A potent nonapeptide inhibitor for the CXCL12/HMGB1 heterocomplex: A computational and experimental approach

Inflammation is a vital defense mechanism activated in response to injury or infection, which induces the release of cytokines and chemokines to promote tissue repair. However, persistent inflammation may result in the development of autoimmune diseases. During pathological conditions such as rheumatoid arthritis, HMGB1 is kept in its reduced isoform and can complex with CXCL12 enhancing cell migration and exacerbating the immune responses. Small organic compounds selective for HMGB1 have been previously reported to be able to disrupt the CXCL12/HMGB1 heterocomplex, but due to their low affinity, they are unsuitable for further development as novel anti-inflammatory drugs. We previously reported a peptide (HBP08) that binds to HMGB1 with high affinity (K_d = 0.8 µM), and blocks the activity of the heterocomplex, in line with a wide literature that supports the use of peptides to design protein-protein interaction inhibitors. In the present work, we computationally optimized the HBP08 peptide sequence, finding new analogues endowed with improved affinity for HMGB1. In particular, HBP08-2 inhibited the activity of the CXCL12/HMGB1 heterocomplex with an IC₅₀ 15-fold lower (3.31 µM) and displayed a K_d 28-fold lower (28.1 ± 7.0 nM) than the parent peptide HBP08. HBP08-2 may represent a valuable tool to investigate CXCL12/HMGB1-driven inflammatory pathways and to develop peptidomimetics with optimized pharmacokinetic profiles, thereby offering a promising strategy for the treatment of chronic inflammatory diseases, such as rheumatoid arthritis, where the CXCL12/HMGB1 heterocomplex plays a pivotal pro-inflammatory role.