Discovery and Optimization of Phosphopantetheine Adenylyltransferase Inhibitors with Gram-Negative Antibacterial Activity

Abstract

In the preceding manuscript [ Moreau et al. 2018 , 10.1021/acs.jmedchem.7b01691 ] we described a successful fragment-based lead discovery (FBLD) strategy for discovery of bacterial phosphopantetheine adenylyltransferase inhibitors (PPAT, CoaD). Following several rounds of optimization two promising lead compounds were identified: triazolopyrimidinone 3 and 4-azabenzimidazole 4. Here we disclose our efforts to further optimize these two leads for on-target potency and Gram-negative cellular activity. Enabled by a robust X-ray crystallography system, our structure-based inhibitor design approach delivered compounds with biochemical potencies 4-5 orders of magnitude greater than their respective fragment starting points. Additional optimization was guided by observations on bacterial permeability and physicochemical properties, which ultimately led to the identification of PPAT inhibitors with cellular activity against wild-type E. coli.

Vitamin in the Crosshairs: Targeting Pantothenate and Coenzyme A Biosynthesis for New Antituberculosis Agents

“…While no inhibitors have been reported for MtPPAT using structure-based approaches, inhibitors have been described for EcPPAT, its homologue from E. coli (Zhao et al, 2003;Miller et al, 2010;Moreau et al, 2018;Skepper et al, 2018;Liyanage et al, 2019;Wang et al, 2020). The two homologues share 44% identity and 77% similarity in their amino acid sequences, and both exist as hexamers in their active form (Timofeev et al, 2010).…”

“…While several differences are present between the two structures, such as the number of subunits involved in ligand binding, evaluation of EcPPAT inhibitors can guide future design of compounds to inhibit MtPPAT. Recently, a fragment-based strategy facilitated by crystallography revealed several highly potent compounds (83 and 84, Figure 8C) with nanomolar activity against EcPPAT and P. aeruginosa PPAT (PaPPAT), and moderate cellular activity against the efflux-deficient E. coli DtolC mutant (Moreau et al, 2018;Skepper et al, 2018). Inhibitor development studies of other bacterial PPAT enzymes, including those from S. aureus, S. pneumoniae, and H. pylori have also been performed, but none of these have led to the development of clinical candidates (De Jonge et al, 2013;Cheng et al, 2013).…”

Comprehensive and High‐Throughput Exploration of Chemical Space Using Broadband 19F NMR‐Based Screening

“…[31] As the equivalent human orthologue does not share sequence homology with the bacterial enzyme,C oaD has been evaluated as an antibacterial drug target. [32][33][34] Assessing the entire LEF4000 library in the absence (negative control) and presence of CoaD (binding experiments) required only atotal of 48 hours (comprised of two times 24 hours for the control and binding experiments, respectively). Due to the clear separation of 19 Fs ignals,h it identification could be performed in an automated manner using Mnova Screen within 3.5 hours on astandard Windows PC laptop.…”

Addressing Antimicrobial Resistance through New Medicinal and Synthetic Chemistry Strategies

“…4 ) because of its ability to overlap with compound 23a but to bind differently with the enzyme. 92 Incorporation of a weakly basic amine in the lead structures led to enhanced gram-negative permeability (compound 24 ; Fig. 4 ).…”

“…4 ) represent the first known bacterial PPAT inhibitors with cellular activity against wilt-type gram-negative bacteria. 92 However, these series of compounds has not been developed further, because of the lack of sufficient potency against other gram-negative species.…”