Synthesis, structure and properties of 7’-((4-amino-5-thio-1,2,4- triazole-3-yl)methyl)theophylline derivatives

Materials and methods. Theophylline was used as the starting material. Using alkylation reactions, hydrazinolysis, interaction with a carbon disulfide followed by heterocyclization with an excess of hydrazine hydrate, 7’-((4-amino-5-thio-1,2,4-triazole-3-yl)methyl) theophylline was obtained. The following stages of the chemical conversion included alkylation reactions with haloalkanes, the formation of azomethine compounds by reaction with aromatic aldehydes, and the reaction with aromatic carboxylic acid chlorides. The structure of the obtained compounds was confirmed by data of elemental analysis, 1H NMR spectroscopy and IR-spectrophotometry. The individuality of substances was established by using high performance liquid chromatography with diode-array and mass spectrometric detection.

Methods of combining these compounds involve the formation of a 1,2,4-triazole moiety using xanthine as the starting compound. The literature describes various approaches to the conditions of this transformation [7]. But determining the optimal conditions for this process remains relevant despite the advances in research in this direction.

Aim
The aim of our work was to search for promising compounds from the point of biological activity in a series of derivatives that combine heterocyclic fragments of theophylline and 1,2,4-triazole.

Materials and methods
The implementation of the experimental part of the work has been accompanied by the use of traditional methods of organic synthesis [2,4,8]. Melting points were determined in open capillary tubes in a "MPA 100". The elemental analysis (C, H, N) were performed through the "Elementar vario EL cube" analyzer. IR spectra (4000-400 cm -1 ) were taken using "ALPHA FT-IR spectrometer". 1 H NMR spectra (400 MHz) were recorded at "Varian-Mercury 400" spectrometer with SiMe 4 as internal standard in DMSO-d 6 solution. Chromatography-mass spectral studies were conducted on the instrument "Agilent 1260 Infinity HPLC" equipped with a mass spectrometer "Agilent 6120" (method of ionizationelectrospray (ESI)).

Chemistry
In the primary stage, the synthesis of theophylline ester was performed using 2-chloroacetic acid, followed by hydrazinolysis and heterocyclization in excess of hydrazine. The formed thiols were used in the reactions of alkylation, synthesis of Schiff bases and carboxamides. The influence of the nature of the solvent and the duration of heating on the yield of the reaction products were investigated. Sodium salt, ester and hydrazide were prepared according to known methods [7]. Hydrazinolysis and subsequent heterocyclization carried out using traditional methods of organic synthesis. The resulting 7'-((4-amino-5-thio-1,2,4-triazole-3-yl)methyl)theophylline 7'-((4-Amino-3-thio-1,2,4-triazole-5-yl)methyl)theophylline (7). 1 g (0.02 mol) N 2 H 4 ·H 2 O was added to a solution of 3.66 g (0.01 mol) of the potassium 2-(2-(theophylline-7-yl) acetyl)hydrazine-1-carbodithioate dissolved in 3 ml of water. The mixture was refluxed for 2 h, cooled, diluted with water and acidified with CH 3 COOH. The product was crystallized from ethanol and isolated as a white solid.

Molecular docking
Molecular docking was performed to obtain structural information on the interaction of the synthesized compounds and the corresponding biological structure [5]. The Х-ray crystal structures of the corresponding biological targets from the protein database (PDB-ID) in complex with the standard ligand were previously downloaded: kinases of anaplastic lymphoma in the complex of crizotinib (2XP2), lanosterol 14-α-demethylase with ketoconazole (3LD6), cyclooxygenase-2 with diclofenac (4Z0L). The ligands (crizotinib, ketoconazole, diclofenac) were previously removed from the primary structures. Carried out the joining of different ligands to the protein using AUTODOCK. The conformations of the ligand were analyzed in terms of energy, hydrogen bonding and hydrophobic interaction between the ligand and
In obedience to the IR spectroscopic data of the compounds 7, 7.5-7.8 the observation of C=S stretching bands at 1203-1217 cm -1 . Valence vibrations of bonds of C-H alkyl groups form bands in area 2935-2850 cm -1 . The synthesized compounds are also characterized by valence vibrations of the C=C bond of the aromatic rings at 1468-1453 cm -1 .
In the 1 H NMR spectra of compounds (7.1-7.4) protons of the S-alkyl fragments resonate in a strong field as a singlet, a triplet or a multiplet in area 3.17-0.97 ppm. Proton of the N=CH fragment forms a signal in the form of the singlet at 8.82-8.71 ppm. The signal in the spectrum of compound 7.8 at 8.09 ppm corresponds to the proton of the CONH fragment and resonates in the form of a singlet.
In the chromatic mass spectra, individual peaks of the molecular ion and peaks of the fragment ions are recorded, which have a high intensity, which confirms the structure and identity of the compounds 7, 7.1-7.8.   The methodology for rational drug development involves the use of molecular docking. Docking experiments of synthesized compounds (7, 7.1-7.8) with the 2XP2 (ALK tyrosine kinase receptor) receptor revealed that compound 7.8 is the most active with a calculated binding energy of 8.1 kcal/mol (Table 1) [3,6,[8][9][10].
The next stage is reaching at the base of the specified disparity of synthesizing compounds to the site of the enzyme's link cyclooxygenase-2 (COX-2) ( Table 2) [9]. Visualization of the interaction of the most active compound (7) with the center of COX-2 allowed to establish that it has a hydrogen bond with the amino acid residue D: TYR 3355, in addition, three pi-alkylhydrophobic interactions with D: LEU 3531, D: 3523, D: 3352.
Analysis of the complex of the most active compound with lanosterol-14α-demethylase showed interactions with the following amino acid residues: В:  Crizotinib -9,4 *E min : The minimum energy of complex formation, kcal/mol.