Novel N-Arylmethyl-aniline/chalcone hybrids as potential VEGFR inhibitors: synthesis, biological evaluations, and molecular dynamic simulations

Abstract

Significant advancements have been made in the domain of targeted anticancer therapy for the management of malignancies in recent times. VEGFR-2 is characterised by its pivotal involvement in angiogenesis and subsequent mechanisms that promote tumour cells survival. Herein, novel N-arylmethyl-aniline/chalcone hybrids 5a-5n were designed and synthesised as potential anticancer and VEGFR-2 inhibitors. The anticancer activity was evaluated at the NCI-USA, resulting in the identification of 10 remarkably potent molecules 5a-5j that were further subjected to the five-dose assays. Thereafter, they were explored for their VEGFR-2 inhibitory activity where 5e and 5h emerged as the most potent inhibitors. 5e and 5h induced apoptosis with cell cycle arrest at the SubG₀-G₁ phase within HCT-116 cells. Moreover, their impact on some key apoptotic genes was assessed, suggesting caspase-dependent apoptosis. Furthermore, molecular docking and molecular dynamics simulations were conducted to explore the binding modes and stability of the protein-ligand complexes.

Keywords: VEGFR-2; anticancer agents; apoptosis induction; molecular docking.

Figure 1.

Representative examples of FDA-approved VEGFR inhibitors.

Figure 2.

Examples of reported chalcone VEGFR inhibitors.

Figure 3.

New synthesised compound rational design.

Scheme 1.

Synthesis of target derivatives 5a–5n

Figure 4.

Cell line sensitivity to compounds 5a–5n. The average of growth inhibition (GI) obtained with all compounds of the series was calculated for each cell line of the panel. Red bars correspond to an average of GI greater than 100%.

Figure 5.

The dose response curves of NCI full panel for compound 5h (the graphs of the remaining compounds are presented in the supplementary data).

Figure 6.

Structure–activity relationship for compounds 5a–5n.

Figure 7.

Contour plots measuring the percentage of viable (LL), early apoptotic (LR), late apoptotic (UL), and necrotic cells (UR) by AV/PI assay using flow cytometry. The assay was performed after the treatment of HCT-116 (colon cancer) for 24h with doxorubicin, 5e and 5h compared to 0.1% DMSO negative control.

Figure 8.

Histograms for cell cycle analysis measuring the percentage of SubG₀-G₁, G₀-G₁, S-, and G₂M phases by PI assay using flow cytometry. The assay was performed after the treatment of both HCT-116 (colon cancer) for 24h with doxorubicin, 5e and 5h compared to 0.1% DMSO negative control. Data represented as mean ± standard error of the mean (SEM), n = 3.

Figure 9.

Measurement of caspase-3 (Casp-3) and Bax protein levels after treatment of compounds 5e and 5h in HCT-116 cancer cells compared to 0.1% DMSO (negative control). The cells were treated with the IC₅₀ concentrations of compounds for 24h and data were shown as mean ± SEM of three independent experiments (n = 3).

Figure 10.

Docking interaction (2D) with the VEGFR-2 enzyme for (A) Sorafenib, (B) 5e, and (C) 5h.

Figure 11.

Docking interaction (3D form) for compounds (A) 5e and (B) 5h.

Figure 12.

Molecular dynamic simulation of two stable complexes of 5h-VEGFR-2 and Sorafenib-VEGFR-2; (A) RMSD analysis for the MD simulations of 5h–VEGFR-2 and sorafenib–VEGFR-2, in addition to the Apo form of VEGFR-2, (B) RMSF analysis for the MD simulations of 5h–VEGFR-2 and Sorafenib–VEGFR-2, in addition to the Apo form of VEGFR-2.