Design, synthesis, and biological evaluation of biotinylated colchicine derivatives as potential antitumor agents

Abstract

Chemical drug design based on the biochemical characteristics of cancer cells has become an important strategy for discovering new anti-tumour drugs to improve tumour targeting effects and reduce off-target toxicities. Colchicine is one of the most prominent and historically microtubule-targeting drugs, but its clinical applications are hindered by notorious adverse effects. In this study, we presented a novel tumour-specific conjugate 9 that consists of deacetylcolchicine (Deac), biotin, and a cleavable disulphide linker. 9 was found to exhibit potent anti-tumour activity and exerted higher selectivity between tumour and nontarget cells than Deac. The targeting moiety biotin might enhance the transport capability and selectivity of 9 to tumour cells via biotin receptor-mediated endocytosis. The tubulin polymerisation activity of 9 (with DTT) was close to the parent drug Deac. These preliminary results suggested that 9 is a high potency and reduced toxicity antitumor agent and worthy of further investigation.

Keywords: Colchicine; adverse effects; biotin; conjugate; disulphide bond.

Figure 1.

Chemical structures of biotin and biotin conjugates.

Figure 2.

Chemical structures of colchicine and its derivatives.

Figure 3.

Reduction-sensitive drug release mechanism of Deac-SS-Biotin triggered by DTT (Glutathione mimetic).

Scheme 1.

Reagents and conditions: (a) 4-nitrophenyl carbonochloridate, Et₃N, THF, rt., 6 h; (b) THF, rt., 4 h; (c) 1, DCC, DMAP, rt., 20 h; (d) HATU, Et₃N, rt., 8 h; (e) anhydrides, NMM, DMSO, rt., 45 min; (f) (i) SOCl₂, MeOH, rt., overnight; (ii) NH₂NH₂, rt., 17 h; (g) EDCI, HOBt, DMAP, Et₃N, rt., 10 h.

Figure 4.

Stabilities of Deac-SS-Biotin (9, 5 µM) in water, and PBS cell culture fluid were investigated by HPLC analysis. (A) HPLC analysis of Deac-SS-Biotin (9) in water after incubation for and 72 h; (B) HPLC analysis of Deac-SS-Biotin (9) in PBS after incubation for and 72 h; (C) HPLC analysis of Deac-SS-Biotin (9) in cell culture fluid after incubation for and 72 h.

Figure 5.

In vitro release of Deac from prodrugs. (A) Release profiles of Deac from Deac-SS-Biotin (9, 5 μM) with 0 μM, 5 μM, 10 μM and 20 μM DTT in PBS (pH 7.2–7.4) (n = 3). (B) Release profiles of Deac from Deac-Biotin (10, 5 μM) with 0 μM, 5 μM, 10 μM and 20 μM DTT in PBS (pH 7.2–7.4) (n = 3). (C) HPLC spectra of Deac-SS-Biotin (9, 5 μM) with 10 μM DTT at 0 and 24 h.

Figure 6.

In vitro cytotoxicity of biotin, Deac (13), and Deac-SS-Biotin (9) in A549 cells. Cytotoxicity of biotin, Deac + biotin, and Deac-SS-Biotin + biotin in A549 cells (n = 3).

Figure 7.

The effect of Deac-SS-Biotin (9) on tubulin polymerisation. The tubulin had been pre-incubated for 1 min with Deac (13) at 5 µM, Deac-SS-Biotin (9) at 5 µM, Deac-SS-Biotin (9) at 5 µM and DTT at 10 µM, Deac (13) at 5 µM, Colchicine at 5 µM, Paclitaxel at 5 µM or vehicle DMSO at room temperature before GTP was added to start the tubulin polymerisation reactions. The reaction was monitored at 37 °C.