Fragment-Based <i>De Novo</i> Discovery of Fourth-Generation EGFR-L858R/T790M/C797S-Tyrosine Kinase Inhibitors Targeting Catalytic Lys745 Residue to Overcome the Osimertinib Resistance

The C797S mutation in epidermal growth factor receptor (EGFR) presents a major resistance mechanism against third-generation inhibitors such as Osimertinib in non-small cell lung cancer (NSCLC). A structure-guided fragment-based de novo design strategy was employed to develop inhibitors targeting the conserved Lys745 residue. A starting library of 87,412 lysine-reactive fragments was filtered using the Rule of Three, resulting in 28,829 drug-like candidates. These were screened through constraint-based molecular docking, identifying 2821 fragments that interacted directly with Lys745. These fragments were recombined using Schrödinger[Formula: see text]s Breed module, yielding 6211 novel compounds, of which Breed 2018, Breed 2066 and Breed 1958 displayed docking scores [Formula: see text]9.00 kcal/mol and formed stable interactions with Lys745 and Met793. Molecular dynamics simulations over 200 ns confirmed complex stability, and MM/GBSA calculations showed binding free energies reaching −217.56 kcal/mol. DFT analysis indicated strong electrophilic potential, while ADME profiling predicted &gt;88% oral absorption, no Lipinski violations and high membrane permeability. The results identify Breed 2018 as a promising scaffold for the development of fourth-generation EGFR inhibitors capable of overcoming C797S-mediated drug resistance.