We further confirmed mitotic arrest in the presence of 2a and 3a by microscopic analysis of A549, MCF-7 and HeLa cell lines (Fig. ). We used molecular docking approach to identify the potential binding mechanism of compounds 2a and 3a. The docking procedure requires the selection of the tubulin dimer structure to be used as the receptor for the ligand of interest. Available structures of tubulin from the Protein Data Bank (PDB), containing different ligands at the colchicine site, display slightly different conformations of the ligand-binding pocket. This flexibility of the binding site produces a problem for standard 'rigid' molecular docking methods, since the choice of tubulin structure can significantly affect the docking score and hence the ligand pose prediction. We found that this problem can be partially remedied using the 'induced fit' docking approach, incorporated in the Schrödinger software package, which can account for the flexibility of both ligand and receptor. However, even with this approach, the docking score alone is not a reliable predictor of the correct binding pose, as was confirmed by cross-docking of randomly selected colchicine site ligands into tubulin structures co-crystalized with other diverse ligands of the same site (Fig. ).