Making epothilones fluoresce: design, synthesis, and biological characterization of a fluorescent N12-aza-epothilone (azathilone).
A green fluorescent 12-aza-epothilone (azathilone) derivative has been prepared through the attachment of the 4-nitro-2,1,3-benzoxadiazole (NBD) fluorophore to the 12-nitrogen atom of the azamacrolide core structure. While less potent than natural epothilones or different N12-acylated azathilone derivatives, NBD-azathilone (3) promotes tubulin assembly, inhibits cancer cell proliferation in vitro and arrests the cell cycle at the G2/M transition. Most significantly, the binding of 3 to cellular microtubules (MTs) could be directly visualized by confocal fluorescence microscopy. Based on competition binding experiments with laulimalide-stabilized MTs in vitro, the N12-Boc substituted azathilone 1, Epo A, and NBD-azathilone (3) all interact with the same tubulin-binding site. Computational studies provided a structural model of the complexes between beta-tubulin and 1 or 3, respectively, in which the NBD moiety of 3 or the BOC moiety of 1 directly and specifically contribute to MT binding. Collectively, these data demonstrate that the cellular effects of 3 and, by inference, also of other azathilones are the result of their interactions with the cellular MT network.
Gertsch
Jürg
J
Department of Chemistry and Applied Biosciences, Institute of Pharmaceutical Sciences, Swiss Federal Institute of Technology (ETH) Zürich, HCI H405, Wolfgang-Pauli-Strasse 10, 8093 Zürich, Switzerland.
Feyen
Fabian
F
Bützberger
Alexander
A
Gerber
Barbara
B
Pfeiffer
Bernhard
B
Altmann
Karl-Heinz
KH
D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D015195:Drug Design; D034261:Epothilones / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008870:Microtubules / Q000378:metabolism; D013057:Spectrum Analysis; D014404:Tubulin / Q000378:metabolism
C547066:10,14-dihydroxy-9,11,13,13-tetramethyl-2-(1-methyl-2-(2-methyl-thiazol-4-yl)-vinyl)-5-(7-nitro-benzo(1,2,5)oxadiazol-4-yl)-1-oxa-5-aza-cyclohexadecane-12,16-dione; D034261:Epothilones; D005456:Fluorescent Dyes; D014404:Tubulin; D009327:4-Chloro-7-nitrobenzofurazan