A structure-guided fragment-based approach for the discovery of allosteric inhibitors targeting the lipophilic binding site of transcription factor EthR

Abstract

A structure-guided fragment-based approach was used to target the lipophilic allosteric binding site of Mycobacterium tuberculosis EthR. This elongated channel has many hydrophobic residues lining the binding site, with few opportunities for hydrogen bonding. We demonstrate that a fragment-based approach involving the inclusion of flexible fragments in the library leads to an efficient exploration of chemical space, that fragment binding can lead to an extension of the cavity, and that fragments are able to identify hydrogen-bonding opportunities in this hydrophobic environment that are not exploited in Nature. In the present paper, we report the identification of a 1 μM affinity ligand obtained by structure-guided fragment linking.

Harnessing Allostery: A Novel Approach to Drug Discovery

“…These data suggest that the chemical leads identified using the FBS method provide new opportunities to develop new antiviral drugs. In fact, this method has been demonstrated to be successful to elucidate new allosteric sites on a number of proteins, including HIV protease and integrase, glycogen phosphorylase, oncogenic Ras protein, and transcription factor EthR …”

Fragment Screening against the EthR–DNA Interaction by Native Mass Spectrometry

“…Subsequent fragment elaboration furnished analogues that disrupted the EthR–DNA interaction and boosted ETH activity in Mtb‐infected macrophages with submicromolar potency . Our group has previously employed DSF to screen a 1250 member fragment library against EthR, with fragments showing Δ T m ≥1 °C at 10 m m concentration being classed as hits . Fragment‐merging and ‐linking strategies subsequently generated improved analogues with low‐micromolar IC 50 values against the EthR–DNA interaction …”

“…Our group has previously employed DSF to screen a 1250 member fragment library against EthR, with fragments showing Δ T m ≥1 °C at 10 m m concentration being classed as hits . Fragment‐merging and ‐linking strategies subsequently generated improved analogues with low‐micromolar IC 50 values against the EthR–DNA interaction …”

Fragment-Sized EthR Inhibitors Exhibit Exceptionally Strong Ethionamide Boosting Effect in Whole-Cell Mycobacterium tuberculosis Assays

“…19 The presence of fragment 1 at a concentration of 1 μM was shown to reduce the minimum inhibitory concentration (MIC) of ethionamide from 15 μM to approximately 2 μM under the conditions of the resazurin reduction microplate assay used. 19 An X-ray crystal structure of fragment 1 bound to EthR (Figure 2) was obtained that provided a good starting point for the design of a small focused library of compounds containing hydrogen-bond donor or acceptor groups at appropriate positions within the scaffold of compound 1 . The distances between the primary amide of Asn176 and position X of 1 , and between the hydroxyl oxygen atom of Thr149 and position Y of 1 , are shown in Figure 2.…”