; Bile Ducts; Cell Line; Cell Line, Tumor; Cholangiocarcinoma; Humans; MAP Kinase Kinase 4; Naphthoquinones; Reactive Oxygen Species; Signal Transduction; TNF-Related Apoptosis-Inducing Ligand