Rational discovery of peptide inhibitors targeting Keap1: a focus on neurodegenerative diseases

Introduction: The Kelch-like ECH-associated protein 1 (Keap1)/Nuclear factor erythroid 2-related factor 2 (Nrf2) pathway is a promising therapeutic target for neurodegenerative diseases (NDDs). Nrf2 activation mitigates key pathogenic mechanisms in NDDs by enhancing antioxidant defenses, suppressing neuroinflammation, improving mitochondrial function, inhibiting ferroptosis, and maintaining proteostasis. Aim: To design and identify peptide inhibitors capable of disrupting the Keap1/Nrf2 interaction through a rational computational approach. Methods: A combinatorial peptide library comprising 13,167 peptides was generated from an Nrf2-derived template peptide and subjected to rigorous multi-tier screening based on predicted blood-brain barrier permeability, toxicity, allergenicity, antioxidant and anti-inflammatory potential, and physicochemical properties. Following comprehensive safety assessments, 11 promising peptides were shortlisted and evaluated using molecular docking, MM-GBSA binding free-energy calculations, and per-residue energy decomposition analysis. The top-ranked peptide candidates were subsequently subjected to 200 ns molecular dynamics (MD) simulations to assess complex stability and binding behavior through analyses of structural stability, flexibility, principal component analysis (PCA), hydrogen-bond occupancy, and free-energy landscape analyses. Results: Among the shortlisted peptides, LMEETYEFL (pep4369) and LDYETYEFL (pep6649) exhibited stronger binding affinities toward Keap1 than the template peptide in docking and MM-GBSA analyses. MD simulations confirmed the structural stability and favorable conformational behavior of the peptide-Keap1 complexes throughout the simulation period. Conclusion: These findings suggest that LMEETYEFL and LDYETYEFL are promising candidates for disrupting the Keap1/Nrf2 interaction. However, experimental validation is necessary to confirm their therapeutic potential and facilitate further biological evaluation.