Novel Pyridazin-3(2H)-one-Based Guanidine Derivatives as Potential DNA Minor Groove Binders with Anticancer Activity

Novel aryl guanidinium analogues containing the pyridazin-3(2H)-one core were proposed as minor groove binders (MGBs) with the support of molecular docking studies. The target dicationic or monocationic compounds, which show the guanidium group at different positions of the pyridazinone moiety, were synthesized using the corresponding silyl-protected pyridazinones as key intermediates. Pyridazinone scaffolds were converted into the adequate bromoalkyl derivatives, which by reaction with N,N’-di-Boc-protected guanidine followed by acid hydrolysis provided the hydrochloride salts 1–14 in good yields. The ability of new pyridazin-3(2H)-one-based guanidines as DNA binders was studied by means of DNA UV-thermal denaturation experiments. Their antiproliferative activity was also explored in three cancer cell lines (NCI-H460, A2780, and MCF-7). Compounds 1–4 with a bis-guanidinium structure display a weak DNA binding affinity and exhibit a reasonable cellular viability inhibition percentage in the three cancer cell lines studied.

S2 SCRF-SMD approach implemented in the Gaussian16 package including dispersing, repulsing and cavitation energy terms of the solvent in the optimization. Optimised structures of all the ligands studied are shown in Figure S1.

Docking experiments
The program Autodock Vina 4.2 was used to carry out docking studies [2]. The ligands were flexibly docked into the rigid DNA minor groove model (crystal structure of a pentamidine-oligonucleotide complex, PDB: 1D64 [3]). Scores (G-scores) were measured in kcal/mol and are only indicative of the quality of the interaction with the target; they do not provide a quantitative measure of binding. Poses obtained from the docking were visualised with VMD [4].

General methods
The starting chemical materials were of commercial sources and used as provided. The solvents were purified and dried following standard protocols.  [5,6,7].   in hexane (0.78 mmol). The reaction mixture was stirred at this temperature for 3 h, quenched with t BuOMe (0.40 mL), H 2 O (60 µL) and NaOH 4M (30 µL) and stirred overnight at room temperature. The resulting white precipitate was filtered off, the solvent was evaporated under reduce pressure and the residue was purified by column chromatography to afford the desired compound.

S27
Compound 46 was purified by column chromatography on silica gel (EtOAc
The cytotoxic potency of compounds, measured as 50% inhibitory concentrations (IC 50 ), was calculated from concentration-effect curves by using GraphPad Prism software, version 2.01. Correlation coefficients (r 2 ) were higher than 0.995 for the compounds tested.