PK 11195 derivatives: exploring the influence of amide and heterocyclic substitution on A147T TSPO discrimination

The translocator protein (TSPO) is an 18 kDa protein on the outer mitochondrial membrane. It has gained significant interest in recent years for its potential as a therapeutic target and imaging biomarker, particularly in neuroinflammation, cancer, and central nervous system disorders (CNS). Clinical translation of ligands has been complicated by the presence of a common single nucleotide polymorphism (A147T TSPO), at which many disclosed TSPO ligands lose affinity. One exception is the first-generation TSPO ligand PK 11195, however, this ligand possesses unfavourable pharmacokinetic properties limiting translation in CNS applications. We aimed to investigate which motifs of this ligand contribute to high binding at the wild type (WT) and A147T TSPO isoforms with the aim of identifying elements that may tolerate lipophilicity-reducing substitutions. Affinities for a small library of isoquinoline, quinazoline, indole and azaindole carboxamides with varying aliphatic and aromatic substituents were measured using radioligand binding at both TSPO isoforms, with computational studies performed to rationalise the experimentally measured binding. The heterocycle and acetamide substituents of PK 11195 were found to play a role in its non-discriminating nature. In addition, the study yielded 2a, a high affinity, non-discriminating TSPO ligand. Modelling suggests its high affinity and lack of discrimination results from additional π-π interactions introduced at the binding site of both TSPO isoforms. These findings provide a foundation for developing TSPO ligands with improved clinical properties and insensitivity to A147T polymorphism.