Synthesis, anti-HIV activity and molecular modeling study of 3-aryl-6-adamantylmethyl-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole derivatives

2.1 In vitro anti-HIV activity

Compounds 6a–l were tested for their inhibitory activity against HIV-1 (strain III_B) and HIV-2 (strain ROD) I human MT-4 cell cultures by the MTT assay [32, 33]. The results are compiled in Table 3, where the data for nevirapine [34] were included for comparison purposes. According to the results, four analogs, namely 6a, 6b, 6d and 6g, inhibited the replication of HIV-1 and HIV-2 with EC₅₀ values of >11.50, >14.90 and >12.40 μg mL^–1, respectively, but no selectivity could be witnessed. From the SAR analysis, we found that the halogen residues on the aromatic ring at ortho-position, e.g. in 6d and 6g or alkyl group at ortho- or meta-position as in 6a and 6b, were well tolerated in the hydrophobic region of HIV reverse transcriptase and then showed higher activity than those of the derivatives with other substituents of the same series of Scheme 1. However, the anti-HIV activity and the selectivity of these compounds are too limited to perform extensive mode-of-action studies, and 6b might be considered as a new lead in the development of antiviral agents as a non-nucleoside reverse transcriptase inhibitor.

2.2 In vitro antimicrobial activity

Compounds 6a–l were tested for their antimicrobial potential against different fungal and bacterial strains. Six fungal strains, namely Trichphyton longifusus, Candida albicans, Aspergillus flavus, Microsporum canis, Fusarium solani and Candida glabrata, were used to study antifungal activity. Antibacterial activity was evaluated against six bacterial strains, i.e. Escherichia coli, Bacillus subtilis, Shigella flexneri, Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella typhi. The activities of the synthesized compounds were found to be non-significant at non-toxic concentrations against the tested fungi and bacteria.

2.3 Molecular modeling study

The molecular docking was performed using Sybyl-X 1.1 [35] and the docking results were shown by PyMOL [36]. In the docking study, X-ray crystal structure of HIV reverse transcriptase (RT) enzyme (PDB: 1dtt) was obtained from Protein Data Bank (http://www.rcsb.org) [37].

Compound 6g has been selected for the docking modeling study, since its binding energy score is –9.2, indicating a selectivity of adamantyl-triazolothiadiazole 6g in its binding to the enzyme pocket (Fig. 3). Detailed analysis of the binding mode showed that the aromatic ring of 6g points toward the aromatic ring of the Tyr179 residue apparently developing π–π stacking interactions with the two residues. The triazolothiadiazole backbone is located in the middle of the binding pocket, anchoring the nitrogen atom N-3 and the sulfur atom of 6g in a favorable position for hydrogen bonding with Lys101 and Lys100, respectively, of the reverse transcriptase enzyme. Overall, the combination of hydrophobic interaction and π stacking appears to govern the binding of 6g with HIV reverse transcriptase.

Fig. 3:

Docked conformation of 6g showing two hydrogen bondings: Lys101 with N3 of the thiadiazole ring and Lys100 with the sulfur atom of the same ring. It also exhibited hydrophobic interactions between the phenyl group at C-3′ of the triazole moiety and Tyr179 of the reverse transcriptase enzyme.

4.1 General

Melting points of the synthesized compounds were measured in open capillaries using Stuart SMP3 melting point apparatus (Barloworld Scientific Ltd, Staffordshire, UK) and are uncorrected. The IR spectra were recorded on a Schimadzu Fourier Transform Infrared spectrophotometer (Model 270, Japan). The ¹H and ¹³CNMR spectra were recorded on a Bruker Avance 300 MHz NMR spectrometer (Bruker, Reinstetten, Germany) and signals calibrated to the residual solvent signals. The EI mass spectrometry was carried out using Agilent Technologies 6890N (GC) with an inert selective detector 5973 mass spectrometer (Agilent Technologies, San Diego, USA). The reagents used were of analytical grade, while the solvents were purified before use.

4.2 General procedure for the synthesis of 3-aryl-6-adamantylmethyl-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazoles 6a–l

A mixture of 4a–l (0.90 mmol) and (adamant-1-yl)acetic acid 5 (175 mg, 0.90 mmol) in phosphorus oxychloride (5 mL) was heated under reflux for 4 h. The reaction mixture was cooled to room temperature, poured into crushed ice and neutralized with solid potassium carbonate and potassium hydroxide until the pH was 8. The precipitated products were filtered, washed with excess water and purified by silica gel column chromatography, using n-hexane-ethyl acetate (7:3) as eluent to give the desired products 6a–l.

4.3 Crystal structure determination of 6a

Suitable single crystals of compound 6a were obtained as colorless blocks from chloroform. Data were collected on a Stoe Image Plate Diffraction System diffractometer with graphite-monochromatized MoK_α radiation (λ = 0.71073 Å). The structure was solved by Direct Methods and refined by full-matrix least-squares procedures on F² with the Shelxs/l-97 software package. Hydrogen atoms were placed in idealized positions and included as constrained into the refinement, while all other atoms were refined with anisotropic displacement parameters.

Crystal structure data: C₂₁H₂₄N₄S, M_r = 364.50, colorless plates, crystal dimensions = 0.38 × 0.27 × 0.27 mm³, monoclinic space group P2₁, a = 6.7727(5), b = 11.0272(9), c = 12.2678(8) Å, V = 900.04 (12) ų, β = 100.78(8)°, Z = 2, D_calcd. = 1.345 mg m^–3, μ(MoK_α) = 0.193 mm^–1, F(000) = 388 e, T = 173 K. 6869 measured refl., hkl range ± 8, ± 13, ± 15, θ_max = 26.0°, 3433 unique refl., R_int = 0.022, 237 ref. parameters, R/wR (all data) = 0.0276/0.0721, GoF = 1.096, x(Flack) = 0.02(5), Δρ_fin (max/min) = 0.25/–0.19 e Å^–3.

CCDC 940854 contains the supplementary crystallographic data for this paper. These data can be obtained free of charge from the Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/datarequest/cif.