Characterisation of CTR-17 and CTR-20, novel chalcone derivatives that inhibit tubulin polymerisation activity

Agents targeting colchicine-binding sites are recognised as valuable lead compounds in the development of new anticancer drugs. Although colchicine can effectively inhibit cell proliferation, its use as an anticancer agent has not been approved by the FDA due to its inherent toxicity. To develop colchicine-binding site targeting agents with low or no toxicity, in collaboration with Rajiv Gandhi Technical University of India, several chalcone derivatives were created and examined. Preliminary studies at the Lee Lab identified CTR-17 and CTR-20 as promising leads. Their anti-proliferative activities using three human breast cancer cell lines (MDA-MB468, MDA-MB231 and MCF-7) and two matching noncancer breast cell lines (184B5 and MCF10A) were initially determined. Subsequently, nine other cancer cell lines were used to assess the broad spectrum anti-proliferative effects of the CTR compounds. Data from this study showed that CTR-17 and CTR-20 preferentially kill cancer cells 10-25 times over non-cancer cells. Data obtained from flow cytometry, confocal microscopy and Western blotting showed that CTR-17 induced a prolonged mitotic arrest, leading to cancer cell death probably via apoptosis. I also found that both CTR-17 and CTR-20 inhibited tubulin polymerisation and bound to purified tubulin fibers with a dissociation constant of 4.58±0.95 μM and 5.09±0.49 μM, respectively. CTR-17 and CTR-20 competitively inhibited the binding of colchicine to tubulin with an inhibitory concentration of 5.68±0.35 μM and 1.05±0.39 μM, respectively, suggesting that the CTR compounds bind to tubulin at a site partially overlapping the colchicine-binding site. Molecular docking studies confirmed this binding to occur via two and one hydrogen bonds between tubulin and CTR-20 and CTRiv 17, respectively. More interestingly, CTR compounds inhibit the proliferation of multi-drug resistant cell lines, which overexpress drug transporters involved in the efflux of clinically available microtubule targeting agents. In addition, the CTR compounds exhibit a synergistic relationship with paclitaxel in causing cytotoxicity to a P-glycoprotein overexpressing cell line. Therefore, these novel chalcone derivatives not only possess cancer-specific cell killing property but also the ability to exhibit similar cytotoxicity to both the multi-drug sensitive and resistant cells. Hence, CTR compounds possess substantial potential as safe and effective anticancer drugs.Doctor of Philosophy (PhD.) in Biomolecular Science