Reversible Action of Diaminothiazoles in Cancer Cells Is Implicated by the Induction of a Fast Conformational Change of Tubulin and Suppression of Microtubule Dynamics

Abstract

Diaminothiazoles are novel cytotoxic compounds that have shown efficacy toward different cancer cell lines. They show potent antimitotic and antiangiogenic activity upon binding to the colchicine-binding site of tubulin. However, the mechanism of action of diaminothiazoles at the molecular level is not known. Here, we show a reversible binding to tubulin with a fast conformational change that allows the lead diaminothiazole DAT1 [4-amino-5-benzoyl-2-(4-methoxy phenyl amino)thiazole] to cause a reversible mitotic block. DAT1 also suppresses microtubule dynamic instability at much lower concentration than its IC 50 value in cancer cells. Both growth and shortening events were reduced by DAT1 in a concentration-dependent way. Colchicine, the long-studied tubulin-binding drug, has previously failed in the treatment of cancer due to its toxicity, even though it generates a strong apoptotic response. The toxicity is attributable to its slow removal from the cell due to irreversible tubulin binding caused by a slow conformational change. DAT1 binds to tubulin at an optimal pH lower than colchicine. Tubulin conformational studies showed that the binding environments of DAT1 and colchicine are different. Molecular dynamic simulations showed a difference in the number of H-bonding interactions that accounts for the different pH optima. This study gives an insight of the action of compounds targeting tubulin's colchicine-binding site, as many such compounds have entered into clinical trials recently.

SKLB060 Reversibly Binds to Colchicine Site of Tubulin and Possesses Efficacy in Multidrug-Resistant Cell Lines

“…Thus, this could provide an explanation for the different levels of reversibility on tubulin binding by MPC-6827 and SKLB060. As the reversibility of tubulin inhibitors is important for predicting the efficacy and side effects [26], we believe that SKLB060 might be less toxic and have more potential applications.…”

“…Since the reversibility of tubulin inhibitors is an important parameter in terms of predicting in vivo efficacy and any undesirable side effects [26], we attempted to measure how SKLB060 and MPC-6827 binds to tubulin via a reported method [8]. A2780 cells were incubated for 8 h with 1 μM SKLB060 or 1 μM MPC-6827 (high enough concentrations to ensure tubulin was completely bound to the compounds), thoroughly washed, and then incubated for 24 h and 48 h (Fig.…”

N-(2-methyl-indol-1H-5-yl)-1-naphthalenesulfonamide: A novel reversible antimitotic agent inhibiting cancer cell motility

“…This property of mitotic poisons is not only compound-specific but also difficult to explain in terms of SAR: even small structural changes in compounds within a given class can lead to profound differences in mitotic block reversibility [41]. Reversibility is an important parameter in terms of predicting in vivo efficacy and undesirable side effects, as illustrated by the fact that slow dissociation from tubulin is thought to be the main reason behind colchicine's toxicity, which has led to its failure in the clinic [28]. Taking this into account, we investigated the cellular response upon 8e removal.…”

“…Found: C,66.86,H,4.65,N,8.48,S,9.59. 28,H,4.95,N,15.29,S,8.75. Found: C,62.24,H,5.24,N,14.71,S,8.60.…”

“…Since reversibility of antimitotic compounds is an important parameter in terms of predicting in vivo efficacy and undesirable side effects [28], we investigated the reversibility of the cellular effects of sulfonamide 8e. We first analyzed whether disrupting the treatment with 8e would lead to the proper re-assembly of the microtubule network.…”