Microtubule targeting agents: from biophysics to proteomics

Abstract

This review explores various aspects of the interaction between microtubule targeting agents and tubulin, including binding site, affinity, and drug resistance. Starting with the basics of tubulin polymerization and microtubule targeting agent binding, we then highlight how the three-dimensional structures of drug-tubulin complexes obtained on stabilized tubulin are seeded by precise biological and biophysical data. New avenues opened by thermodynamics analysis, high throughput screening, and proteomics for the molecular pharmacology of these drugs are presented. The amount of data generated by biophysical, proteomic and cellular techniques shed more light onto the microtubule-tubulin equilibrium and tubulin-drug interaction. Combining these approaches provides new insight into the mechanism of action of known microtubule interacting agents and rapid in-depth characterization of next generation molecules targeting the interaction between microtubules and associated modulators of their dynamics. This will facilitate the design of improved and/or alternative chemotherapies targeting the microtubule cytoskeleton.

Fig. 1

Microtubule polymerization phases. Microtubules grow by addition of GTP-tubulin subunits. After their incorporation, subunits are hydrolyzed to become GDP-tubulin. The straight tubulin conformation within the microtubule lattice is stabilized by a ‘cap’ of tubulin-GTP subunits. Closure of the terminal sheet structure generates a metastable, blunt-ended microtubule intermediate, which might pause, undergo further growth, or switch to the depolymerization phase possibly by loss of the GTP-tubulin at the end. GDP-tubulin is free to splay out and the microtubule rapidly shrinks. Microtubules can transition from growth to shrinking phases called catastrophes. Protection of microtubules from shrinking by adding GTP-tubulin at the tip of microtubule is referred to as rescue

Fig. 2

Structure of microtubule destabilizers and stabilizers

Fig. 3

Tubulin binding site of MTAs

Fig. 4

MALDI-MS/MS and MALDI-IMS-MS/MS whole body imaging of rats dosed at 6 mg/kg IV with vinblastine. Distribution of the precursor ion m/z 811.4 (vinblastine) and production m/z 751 (vinblastine dissociation product) is shown. Comparison of a m/z 811 and b m/z 811-751 images obtained using MALDI-IMS-MS/MS and conventional MALDI-MS/MS clearly demonstrate the advantages of ion mobility separation within MALDI xenobiotic imaging. The main difference observed is indicated; the distribution of the ions of interest within the renal pelvis. White contrast indicates vinblastine concentration. c Whole body autoradiograph showing the distribution of ³H in a 1-h post-dose rat dosed with 6 mg/kg IV ³H vinblastine, confirming the distribution observed with MALDI-IMS-MS/MS. Reprinted with the authorization of the American Chemical Society