Investigation of anti-tumor mechanisms of K2154: characterization of tubulin isotypes, mitotic arrest and apoptotic machinery.

Abstract

Microtubules are crucial targets for cancer chemotherapeutic drugs, and new microtubule-directed agents are of continued interest in drug development. A novel microtubule-directed agent, ethyl-2-[N-rho-chlorobenzyl-(2'-methoxy)]-anilino-4-oxo -4, 5-dihydro-furan-3-carboxylate, was identified. The compound, designated K2154, inhibited cell proliferation, with IC(50) values of 10.3, 15.3, 9.6, 11.2, 12.8 and 12.1 muM in prostate cancer PC-3, hepatocellular carcinoma Hep3B, non-small cell lung cancer A549, colorectal cancer HT29 and HCT116, and P-glycoprotein-rich breast cancer NCI/ADR-RES cells, respectively. Because NCI/ADR-RES cells were susceptible to inhibition by K2154, it indicated that this compound is a poor substrate for P-glycoprotein. In this study, PC-3 cells were used to identify the anticancer mechanisms of K2154. K2154 induced an arrest of the cell cycle at G2/M phase and a subsequent increase of hypodiploid phase in PC-3 cells, whereas it only induced a moderate level of G2/M arrest with little increase of hypodiploid phase in normal prostate cells. K2154 inhibited microtubule assembly in both in vitro turbidity assay and in vivo microtubule spin-down experiment. Immunochemical examination showed that K2154 caused formation of abnormal mitotic characteristics with bipolar spindles, particularly, in beta(II)- and beta(III)-tubulin staining. It also induced several pathways, including cyclin B1 up-regulation, dephosphorylation on Tyr(15) and phosphorylation on Thr(161) of Cdk1 and Cdc25C phosphorylation, and roscovitine (a Cdk1 inhibitor) significantly inhibited K2154-induced apoptosis, suggesting a pro-apoptotic role of Cdk1. Phosphorylation of Bcl-2 and Bcl-xL and cleavage of Mcl-1, together with activation of caspase-9 and -3, indicated that mitochondrial pathway played a central role in K2154-mediated apoptotic cell death. Additionally, AIF contributed to a late phase of K2154-induced apoptotic pathway. In conclusion, it is suggested that K2154 displays an anticancer activity through a target on microtubules and a subsequent signaling cascade on cell cycle regulation and apoptotic machinery.

• Authors: Pin-Hsuan Lu, Fan-Lu Kung, Sheng-Chu Kuo, Shih-Chieh Chueh, Jih-Hwa Guh
• Journal: Naunyn-Schmiedeberg's archives of pharmacology (Naunyn Schmiedebergs Arch Pharmacol) Vol. 374 Issue 3 Pg. 223-33 (Dec 2006) ISSN: 0028-1298 [Print] Germany
• PMID: 17102938 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• ATP Binding Cassette Transporter, Subfamily B, Member 1
• Aniline Compounds
• Antineoplastic Agents
• Apoptosis Inducing Factor
• CCNB1 protein, human
• Cyclin B
• Cyclin B1
• Furans
• Tubulin
• ethyl 2-(N-p-chlorobenzyl-(2'-methoxy))anilino-4-oxo-4,5-dihydrofuran-3-carboxylate
• Cytochromes c
• Caspases

Topics

• ATP Binding Cassette Transporter, Subfamily B, Member 1
• Aniline Compounds (pharmacology)
• Antineoplastic Agents (pharmacology)
• Apoptosis (drug effects)
• Apoptosis Inducing Factor (metabolism)
• Caspases (drug effects, metabolism)
• Cell Cycle (drug effects)
• Cell Line, Tumor
• Cell Proliferation (drug effects)
• Cyclin B (drug effects, metabolism)
• Cyclin B1
• Cytochromes c (metabolism)
• Drug Delivery Systems
• Furans (pharmacology)
• Humans
• Inhibitory Concentration 50
• Microtubules (drug effects)
• Mitochondria (metabolism)
• Mitosis (drug effects)
• Phosphorylation
• Tubulin (drug effects, metabolism)
• Up-Regulation