R. Silvestri - Indole nucleus as a selected pharmacophore for the design of novel highly potent anti-viral agents active against HIV-1 drawing substituent(s) at position(s) 4 and 5 of the indole is currently in progress. Inhibitors of Tubulin Polymerization. Microtubules (MTs) are essential cytoskeletal polymers that are made of repeating α, β-tubulin (TB) heterodimers and are present in all eukaryotes. The crucial role of MTs in vital cellular functions for tumor cells, including mitosis, motility and cell-cell contacts, has made of MT interfering agents (MIAs) one of the best classes of cancer chemotherapeutic drugs available to date. A large number of chemically diverse compounds are able to bind TB or MTs and inhibit proliferation by acting on the mitotic spindle. Some of these compounds (vinca alkaloids, colchicine) inhibit MT polymerization, whereas others (taxanes) stabilize MTs. While drugs that act on the vinca and taxane sites have well-established roles in the treatment of human cancers, the therapeutic potential of the colchicine site in cancer treatment has yet to be realized. Recently, we have <strong>report</strong>ed arylthioindoles (ATIs) as a new class of potent inhibitors of tubulin polymerization and the growth of some carcinoma cells that bind to colchicine site on β-tubulin close to its interface with α-tubulin. Fist generation ATIs were characterized by (i) an ester function at position 2 of the indole nucleus; (ii) a 3-arylthio group; (iii) an appropriate substituent on the indole nucleus. Focusing our attention on amino derivatives related to combretastatin A-4, we designed and synthesized second generation of ATI derivatives, bearing a hydrogen atom or a methyl group at position 2 of the indole nucleus. New ATIs were synthesized by a two-step procedure. O-Ethyl-S-(3,4,5-trimethoxyphenyl) carbonodithioate was transformed into 3,4,5trimethoxythiophenol by heating at 65 °C in aqueous ethanol in the presence of sodium hydroxide. This mixture was made acidic with 6 N HCl and treated at 25°C with the appropriate indole while adding dropwise an aqueous iodine-potassium iodide solution. The new compounds strongly inhibited tubulin polymerization, with activity in the low micromolar range, comparable to the effects of colchicine and combretastatin A-4. Derivatives bearing a halogen atom or a small alkyl or ether group at position 5 of the indole nucleus, were also potent inhibitors of MCF-7 124 cell growth. For example, 5-bromo-3-[(3,4,5trimethoxyphenyl)thio]-1H-indole showed tubulin polymerization IC 50 = 1.6 µM and growth of MCF- 7 cells IC 50 = 43 nM, and inhibited [ 3 H]colchicine binding at 65%. Cell cycle analysis revealed an accumulation of HeLa cells in the G2/M phase at 24 h and polyploidization at 48 h. At 24 h, inhibition of tubulin polymerization had not caused extensive apoptosis, suggesting that impaired cell viability might occur through a “mitotic catastrophe”. Molecular modelling studies showed that, despite the absence of the ester moiety, the second generation ATIs bind into the colchicine site in the same orientation as the first generation ones. Binding to β-tubulin involved formation of a hydrogen bond between the indole and Thr179 and positioning of the trimethoxy phenyl group in a hydrophobic pocket near Cys241. The synthesis of new derivatives obtained by bioisosteric replacement of the sulfur bridge with either carbonyl or methylene, and some related groups, and the indole nucleus with differently substituted pyrrole and imidazole rings is currently in progress. Selected publications La Regina G, Coluccia A, Piscitelli F, Bergamini A, Sinistro A, Cavazza A, Maga G, Samuele A, Zanoli S, Novellino E, Artico M, Silvestri R. Indolyl aryl sulfones as HIV-1 non-nucleoside reverse transcriptase inhibitors: role of two halogen atoms at the indole ring in developing new analogues with improved antiviral activity. J Med Chem. 2007, 50:5034-8. La Regina G, Edler MC, Brancale A, Kandil S, Coluccia A, Piscitelli F, Hamel E, De Martino G, Matesanz R, Díaz JF, Scovassi AI, Prosperi E, Lavecchia A, Novellino E, Artico M, Silvestri R. Arylthioindole inhibitors of tubulin polymerization. 3. Biological evaluation, structure-activity relationships and molecular modeling studies. J Med Chem. 2007, 50: 2865-74. Ragno R, Coluccia A, La Regina G, Silvestri R. Indolyl aryl sulphones as HIV-1 reverse transcriptase inhibitors: docking and 3D QSAR studies. Exp Opin Drug Discovery 2007, 2:87-114.
AREA7 Biology of malaria and other vector-borne diseases
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