Expanding the chemical and therapeutic landscape of 5H-Indeno[1,2-b]pyridin-5-one derivatives: Novel anticancer activity, EGFR inhibition, and modulation of HIF-VEGF and PI3K/AKT/mTOR pathways supported by computational insights.

A novel series of eleven 5H-indeno[1,2-b]pyridin-5-one derivatives (1-11) was rationally designed and synthesized to explore their anticancer potential. The compounds were screened for cytotoxicity against PC-3, HePG-2, and MCF-7 cancer cell lines, and the normal WI-38 line using the MTT assay. Among them, compounds 5 and 7 exhibited the strongest cytotoxicity, with compound 7 showing remarkable potency toward MCF-7 cells (IC₅₀ = 5.53 μM). Target prediction and enrichment analyses identified EGFR as the most relevant molecular target associated with the observed activity. Compound 7 significantly inhibited EGFR (IC₅₀ = 0.12 ± 0.004 μg mL), comparable to erlotinib, confirming its mechanism-based action. Flow cytometry analysis revealed G₂/M cell-cycle arrest and a marked increase in apoptotic cell populations, indicating apoptosis-mediated cytotoxicity. Furthermore, gene expression analysis of MCF-7 cells treated with compound 7 demonstrated a significant down-regulation of HIF-1α and VEGF genes, accompanied by modulation of PI3K, AKT, mTOR, and PTEN expression levels, suggesting suppression of the HIF-VEGF signaling crosstalk that governs tumor angiogenesis and proliferation. Molecular docking against EGFR (PDB ID: 4I23) showed a strong binding affinity (-7.85 kcal/mol) through hydrogen bonding with Met793 and hydrophobic interactions with Lys745 and Gly796, consistent with the biological findings. DFT analysis (ΔE = 0.069 eV; η = 0.0347) supported the molecule's high reactivity and optimal donor acceptor features. Molecular dynamics confirmed stable complex formation with favorable flexibility, while ADMET profiling indicated acceptable drug-likeness but limited oral absorption and potential metabolic liabilities. Overall, compound 7 emerged as the most promising EGFR-targeted lead with strong selectivity toward MCF-7 cells, induction of apoptosis, inhibition of HIF-VEGF signaling, and favorable electronic behavior, highlighting its ability to modulate the EGFR-PI3K/AKT-HIF-VEGF axis as a potential multi-pathway strategy for breast cancer therapy.