Novel macrocyclic peptidomimetics targeting the insulin-regulated aminopeptidase (IRAP): design, synthesis and evaluation

Abstract

Inhibition of the insulin-regulated aminopeptidase (IRAP) is a promising therapeutic strategy for neurodegenerative disorders such as Alzheimer's disease, due to its role in cognitive processes. HA08, a macrocyclic peptidomimetic derived from angiotensin IV, is among the most potent known IRAP inhibitors (IC₅₀ = 18 nM). However, detailed structure–activity relationship (SAR) studies at its C-terminus have been limited by synthetic constraints. Herein, we report the design, synthesis, and biological evaluation of a focused series of HA08 analogues to explore the impact of C-terminal modifications on IRAP inhibition. An improved divergent synthetic route was established via a common macrocyclic intermediate, enabling late-stage diversification through coupling with non-natural amino acids which led to the synthesis of twelve novel peptidomimetic scaffolds. Several analogues retained high potency, with one-carbon elongation next to the carboxylic acid moiety or secondary amine being well tolerated. In contrast, aliphatic analogues exhibited markedly reduced potency, highlighting the importance of π–π interactions, while the low activity of phenoxyacetic acid derivatives likely reflects altered geometry within the binding pocket. The most potent inhibitor in the series featured a C-terminal benzyl alcohol (IC₅₀ = 59 nM), approaching the activity of HA08. To rationalise these SAR trends, molecular dynamics simulations were performed based on the IRAP–HA08 co-crystal structure. Partial least squares analysis of protein–ligand contact patterns revealed that sustained interactions between the C-terminal carboxylate and Arg929 correlated with lower potency, whereas interaction with Arg439 was associated with enhanced activity. These findings suggest that subtle shifts in C-terminal positioning influence binding mode and potency and provides valuable insights for the design of future IRAP inhibitors.