Arylthioindoles (ATIs) and aroylindoles (AIs) are two classes of modern and potent colchicine-binding site inhibitors of tubulin polymerization and cancer cell growth, recently developed by our research group. Unpublished structure-activity relationship studies pointed out that the introduction of a triazole ring at position 6 or 7 of the indole nucleus improved both water solubility and anticancer activity. Based upon these promising results, aim of this research project is a hit lo lead optimization process by introducing different basic groups (e.g., methylamino, dimethylamino, azetidin-1-yl, pyrrolidin-1-yl, piperidin-1-yl, piperazin-1-yl, 4-methylpiperazin-1-yl and morpholino) at position 6 or 7 of the indole nucleus. In addition, the bioisosteric substitution of the sulphur atom with a ketone bridge and the introduction of a phenyl group at position 2 of the indole nucleus will be explored in all the new derivatives.
Molecular modelling studies will be carried out to clarify the binding mode of the new derivatives into the colchicine-binding site of tubulin, and according to the obtained results the most promising compounds will be prepared. The use of both microwave-assisted synthesis and fully automated flash chromatography system will allow us to slash both reaction and purification times. All the newly synthesized compounds, prepared as free bases and ionic salts, will be tested for determining their ability to inhibit tubulin assembly, binding of colchicine to tubulin as well as MCF-7 human breast carcinoma cell growth. Then, the most active compounds will be evaluated using different cancer cell lines and fully characterized in terms of effects on cell cycle progression, inhibition of microtubule assembly and induction of mitotic arrest or death, as well as pharmacokinetic profile.
Colchicine, an alkaloid extracted from the poisonous meadow saffron Colchicum autumnale L., is the first identified tubulin destabilizing agent. It has been used for many years as an unapproved drug to treat gout, familial Mediterranean fever, pericarditis and Behcet's disease. In 2009, U.S. Food and Drug Administration (FDA) approved colchicine as a monotherapy drug to treat familial Mediterranean fever and acute gout flares. Colchicine can effectively inhibit mitosis, and since cancer cells undergo mitosis at a significantly increased rate they are more susceptible to colchicine poisoning than normal cells. Therefore, colchicine is also being investigated as an anticancer drug. However, colchicine shows limited clinical application for treating cancer because of its low therapeutic index and toxic effects such as neutropenia, gastrointestinal upset, bone marrow damage and anaemia.
Although colchicine is not employed as an anticancer agent, there have been multiple efforts to clinically develop colchicine-binding site inhibitors (CBSIs). As microtubules are important regulators of endothelial cell biology, one advantage of the mechanism of actions of CBSIs is targeting the tumour vasculature. CBSIs can prevent new blood vessels formation by outgrowth from pre-existing ones (angiogenesis inhibitors) or destroy the existing tumour vasculature (vascular disrupting agents, VDA). The targeting of tumour blood vessels introduces a therapeutically promising application for these compounds. Another favourable factor is that most of these drugs have no multidrug resistance (MDR) issues. The major limitation of using microtubule-targeting agents clinically is innate and acquired drug resistance. The most common form of clinical resistance is overexpression of the MDR1 gene, which encodes the P-glycoprotein (Pgp) drug efflux pump. This membrane-associated ATP-binding cassette (ABC) transporter is overexpressed in many tumour cell lines, including tissues of the liver, kidney, and gastrointestinal tract. Over-expression of Pgp decreases intracellular drug levels, consequently limiting drug cytotoxicity. In addition, over-expression of Pgp is associated with poor response to microtubule-targeted agents including taxanes and vinca alkaloids and subsequent treatment failure.
Besides over-expression of ABC transporters, other significant mechanisms of resistance include mutations in tubulin and overexpression of ßIII-tubulin isoform. Among eight identified ß-tubulin isotypes in human, overexpression of class III ß-tubulin is an indicator of resistance to tubulin targeting agents such as paclitaxel and vinorelbine. While the efficacies of some CBSIs such as colchicines and 2-methoxyestradiol were not affected by the expression pattern of ß-tubulin. Therefore, the clinical development of a microtubule-targeting agent that circumvents both of these drug resistance mechanisms could have advantages for patients with drug resistant tumours.
Fueled by the purpose of developing novel chemotherapeutics and with the aim of establishing the benefits of tubulin inhibition, we have recently developed arylthioindoles (ATIs) and aroylindoles (AIs) as modern and potent CBSIs of tubulin polymerization and cancer cell growth. Both ATIs and AIs proved to be much more active than vinorelbine, vinblastine, and paclitaxel in P-glycoprotein overexpressing NCI/ADR-RES and Messa/Dx5 cancer cell lines. Among them, 2-(1H-imidazol-1-yl)-3-((3,4,5-trimethoxyphenyl)thio)-1H-indole showed vascular disrupting effects, good water solubility and interesting metabolic stability. To date, ATIs and AIs not only show greater anticancer profiles but also have smaller molecular weight with chemically accessible structures with respect to both taxanes and Vinca alkaloids; this makes them attractive chemical classes to work with for achieving improved pharmacokinetic properties and efficacy as well as reduced toxicity.
CBSIs as well as VDAs appear to be the most likely agents for future clinical interest. Therefore, the project proposed here is an ambitious medicinal chemistry process which is expected to make a difference in the field of new anticancer drug discovery. Indeed, the results obtained by us so far, open a new window for developing new, potent and selective CBSIs as potential drug candidate for treatment of human cancer.