Beyond Antibodies: Leveraging Trastuzumab Structural Insights for Next-Generation HER۲-Positive Cancer Peptide Therapeutics

Trastuzumab (Herceptin) is a humanized monoclonal antibody targeting HER۲ receptor, approved for HER۲-positive breast cancer treatment. Despite clinical success, antibody therapeutics face limitations including high production costs, poor tissue penetration, and immunogenicity. Peptide-based therapeutics offer advantageous pharmacokinetic profiles while maintaining target specificity. Understanding molecular determinants of Trastuzumab-HER۲ recognition is essential for rational peptide design. This study aimed to identify critical binding hotspots in the Trastuzumab-HER۲ interface using computational alanine scanning mutagenesis, establishing a foundation for developing novel HER۲-targeting peptides that mimic the functional paratope of Trastuzumab with improved druglike properties.Methods (۱۵۰ words)The crystal structure of Trastuzumab Fab-HER۲ extracellular domain complex (PDB: ۱N۸Z, ۲.۵۲ Å resolution) served as template. Interface residues were identified and analyzed using PDBsum. Five CDR residues were selected for alanine substitution: Y۹۲ (CDR-L۳), R۵۰, R۵۹ (CDR-H۲), and W۹۹, Y۱۰۵ (CDR-H۳). Computational mutagenesis and protein-protein docking were performed using HADDOCK ۲.۴ with default parameters. The top-ranked clusters were analyzed, and mutant-derived complexes were further evaluated using PDBsum for interface characterization. Delta values (Mutant−WT) were calculated for HADDOCK scores and energy components including van der Waals, electrostatic, and desolvation energies. Identified hotspot residues and their spatial arrangements will guide peptide scaffold design using a hybrid approach combining structure-based modeling with artificial intelligence-driven sequence optimization. Molecular dynamics simulations will subsequently validate designed peptide-HER۲ complexes for stability and binding persistence.Results Wild-type Trastuzumab-HER۲ achieved HADDOCK score of -۱۳۵.۶±۲.۰with RMSD ۰.۷±۰.۵Å. All mutations resulted in binding destabilization. Y۹۲A demonstrated the largest impact (ΔScore = +۱۹.۸) with substantial RMSD increase to ۳.۵±۲.۱Å and significant desolvation penalty (ΔDesolv = +۱۲.۸), indicating a key role in maintaining binding geometry. R۵۰A exhibited the most pronounced electrostatic penalty (ΔElec = +۱۱۱.۶, ΔScore = +۱۶.۲), suggesting involvement in important charge-mediated interactions. Y۱۰۵A (ΔScore = +۱۶.۳) showed combined electrostatic (ΔElec = +۴۶.۱) and desolvation (ΔDesolv = +۶.۶) contributions. R۵۹A contributed moderately (ΔScore = +۱۳.۲) primarily through electrostatic interactions. Notably, W۹۹A showed minimal binding perturbation (ΔScore = +۴.۸) with favorable van der Waals adjustment (ΔVdW = −۸.۵), suggesting this position may tolerate modification. These results identify Y۹۲, R۵۰, and Y۱۰۵ as priority targets for peptide design.Conclusion Computational alanine scanning identified Y۹۲ (CDR-L۳) as a primary binding hotspot associated with significant geometric rearrangement upon mutation, while R۵۰ (CDR-H۲) serves as a key electrostatic contributor. The relative tolerance of W۹۹ mutation suggests CDR-H۳ may accommodate sequence modifications, providing design flexibility. These findings establish a minimal functional paratope comprising Y۹۲, R۵۰, and Y۱۰۵ as priority pharmacophoric elements for HER۲-targeting peptide development. Subsequent molecular dynamics validation and AI-assisted sequence optimization will enable rational design of constrained peptides mimicking Trastuzumab’s binding mechanism with enhanced pharmacokinetic properties

کلیدواژه ها:

HER۲-positive breast cancer ، Trastuzumab ، Peptide drug design ، Alanine scanning mutagenesis ، HADDOCK