Targeting Conformational Flexibility of Reactive Intermediate to Enhance Selectivity of a GABA Aminotransferase Inactivator

Currently, mechanism-based inactivators (MBIs) are the only available therapeutic option to target γ-aminobutyric acid aminotransferase (GABA-AT). However, off-target activity against homologous enzymes has posed major challenges for the clinical use of MBIs. For example, CPP-115, a MBI of GABA-AT that completed a Phase I clinical trial, also inactivates ornithine aminotransferase (OAT). Here, we present an OAT-specific inactivation mechanism for CPP-115, enabled by the dynamic motion of a key reactive intermediate. Based on this, to selectively reduce the off-target activity, a proof-of-concept molecule that regulates the intermediate conformational flexibility was designed and synthesized. The resulting inactivator achieved the greatly enhanced GABA-AT selectivity over OAT and demonstrated therapeutic efficacy in an inflammatory pain animal model. Our strategy in this study, targeting dynamics of a reactive intermediate based on precise mechanistic understanding, could serve as a generalizable design principle for fine-tuning the selectivity of MBIs, particularly for other aminotransferases.