Targeting DNA G-Quadruplexes with Helical Small Molecules

We previously identified quinoline-based oligoamide helical foldamers and a trimeric macrocycle as selective ligands of DNA quadruplexes. Their helical structures might permit targeting of the backbone loops and grooves of G-quadruplexes instead of the G-tetrads. Given the vast array of morphologies G-quadruplex structures can adopt, this might be a way to achieve sequence selective binding. Here, we describe the design and synthesis of molecules based on macrocyclic and helically folded oligoamides. We tested their ability to interact with the human telomeric G-quadruplex and an array of promoter G-quadruplexes by using FRET melting assay and single-molecule FRET. Our results show that they constitute very potent ligands—comparable to the best so far reported. Their modes of interaction differ from those of traditional tetrad binders, thus opening avenues for the development of molecules specific for certain G-quadruplex conformations.


Synthetic Schemes
S7 remove all traces of water, characterized by 1 H NMR, and used in the next step without further purification.
General procedure for trimethylsilyl ethanol ester (TMSE) deprotection. The TMSE ester (typically 1 mmol) was dissolved in dry THF (10 mL). Anhydrous TBAF (5 mmol) was added dropwise at room temperature. The reaction mixture was stirred at room temperature for 12h. Reaction progress was monitored by TLC, and additional TBAF was added if necessary to complete the reaction. All volatiles were then evaporated under reduced pressure and the acid was purified by column chromatography on silica gel.
General procedure for nitro group reduction. The nitro precursor (typically 2 mmol) was dissolved in 100 mL of EtOAc, and 200 mg of 10 % wt Pd/C was carefully added under a nitrogen atmosphere. The reaction was stirred under a hydrogen atmosphere for 12 h (pressure provided by a balloon). Reaction progress was monitored by TLC and by 1 H NMR. Additional Pd/C and hydrogen were added if necessary to complete the reaction. Upon completion the catalyst was removed by filtration through celite and the filtrate was evaporated to provide the crude amine. The product was characterized by 1 H NMR and used in the next step without further purification.
General procedure for the removal of the Boc protecting groups. The Boc protected oligomer (typically 1.0 mmol) was dissolved in 4 mL of CH 2 Cl 2 /TFA (1:1 vol/vol) and the solution was stirred at room temperature for 2 h. The solvents were evaporated to provide the crude product. The product was purified by multiple semipreparative HPLC runs using a microsorb column (Microsorb C18, 100Å, 5µ, 250*21.4 mm) and a water/acetonitrile gradient with 0.1% TFA.
General procedure for coupling an amine and an acid. The acid (typically 1.0 mmol) was suspended in 20 mL of dry CH 2 Cl 2 under an argon atmosphere and 1-chloro-N,N-2-trimethylpropenylamine (2.0 mmol) was added. The reaction mixture was stirred at 25 °C for 2 h resulting in a homogeneous solution, then evaporated to provide the corresponding acid chloride. To a solution of the amine (1.1 mmol) in 20 mL of dry CH 2 Cl 2 containing DIPEA (3.7 mmol), the acid chloride in dry CH 2 Cl 2 was added. The reaction mixture was stirred at 25°C for 12 h, then the solvent was evaporated.
The reaction mixture was stirred for 12 h at 25 °C. Water was then added and the aqueous layer was extracted with CH 2 Cl 2 (2 x 5 mL). The combined organic layers were washed with brine (5 mL), dried over anhydrous MgSO 4 and filtered. The filtrate was concentrated under reduced pressure. The resulting crude pentafluorophenyl ester was taken directly in THF (5 mL) and a catalytic amount of 10% Pd/C (10 mg) was carefully added under an oxygen free atmosphere. The reaction mixture was stirred under a hydrogen atmosphere at 25°C for 12 h. The reaction progress was monitored by TLC. The reaction mixture was filtered over celite. Solids were washed with CH 2 Cl 2 (2 x 4 mL). Solvents were removed under reduced pressure, to obtain the crude amino-pentafluoropenyl ester which was set to react immediately without further purification. This material (70 mg, 0.05 mmol) was dissolved in dry toluene (5 mL). A catalytic amount of DMAP (5 mg) was added and the mixture was stirred at 100 o C for 4 h.