Synthesis and in vitro antitumor evaluation of new thieno[2,3-d]pyrimidine derivatives as EGFR and DHFR inhibitors

Abstract

New thienopyrimidine derivatives 2–16 have been synthesized and their in vitro cytotoxicity was evaluated against five different human cancer cell lines HCT-116, Hela, MDA-MB-231, MCF7 and PC3. Compounds 6e, 7a, 7b, 7d, 10c and 10e displayed the highest antitumor activity against all tested cell lines compared to Doxorubicin. Enzyme inhibition assay revealed that compounds 6e and 10e showed high inhibitory activity against EGFR-TK, with IC50 values of 0.133 and 0.151 µM, compared to Olmutinib (IC50 = 0.028 µM); while the highest DHFR inhibitory activity was shown by compounds 7d and 10e with IC50 values of 0.462 and 0.541 µM, compared to Methotrexate (IC50 = 0.117 µM). Cell cycle analysis following a flow cytometric study using colorectal HCT-116 cancer cell line proved that compound 6e induced cell cycle arrest in G0-G1 phase, while compound 10e arrested the cell cycle at both G0-G1 and S phases. Additionally, both compounds (6e and 10e) were potently able to induce apoptosis in HCT-116 cell line. Docking results of compounds 6e and 10e into the pocket of EGFR active site showed their similar main binding features with Olmutinib, while compounds 7d and 10e showed only moderate fitting into DHFR compared to methotrexate. In silico studies revealed that most of the tested compounds obeyed Lipinski’s RO5 and showed positive drug likeness scores.