Novel chalcone 2-thiopyrimidine conjugates as dual VEGFR-2/BRAF inhibitors: design, synthesis, in vitro cytotoxicity, and molecular docking study

Abstract

Chalcone-based derivatives have shown potential anticancer activity via multiple mechanisms including protein kinase inhibition. In the current study, two series of chalcone/2-thiopyrimidine conjugates 4a–4d and 6a–6i were designed, synthesized and screened for their antiproliferative activity in a single-dose assay against NCI-60 cancer cell lines. Ten compounds, 4a–4d, 6a–6c, 6f, 6h, and 6i, were selected for a five-dose assay and their GI₅₀ values were determined. Compound 4c showed potent anticancer activity against LOX IMVI melanoma cell line with a GI₅₀ value of 0.0128 μM. Seven compounds, 4a, 4c, 4d, 6c, 6f, 6h, and 6i, were found to be non-cytotoxic against fibroblast (hFB) normal cell line. Additionally, investigation of the VEGFR-2 inhibitory activity of the ten promising compounds revealed that 4c, 4d and 6i displayed promising VEGFR-2 inhibition (IC₅₀ = 0.144, 0.105, and 0.072 μM, respectively) compared to sorafenib (IC₅₀ = 0.081 μM). Moreover, 4c inhibited BRAF_WT and BRAF_V600E kinases (IC₅₀ = 0.201 and 0.101 μM, respectively) relative to vemurafenib (IC₅₀ = 0.156 and 0.063 μM, respectively). Furthermore, 4c arrested the cell cycle progression at the G₁ phase and induced late apoptosis in LOX IMVI cells. Moreover, evaluation of the effect of 4c on apoptotic markers in the mentioned cells indicated an increase in the Bax/Bcl-2 ratio by 28.12-fold along with upregulation of caspases-3 and -9 by 7.40- and 5.63-fold, respectively, in addition to anti-migratory effect. Molecular docking study of the most promising derivatives revealed a common binding pattern in the binding site of the target kinases that extends from the hinge region through the gate area towards the allosteric back pocket interacting with the key amino acids in a type II inhibitor-like binding pattern.