Zampanolide, a Potent New Microtubule-Stabilizing Agent, Covalently Reacts with the Taxane Luminal Site in Tubulin α,β-Heterodimers and Microtubules

Abstract

Summary Zampanolide and its less active analog dactylolide compete with paclitaxel for binding to microtubules and represent a new class of microtubule-stabilizing agent (MSA). Mass spectrometry demonstrated that the mechanism of action of both compounds involved covalent binding to β-tubulin at residues N228 and H229 in the taxane site of the microtubule. Alkylation of N228 and H229 was also detected in α,β-tubulin dimers. However, unlike cyclostreptin, the other known MSA that alkylates β-tubulin, zampanolide was a strong MSA. Modeling the structure of the adducts, using the NMR-derived dactylolide conformation, indicated that the stabilizing activity of zampanolide is likely due to interactions with the M-loop. Our results strongly support the existence of the luminal taxane site of microtubules in tubulin dimers and that microtubule nucleation induction by MSAs may proceed through an allosteric mechanism.

SKLB060 Reversibly Binds to Colchicine Site of Tubulin and Possesses Efficacy in Multidrug-Resistant Cell Lines

“…Although MTIs have been demonstrated to exert a high level of anti-cancer activities during clinical treatment or preclinical testing, their effectiveness is limited by multidrug resistance (MDR) [6,7]. To avoid MDR, researchers once focused on the development of irreversible tubulin inhibitors [8,9], because resistant tumor cells cannot escape the effects of irreversibly bound compounds by reducing their affinity for the target or by enhancing drug efflux. However, these irreversible compounds that bind to tubulin usually result in severe toxicity, and thus their clinical use is limited [10].…”

Recent progress with microtubule stabilizers: new compounds, binding modes and cellular activities

“…Interestingly, one analogue that is devoid of the entire tetrahydropyran ring, yet maintains the hemiaminal side chain retains biological activity. 82 The ability of zampanolide to bind covalently to microtubules was identified in a collaborative effort led by Fernando Diaz 83 and is described in detail below in comparison with other microtubule stabilizers and their interactions with tubulin binding sites.…”

“…Compounds like the epothilones, taxanes and discodermolide, which do not bind covalently, bind much tighter to formed microtubules than they do to unassembled tubulin. 83 The higher energy of binding to the formed microtubule shifts the equilibrium towards microtubule polymerization independent of any structural effects caused by the drug on the polymerized microtubule. On the other hand, compounds that bind covalently to microtubules displace this equilibrium through a structural allosteric effect in which the modified tubulin has a higher affinity towards polymerized microtubules than the unmodified fraction of tubulin.…”

“…On the other hand, compounds that bind covalently to microtubules displace this equilibrium through a structural allosteric effect in which the modified tubulin has a higher affinity towards polymerized microtubules than the unmodified fraction of tubulin. 83 …”

Epothilones: From Discovery to Clinical Trials

“…Beside taxanes and epothilones, microtubule stabilizers include discodermolide, 29 eleutherobin 30 and sarcodic-tyins, 31 laulimalide, 32 peloruside 33 and zampanolide. 34 The most prominent microtubule destabilizing agents are colchicine, 35 combretastin A-4, 36 podophyllotoxin, 37 vinblastine 38 and other vinca alkaloids. 39 …”