Trisoxazole Macrolides Potentiate the Microtubule Assembly and Antimitotic Activities of Taxanes

Mycalolides are marine-derived trisoxazole macrolides that exhibit potent actin-depolymerizing activity and inhibit the proliferation, migration, and invasion of various cancer cells. The C25-C35 side-chain moiety of mycalolides is important for their actin-binding properties; however, the biological function of the C1-C24 trisoxazole macrolactone moiety remains largely unexplored. In this study, using a photoaffinity biotin probe, we identified tubulin as a new, actin-independent target of mycalolide C (MyC). Ultracentrifugation, fluorescence-based tubulin polymerization assays, and transmission electron microscopy of negatively stained microtubules (MTs) demonstrated that MyC and trisoxazole macrolactones exert potent synergistic effects on paclitaxel-induced MT assembly and stabilization. These compounds enhanced the antiproliferative activity of paclitaxel by 4.2- to 6.7-fold in HCT-116 human colon cancer cells but showed little synergy in 3T3-L1 murine fibroblast. Molecular modeling studies suggest that MyC stabilizes the protein-protein interactions between α/β-tubulin heterodimers. Given that the C1-C24 trisoxazole macrolactone moiety and its analogues exhibit little cytotoxicity, our findings may contribute to the development of new lead compounds for MT inhibitors with reduced side effects.