Tubulin-binding properties

The cause of the cell cycle specificity of the bisindole alkaloids may be associated with the ability of these compounds to interact with the protein tubulin and thereby inhibit the polymerization (and depolymerization) of microtubules (16,17). In this respect the cellular pharmacology of vinca alkaloids is similar to that of other cytotoxic natural products such as colchicine or podophyllotoxin. On closer inspection, however, Wilson determined that the specific binding site on tubulin occupied by vinblastine or vincristine is chemically distinct from the site occupied by the other natural products (18). Subsequent experiments have determined that the maytansinoids, a class of ansa-macrocycles structurally distinct from the bisindoles, may bind to tubulin at an adjacent (or overlapping) site (19). A partial correlation of the antimitotic activity of these compounds with their tubulin binding properties has been made, but discrepancies in cellular uptake probably preclude any quantitative relationship of these effects (20). 

The relative tubulin binding properties of a collection of bisindole alkaloids reflect the importance of combined modifications at C-4 and N-1 (Table XII) (94). Replacement of the vinblastine C-4 hydroxyl with hydrogen results in reduction in tubulin affinity by over 50%. However, when this substitution is combined with inversion of configuration at C-4, the tubulin binding affinity is increased by 84%. Replacement of the N-1 methyl with formyl increases tubulin affinity an additional 37%. The... 

The tubulin-binding properties of (-)-rhazinilam were discovered through screening of a number of Malaysian plant extracts [60]. Natural (-)-rhazinilam induces tubulin spiralization, inhibiting tubulin assembly in the same way as vinblastine-like alkaloids, and protects microtubules from cold disassembly such as with paclitaxel [67]. This effect has never been observed with other microtubule poisons. For this reason, and despite the in vivo inactivity of (-)-rhazinilam [67], a number of analogues have been prepared by semi-synthesis and total synthesis (see Sections 3.1.3. and 3.2.3.) in order to improve the pharmacological properties of this molecule. 

Singer WD, Himes RH. Cellular uptake and tubulin binding properties of four vinca alkaloids. Biochem Pharmacol 1992 43(3) 545-51. 

It has been shown that combretastatin A-4 disrupts the microtubules of human umbilical vein endothelial cells (HUVECs) in culture, thus confirming that the tubulin binding properties shown in cell-free systems are retained when the compound enters cells and that tubulin binding is a significant component of the biological activity. Also 3-fluoro- and 3-chloro derivatives retained activity in human umbilical vein endothelial cells. This kind of activity against endothelial cells is extremely important, as endothelial cells play a key role in the angiogenic process. 

Another anti-cancer agent in clinical use is podophyllotoxin (3-59) this has an aryl tetrahydronaphthalene lignan lactone skeleton, and demonstrates potent tubulin-binding, anti-mitotic properties (Scheme 3.16) [30]. The Sherburn group [31] prepared this molecule by a tris(trimethylsilyl)silane promoted conversion of thionocarbonate 3-55 into the lactone 3-58, which proceeded with a yield of 38 %. As intermediates, the radicals 3-56 and 3-57 can be assumed. 

Fast binary filtering methods can also be used for scaffold ranking, i.e., the prioritization of combinatorial scaffolds based on predicted properties. Privileged scaffolds were selected to demonstrate this idea [82], Piperazines SI, benzodiazepines S2, and spiroindolines S3 have been described as GPCR-privileged scaffolds [83], Scaffold S4 represents a SPIKET motif for tubulin binding which is effective for inhibiting cellular proliferation [84], Dysidiolide-derived compounds... 

The covalent binding property of benzamides has been exploited in the development of cell-based competitive binding assays that measure the ability of other antitubulin agents to inhibit binding of radiolabeled benzamides to j -tubulin in whole cells. The tritiated S-enantiomer of zoxamide has been used to study the zoxamide binding site in the Oomycete Phytophthora capsid [7]. Tritiated analogs 2 and 3 (RH-4032 and RH-5854, Fig. 16.1.2) have been used in similar assays in plant [4] and mammalian cells [6], respectively. 

The discovery of the tubulin-assembly properties of these natural products have raised the obvious questions as to whether they bind to the same binding site as taxol and whether there is thus a common pharmacophore that can be deduced for all these compounds. These questions have been addressed in various ways. The first attempt at developing a common pharmacophore was by Winkler and Axelsen 499), who simply compared the structures of taxol and epothilone and proposed a pharmacophore based on regions of structural similarity. 

Chalcones (including 11) contain a l,3-diaryl-a,/)-unsaturated ketone moiety and have anti-cancer properties [38]. As analogs of CA-4, 7, the mode of cytotoxic action of chalcones has been shown to be similar to the com-bretastatins. They bind to the colchicine site of tubulin and inhibit tubulin polymerization [39]. 

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