Design and Synthesis of Benzene Homologues Tethered with 1,2,4-Triazole and 1,3,4-Thiadiazole Motifs Revealing Dual MCF-7/HepG2 Cytotoxic Activity with Prominent Selectivity via Histone Demethylase LSD1 Inhibitory Effect

Abstract

In this study, an efficient multistep synthesis of novel aromatic tricyclic hybrids incorporating different biological active moieties, such as 1,3,4-thiadiazole and 1,2,4-triazole, was reported. These target scaffolds are characterized by having terminal lipophilic or hydrophilic parts, and their structures are confirmed by different spectroscopic methods. Further, the cytotoxic activities of the newly synthesized compounds were evaluated using in vitro MTT cytotoxicity screening assay against three different cell lines, including HepG-2, MCF-7, and HCT-116, compared with the reference drug Taxol. The results showed variable performance against cancer cell lines, exhibiting MCF-7 and HepG-2 selectivities by active analogs. Among these derivatives, 1,2,4-triazoles 11 and 13 and 1,3,4-thiadiazole 18 were found to be the most potent compounds against MCF-7 and HepG-2 cancer cells. Moreover, structure-activity relationship (SAR) studies led to the identification of some potent LSD1 inhibitors. The tested compounds showed good LSD1 inhibitory activities, with an IC₅₀ range of 0.04-1.5 μM. Compounds 27, 23, and 22 were found to be the most active analogs with IC₅₀ values of 0.046, 0.065, and 0.074 μM, respectively. In addition, they exhibited prominent selectivity against a MAO target with apparent cancer cell apoptosis, resulting in DNA fragmentation. This research provides some new aromatic-centered 1,2,4-triazole-3-thione and 1,3,4-thiadiazole analogs as highly effective anticancer agents with good LSD1 target selectivity.

Keywords: 1,2,4-triazoles; 1,3,4-thiadiazoles; LSD1; anticancer activity; apoptosis study; melanoma; tris-substituted analogues.

Figure 1

Reported aromatic planar system targeting LSD1 as anticancer agents.

Scheme 1

Synthesis of tris-acid hydrazide 11 and tris-acid thiosemicarbazides 12–17.

Scheme 2

Synthesis of tris-1,2,4-triazoles 18–23 and tris-1,3,4-thidiazoles 24–28.

Figure 2

(Up),percent of DNA content in compound 27-treated cells versus the control at different cell cycle phases. (Down)percent of apoptotic cells in compound 27-treated cells versus control (total, early, late, and necrosis).

Figure 3

Determination of cell cycle arrest by flow cytometry. Effect of compound 27 on the cell cycle distribution of the MCF-7 cell line.

Figure 4

Evaluation of apoptosis induced by compound 27 treatment. Flow cytometry analysis for Annexin V-FITC/PI staining, Right: compound 27-treated MCF-7 cells compared to Left: control MCF-7 cell.

Figure 5

Statistical analysis of the DNA fragmentation percentage of MCF-7 and HepG2 cells after incubation with compounds 20 and 27 for 24 h IC₅₀ value. The data are reported as the mean ± SD of three independent experiments in triplicate.

Figure 6

(a) 2D binding mode and residues involved in the recognition of cocrystallized ligand (LSD₁ inhibitor) from 5YJB PDB Lys661 and Asp 555. (b) 2D binding mode and residues involved in the recognition of FAD; Try 761, Arg 310, Arg 316, Val 333, Met 332, Glu A801, Ser 209, Glu 309, and Val 590 (with yellow highlight).

Figure 7

Two-dimensional binding mode and residues involved in the recognition of compounds (a) 22, (b) 23, (c) 18, and (d) 27 ligands.

Figure 7

Two-dimensional binding mode and residues involved in the recognition of compounds (a) 22, (b) 23, (c) 18, and (d) 27 ligands.

Figure 8

Surface map for the most active derivative 22 inside an active site.

Figure 9

Surface map for the most active derivative (a) 22 and (b) 27, (c) the least active 18, and (d) the reference compound GSK. Pink, hydrogen bond; blue, mild polar; green, hydrophobic.