Macrocyclic FKBP51 Ligands Define a Transient Binding Mode with Enhanced Selectivity

Abstract

Subtype selectivity represents ac hallenge in many drug discovery campaigns.Atypical example is the FK506 binding protein 51 (FKBP51), whichh as emerged as an attractive drug target. The most advanced FKBP51 ligands of the SAFit class are highly selective vs.F KBP52 but poorly discriminate against the homologs and off-targets FKBP12 and FKBP12.6. During am acrocyclization pilot study,w eo bserved that many of these macrocyclic analogs have unanticipated and unprecedented preference for FKBP51 over FKBP12 and FKBP12.6. Structural studies revealed that these macrocycles bind with anew binding mode featuring atransient conformation, which is disfavored for the small FKBPs.Using aconformation-sensitive assayweshow that this binding mode occurs in solution and is characteristic for this new class of compounds.T he discovered macrocycles are non-immunosuppressive,e ngage FKBP51 in cells,a nd blockt he cellular effect of FKBP51 on IKKa.O ur findings provide an ew chemical scaffold for improved FKBP51 ligands and the structural basis for enhanced selectivity.

Picomolar FKBP inhibitors enabled by a single water-displacing methyl group in bicyclic [4.3.1] aza-amides

“…To explore the thermodynamic signature for the enhanced affinity of α-methyl-containing bicyclic [4.3.1] aza amides we used isothermal titration calorimetry (ITC) 13,31 (Table S4 † ). The affinities measured with ITC were fully consistent with the FP-assay data (Fig.…”

Fenton-Chemistry-Based Oxidative Modification of Proteins Reflects Their Conformation

“…Binding affinity (K i ) to FKBP12 is approximately 163 nM for SAFit1 and 1.8 nM for FK[4.3.1]-16h. Binding affinity to FKBP51 is approximately 4 nM for SAFit1 and 57 nM for FK[4.3.1]-16h [ 41 , 49 ].…”

“…An alternative, more intriguing explanation for this behaviour than a simple steric effect involves the fact that binding of SAFit1 to FKBP51FK1 requires the side chain of residue Phe67 to be displaced, and this alternative conformation is at the core of the ability of this type of ligand to distinguish between homologues [ 40 ]. The binding affinity of SAFit1 to FKBP12, where Phe36 is the counterpart to Phe67 in FKBP51FK1 (see Figure 6 ), is an order of magnitude lower than to FKBP51FK1 [ 41 , 49 ]. Interestingly, in both cases the key phenylalanine residue is part of a β-sheet, with the adjacent strand (labelled as ‘βB’ in Figure 6 and Figure 8 ) composed of residues 21–30 in FKBP12 and residues 52–61 in FKBP51FK1.…”

“…Of note is the protection of the peptide with residues 89–98 in FKBP51FK1 and, similarly, that with residues 53–71 in FKBP12. These span an α-helix (αB) containing, or being close to, a key interacting amino acid residue (Ile56 in FKBP12 and Ile87 in FKBP51FK1) that forms a strong hydrogen bond with both ligands through its amide nitrogen atom [ 49 , 52 , 53 , 54 ]. Furthermore, a very reactive tryptophan (based on both the inherent reactivity of the side chain toward oxidative labelling, and the direct observation, as shown in Figure 8 C) that is located within the binding pocket and directly exposed to solvent in the absence of a ligand is located in this region.…”

Frontiers in Medicinal Chemistry 2022 Goes Virtual

“…This approach allowed for addressing FKBP51, a potential target for depression, obesity and chronic pain with high selectivity over FKBP52, which has opposite biological functions but an almost identical ligand binding pocket. Moreover, one of the series gave rise to compounds with a transient binding mode conferring an unprecedented selectivity against the homologs FKBP12 and FKBP12.6 [45b] …”