Fragment-Based De Novo 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 were filtered using the Rule of Three, resulting in 28,829 drug-like candidates. These were screened through constraint-based molecular docking, identifying 2,821 fragments that interacted directly with Lys745. These fragments were recombined using Schrödinger's breed module, yielding 6,211 novel compounds, of which Breed 2018, Breed 2066, and Breed 1958 displayed docking scores ≤ -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 >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.