The epidermal growth factor receptor (EGFR) is a common diver gene for lung cancer (NSCLC), which leads to an increasing death rate worldwide. This study reports the design, synthesis, and biological evaluation of triazole-clubbed pyrimidine derivatives (RDa–RDm) as potential anticancer agents. Thirteen compounds were synthesized and screened against the A549 lung cancer cell line. RDg emerged as the most potent derivative, exhibiting an IC50 of 15.70 µM, compared with the standard drug erlotinib (IC50 = 10.10 µM). Notably, all derivatives displayed moderate to excellent anticancer activity at 100 µM, with IC50 values ranging from 15.70 to 88.27 µM. RDg, characterised by a 4-chlorophenyl group, demonstrated strong in vitro activity and induced cell-cycle arrest at the sub-G0 phase. In vivo study using the Ehrlich ascites carcinoma (EAC) mouse model confirmed the superior anticancer efficacy of RDg. At a 5 mg/kg dose, RDg achieved a 52% reduction in tumour volume and 54% reduction in tumour weight compared with erlotinib 26% tumour volume reduction. Furthermore, RDg demonstrated a 90% tumour inhibition rate compared with erlotinib 75%, attributed to its enhanced cellular uptake and sustained release properties. In silico analyses provided insights into RDg mechanism of action, revealing strong interactions with EGFR binding sites, including hydrogen bonding with Met-793 and π-sulphur interaction with Met-790. Molecular dynamics simulations demonstrated RDg stabilising effect on EGFR, as evidenced by reduced protein flexibility and compact conformational space. The combination of promising in vitro, in vivo and in silico results showed RDg may be used as a lead compound for further development of novel compounds as EGFR inhibitors.
