Macromolecular interaction of halichondrin B analogues eribulin (E7389) and ER-076349 with tubulin by analytical ultracentrifugation.
Halichondrin B is an antimitotic drug that inhibits microtubule assembly. To understand the molecular details of its interaction with tubulin, we investigated the binding of two halichondrin B analogues, eribulin (previously, ER-086526, E7389) and ER-076349, to tubulin by quantitative analytical ultracentrifugation. Eribulin is currently undergoing phase III clinical trials for cancer; ER-076349 is a closely related analogue with C.35 hydroxyl instead of C.35 primary amine [Towle, M. J., et al. (2001) Cancer Res. 61, 1013]. Below the critical concentration for microtubule assembly and in the presence of GDP, tubulin undergoes weak self-association into short curved oligomers. Eribulin inhibits this oligomer formation 4-6-fold, while ER-076349 slightly stimulates oligomer formation by 2-fold. This is in contrast to vinblastine which strongly stimulates large spiral polymers by 1000-fold under these same conditions. Vinblastine-induced spiral formation is strongly inhibited by both eribulin and ER-076349. Colchicine binding to the intradimer interface has no significant effect on small oligomer formation or the inhibitory activity of eribulin on this process. These results suggest that halichondrin B analogues bind to the interdimer interface or to the beta-subunit alone, disrupt polymer stability, and compete with vinblastine-induced spiral formation. Stathmin is known to form a tight 1:2 complex with tubulin. Eribulin strongly inhibits formation of the 1:2 stathmin-tubulin complex (>3.3 kcal/mol), while ER-076349 weakens formation of the 1:2 complex by approximately 1.9 kcal/mol. These results suggest that eribulin is a global inhibitor of tubulin polymer formation, disrupting tubulin-tubulin contacts at the interdimer interface. ER-076349 also perturbs tubulin-tubulin contacts, but in a more polymer specific manner, reflecting adaptability of the interdimer interface to drug and polymer polymorphism. These results suggest halichondrin B analogues exhibit unique tubulin-based activities that may underlie the clinical utility of these compounds.
Alday
P Holland
PH
Department of Biochemistry, University of Mississippi Medical Center, Jackson, Mississippi 39216, USA.
Correia
John J
JJ
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000378:metabolism; D001667:Binding, Competitive; D019281:Dimerization; D005663:Furans / Q000737:chemistry* / Q000378:metabolism; D006576:Heterocyclic Compounds, 4 or More Rings / Q000737:chemistry* / Q000378:metabolism; D007659:Ketones / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D050777:Stathmin / Q000737:chemistry / Q000378:metabolism; D013552:Swine; D014404:Tubulin / Q000737:chemistry* / Q000378:metabolism; D050257:Tubulin Modulators / Q000737:chemistry* / Q000378:metabolism; D014461:Ultracentrifugation; D000095662:Polyether Polyketides
D000970:Antineoplastic Agents; D005663:Furans; D006576:Heterocyclic Compounds, 4 or More Rings; D007659:Ketones; D050777:Stathmin; D014404:Tubulin; D050257:Tubulin Modulators; C421616:ER 076349; C490954:eribulin; D000095662:Polyether Polyketides