Pan-cancer immunogenomics and structure-guided repurposing identify N-Acetyl-L-carnosine as a PAR2 binding modulator

Protease Activated Receptor 2 (PAR2/F2RL1) is a G protein coupled receptor, activated by proteolytic cleavage of its N-terminus tethered peptide. Its role in shaping the tumor immune microenvironment across different cancers remains poorly defined. This study presents an integrated immunogenomic and drug discovery approach to evaluate PAR2's role across 33 cancer types. Pan-cancer analysis revealed significant upregulation of PAR2 in eight tumor types, head and neck squamous cell carcinoma (HNSC) and lung adenocarcinoma (LUAD) showed high immunosuppressive signatures and poor prognostic through depletion of adaptive immune cells and enrichment of alternative checkpoint pathways. These findings suggest PAR2 as a viable target for reversing tumor immune suppression specially in PAR2 over expressing HNSC and LUAD. A structure based virtual screen of 5,954 compounds identified N-Acetyl-L-carnosine (NAC), a clinically used antioxidant not previously studied for GPCR binding as an orthosteric PAR2 binder. Molecular docking and molecular dynamics simulations showed that NAC forms a stable and energetically favourable binding with PAR2 at key residues (TYR326, ASP228), similar to the known antagonist AZ8838. NAC showed a predicted binding energy of -46.08 kcal/mol compared to -42.75 kcal/mol for the known antagonist AZ8838. MTT assay showed no toxicity with NAC treatment or in combination with the PAR2 activator SLIGKV. In wound-healing assays, NAC increased migration at higher concentrations and enhanced SLIGKV-induced motility, indicating that NAC does not inhibit PAR2 activity but may act as a context dependent modulator. Together, these results identify PAR2 as an immunotherapeutic target for cancers of HNSC and LUAD, and highlights NAC as a PAR2-binding molecule whose functional impact on PAR2 signalling requires further validation through direct receptor-signalling assays.