New modified thieno[2,3-d]pyrimidine derivatives as VEGFR-2 inhibitors: Design, synthesis, in vitro anti-cancer evaluation and divers in silico studies

Angiogenesis is a critical regulatory mechanism for tumor development and metastasis. Tumor growth is closely related to the production of the vascular endothelial growth factor (VEGF), which is mediated by the cell-surface kinase VEGFR-2. Accordingly, blocking of VEGFR-2 is an important approach to find new treatments for malignancies. The aim of this research is the synthesis of new thieno[2,3-d]pyrimidine derivatives that could be potential anticancer leads inhibiting VEGFR-2. The prepared thieno[2,3-d]pyrimidine derivatives were tested in vitro for their inhibitory effects against the kinase activity of VEGFR-2. In addition, the synthesized compounds were assessed for their anti-proliferative activities against MCF-7 and HepG2 cell lines. Compound 4c displayed the strongest anti-VEGFR-2 potentiality with an IC50 value of 0.15 mu M and exhibited good anti-proliferative effects against HepG2 and MCF-7 cells with IC50 of 17.14 and 11.56 mu M, respectively. Compound 4c induced cell cycle arrest at the S phase and boosted early and late apoptosis in MCF-7 cells. Additionally, compound 4c increased BAX (4.5-fold), decreased Bcl-2 (3.5-fold), and demonstrated a notable increase in caspase-8 (2.9-fold) and caspase-9 (3.3-fold) levels. Furthermore, compound 4c significantly reduced TNF-alpha (3.6-fold) and IL-6 (3.2fold) levels in MCF-7 cells. Molecular docking studies indicated good binding affinities as well as energies of the thieno[2,3-d]pyrimidine derivatives against the VEGFR-2. Molecular dynamics (MD) simulations were utilized to study the structural, energetic, and conformational changes of the VEGFR-2-4c complex, and the results indicated its stability. The MM-GBSA study of the VEGFR-2-4c complex showed a stable thermodynamic behavior with a binding free energy of -44 kcal/mol. The PLIP analysis identified the 3D interactions and binding conformation through the VEGFR-2-4c complex. Moreover, the DFT studies have been performed to study, Mullikan atomic charge distribution, FMO, ESP, the total density of state, and the QTAIM maps of compound 4c to theoretically verify its reactivity. Finally, computational ADMET and toxicity studies were conducted to assess the drug development potential of the thieno[2,3-d]pyrimidine derivatives.