Abstract
The c-MYC proto-oncogene is a regulator of fundamental cellular processes such as cell cycle progression and apoptosis. The development of novel c-MYC inhibitors that can act by targeting the c-MYC DNA G-quadruplex at the level of transcription would provide potential insight into structure-based design of small molecules and lead to a promising arena for cancer therapy. Herein we report our finding that two simple bis-triazolylcarbazole derivatives can inhibit c-MYC transcription, possibly by stabilizing the c-MYC G-quadruplex. These compounds are prepared using a facile and modular approach based on Cu(I) catalysed azide and alkyne cycloaddition. A carbazole ligand with carboxamide side chains is found to be microenvironment-sensitive and highly selective for “turn-on” detection of c-MYC quadruplex over duplex DNA. This fluorescent probe is applicable to visualize the cellular nucleus in living cells. Interestingly, the ligand binds to c-MYC in an asymmetric fashion and selects the minor-populated conformer via conformational selection.
Highlights
Consecutive 5′ G-runs[9,10]
Ligand BTC f showed the highest Δ Tm for promoter gene quadruplexes that can be measured by this method. It showed a Δ Tm value of 22.7 ± 1.4 K for c-MYC at 100 nM, 39.4 ± 2.5 K for c-KIT1 at 750 nM and 23.2 ± 1.6 K for c-KIT2 at 500 nM ligand concentrations. These results revealed that 5–7 fold higher concentrations of BTC f are required for the c-KIT1 and c-KIT2 to achieve maximum stabilization potential (Δ Tm) compared to the c-MYC quadruplex
Competitive Förster Reson ance Energy Transfer (FRET)-melting experiments (Fig. 2b) clearly showed that the presence of 100 mol equivalent excess ds DNA did not significantly interfere with the stabilization of quadruplexes induced by the ligand BTC f, indicating its high selectivity for the quadruplexes over duplex DNA
Summary
Consecutive 5′ G-runs[9,10]. Various G-quadruplex structures derived from different G-rich tracts of the c-MYC regulatory element NHE III1 have been reported[11,12,13]. The pioneering work of Siddiqui-Jain et al showed that small molecules stabilizing the c-MYC G-quadruplex can reduce c-MYC transcription in cancer cells[14]. Several classes of small molecules, which can bind and stabilize c-MYC quadruplex have been developed[15,16,17,18,19,20,21,22,23,24,25,26,27] and few of them have been structurally characterized in complex with c-MYC G-quadruplex by NMR12,24. We delineate a modular synthetic access to novel bis-triazolyl carbazole derivatives[29] as potent “turn on” G-quadruplex probes, which bind to c-MYC quadruplex via conformational selection with the potential to down-regulate c-MYC transcription in hepatocellular carcinoma cells
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