1,4-Diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126) Enhances the Cytotoxicity of Combretastatin A4 Independently of Mitogen-Activated Protein Kinase Kinase

Combretastatin A4 (CA4) is a novel vascular-disrupting agent that has shown promising anticancer effects through its inhibition of microtubule assembly and subsequent disruption of tumor blood flow. In this report, we demonstrate that 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126), a selective inhibitor of mitogen-activated protein kinase kinase (MEK), significantly enhances the cytotoxicity of CA4 in BEL-7402 cells, independently of MEK inhibition. This independence is evidenced by the fact that another, more specific MEK inhibitor, PD0325901 [N-[(R)-2,3-dihydroxy-propoxy]-3,4-difluoro-2-[2-fluoro-4-iodo-phenylamino]-benzamide], does not have the same effect as U0126. The disassembled microtubules are able to reassemble in the later stages of CA4 treatment, because of the inactivating glucuronidation of CA4. U0126, but not PD0325901, inhibits CA4 glucuronidation, thereby blocking microtubule reassembly and enhancing CA4-induced G(2)/M cell-cycle arrest. Consistent with this, U0126 significantly enhances CA4-induced cytotoxicity for cells in which CA4 glucuronidation occurs, but not for cells in which such glucuronidation does not occur. These results suggest that great caution should be exercised when interpreting data obtained using U0126 or when CA4 is combined with inhibitors of glucuronidation in clinical practice. It is most important to note that these findings indicate that the combination of CA4 with inhibitors of glucuronidation may be a novel and rational strategy for cancer therapy.