Effect of 2'-OH acetylation on the bioactivity and conformation of 7-O-[N-(4'-fluoresceincarbonyl)-L-alanyl]taxol. A NMR-fluorescence microscopy study.
The relationship between conformation, 2'-OH acetylation, and bioactivity of two fluorescent taxoids has been investigated by a combination of NMR and fluorescence microscopy techniques. These taxoids present the structure of taxol with the 7-OH group esterified with the N-(4'-fluoresceincarbonyl)-L-alanine group and with the 2'-OH group free (taxoid 2) or acetylated (taxoid 3). The larger water solubility of 2 and 3 compared with taxol allowed a detailed NMR study in DMSO-d6/D2O (3/7), showing that both taxoids adopt a similar collapsed conformation in which the hydrophobic groups 2-O-benzoyl, 3'-phenyl and 4-O-acetyl are in close proximity, with the fluorescein group displaying unrestricted motion. On the other hand, while taxoid 2 retains essentially the ability of taxol to induce in vitro microtubule assembly and to bind to cell microtubules, the 2'-acetylated derivative 3 does not show immediate activity. However, when taxoid 3 is left in the cell culture, the slow hydrolysis of the 2'-acetate group in the medium liberates the cytotoxic, microtubule-specific taxoid 2. The intense emission of this active derivative (2) allows the accurate recording of the drug-cell interaction from the very initial steps using fluorescence microscopy. These experiments show conclusively, for the first time in cell cultures, that a free 2'-OH group in taxol is essential for the recognition of the drug by the binding site of cellular microtubules.
Jiménez-Barbero
J
J
Instituto de Química Orgánica, CSIC, Madrid, Spain.
Souto
A A
AA
Abal
M
M
Barasoain
I
I
Evangelio
J A
JA
Acuña
A U
AU
Andreu
J M
JM
Amat-Guerri
F
F
D000107:Acetylation; D000818:Animals; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000187:drug effects; D009682:Magnetic Resonance Spectroscopy; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D017239:Paclitaxel / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000494:pharmacology; D043823:Taxoids; D014404:Tubulin / Q000378:metabolism
C116524:2'-O-acetyl-7-O-(N-(4'-fluoresceincarbonyl)alanyl)taxol; C116523:7-O-(N-(4'-fluoresceincarbonyl)alanyl)taxol; D043823:Taxoids; D014404:Tubulin; D017239:Paclitaxel