Microtubules stabilization mechanisms

Taxol s journey to the clinic was slow and arduous. Initial difficulties with aqueous solubility and lack of knowledge regarding its mechanism, of action delayed its development until 1979 when, in another seminal paper in the field, S.B.Horwitz and her collaborators disclosed their findings on the interaction of taxol with microtubules.4 Taxol s unique biological action, which includes promotion of microtubule formation and microtubule stabilization, stimulated a renewed interest in taxol as a potential drug candidate. The problem of procuring adequate supplies of taxol became even... 

Epothilones are naturally occurring cytotoxic macrolides, which were initially isolated from a mycobacterium. Their antitumor activity is similar to that of the clinically established taxoids (Taxol, Taxotere), by interrupting the dynamic mechanism of microtubule assembly/disassembly via microtubule stabilization. In contrast to taxoids, epothilones are remarkably efficient against multidrug resistant cells. 

The third chapter describes a comprehensive study of the mechanisms of activity of microtubules stabilizing drugs. Thermodynamic, kinetic, structural and functional data on microtubules stabilizing drugs are discussed in an interdisciplinary manner to generate a time-resolved picture of the interaction of the drugs with different tubulin forms. 

On the other hand, from a thermodynamical point of view, a common mechanism for assembly induction can be proposed. All microtubule stabilizing agents bind tightly to the assembled form, while they do not bind with a measurable affinity to the dimeric tubulin [33], indicating that the taxane binding pocket is either not formed or not-completely formed in non-microtubular tubulin. 

However, since microtubule stabilizing agents will bind tightly to unoccupied sites, it is reasonable to assume that both mechanisms should work however, in this case the apparent critical concentration will saturate following Eq. (5) (as in Eq. 4) in the case where path 3b is followed the term ([Tub-Lig]+[Tub])[Mtb] should be replaced by ([Mtb-Lig] + [Mtb]) [Tub]) ... 

Peluroside A was the first microtubule stabilizing agent whose conformation has been determined bound to microtubules (those of Paclitaxel and Epothilone were determined in non-microtubular tubulin [5, 12, 38, 91]). In the bound state, the NMR data, assisted by molecular mechanics calculations and docking experiments, indicated that only one (that present in water, B) of the two major conformations existing in water solution is bound to microtubules (a-tubulin). A model of the binding mode to tubulin has also been proposed [27], involving the a-tubulin monomer, in contrast with paclitaxel, which binds to the p-monomcr. 

After the identification of the paclitaxel mechanism of action, several additional microtubule stabilizing agents were discovered from natural sources (Chart 9). Among them, epothilones (12-15) are the most studied and characterized. Epothilones A and B (12,13) were first isolated by Hofle and coworkers [54] from a myxobacterium (Sorangium cellulosum strain 90) in 1993, while their activity in stabilizing microtubule similarly to paclitaxel was reported for the first time by Bollag and co-workers in 1995 [55], Experiments with radio-labeled paclitaxel... 

Apoptosis is another important mechanism for cell poisoning, in addition to tubulin assembly promotion and microtubule stabilization for paclitaxel, especially at a high concentration (sub pM to pM level). It is also observed that in higher concentrations (5 to 50 pM), paclitaxel induced tumor necrosis through microtubules rather than apoptosis. " " Note that paclitaxel concentration in the clinic is lower than pM. 

Tirnauer JS, Grego S, Salmon ED, Mitchison TJ. EBl-microtubule interactions in Xenopus egg extracts role of EBl in microtubule stabilization and mechanisms of targeting to microtubules. Mol. Biol. Cell 2002 13 3614-3626. 

Bollag DM, McQueney PA, Zhu J, Hensens O, Koupal L, Liesch J, Goetz M, Lazarides E, Woods CM. Epothilones, a new class of microtubule-stabilizing agents with a taxol-like mechanism of action. Cancer Res. 1995 55 2325-2333. 

Two kinds of resistant biotypes have been noted one is highly-resistant (R) and is unaffected even by saturated solutions of dinitroaniline herbicide, whereas an intermediate-resistant (I) biotype is only 50X resistant to trifluralin and less than 10X resistant to oryzalin compared with the susceptible (S) biotype. Both R and I biotypes are cross-resistant to phosphoric amide herbicides. Tubulin from the R is able to polymerize into microtubules even in the presence of oryzalin, whereas that of the S biotypes cannot. Western blots of tubulin from the R biotype reveal two -tubulin isotypes whereas only one form is noted in the S biotype. Because the R biotype is hypersensitive to the microtubule-stabilizing agent taxol, it is likely that the R biotype is resistant by having hyperstabilized microtubules. The I biotype has no gross alteration in tubulin nor extreme sensitivity to taxol, indicating that this biotype has a different resistance mechanism than the R. 

Other drugs bind to different sites on tubulin dimers or to microtubules and therefore affect microtubule stability through different mechanisms. For example, at low concentrations, taxol binds to microtubules and stabilizes them by inhibiting their shortening. 

Bramlett HM, Dietrich WD (2004) Pathophysiology of cerebral ischemia and brain trauma similarities and differences. J Cereb Blood Flow Metab 24 133-150 Brown TP, Rumsby PC, Capleton AC, Rushton L, Levy LS (2006) Pesticides and Parkinson s disease - is there a link Environ Health Perspect 114 156-164 Butler D, Bendiske J, Michaelis ML, Karanian DA, Bahr BA (2007) Microtubule-stabilizing agent prevents protein accumulation-induced loss of synaptic markers. Eur J Pharmacol 562 20-27 Campbell A, Smith MA, Sayre LM, Bondy SC, Perry G (2001) Mechanisms by which metals promote events connected to neurodegenerative diseases. Brain Res Bull 55 125-132... 

Eleuthosides. Marine natural products from soft corals (Eleutherobia albiflora) which exhibit stroM cytotoxic and anticancer properties with a taxol -like mechanism of action (tubulin polymerization and microtubule stabilization). Most active is eleutherobin, first isolated in 1994 C35H48N2OK), Mr 656.76, [a] -67° (CH3OH), oil. Closely related are E. A and B from... 

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