Exploring the antimicrobial activity of pantothenamides against uropathogenic Escherichia coli

Metabolic processes required for bacterial pathogens to adapt in the host are potential targets for the development of much-needed new antimicrobial agents. The bacterial central metabolic enzyme 1-deoxy-d-xylulose 5-phosphate synthase (DXPS) is required for the synthesis of essential isoprenoids, thiamin diphosphate (ThDP), and pyridoxal phosphate (PLP) and is believed to function in bacterial adaptations requiring these metabolites. DXPS inhibition impairs a PLP-dependent adaptation of Uropathogenic Escherichia coli (UPEC) to d-serine, a bacteriostatic host metabolite in urine that inhibits pantothenate production in Coenzyme A (CoA) biosynthesis. CoA is required for a functioning tricarboxylic acid (TCA cycle, which is critical for UPEC survival in the urinary tract. Accordingly, inhibition of DXPS also sensitizes UPEC to N-pentylpantothenamide (N5-Pan, 1), an inhibitor of CoA synthesis. However, 1 is enzymatically hydrolyzed by pantetheinases, presenting challenges for its use in in vivo studies. We sought to identify a pantothenamide inhibitor candidate for studies to explore the in vivo efficacy of inhibitor combinations targeting DXPS and CoA syntheses. Here, we describe studies that highlight a truncated analog of 1, pantetheinase-resistant N5-α-Pan (6), as a promising candidate for in vivo studies. Analog 6 exhibited comparable antimicrobial activity with 1 against UPEC grown in nutrient-limited culture conditions, including urine, and displayed enhanced activity in the presence of a DXPS inhibitor. In contrast to 1, analog 6 was stable in the presence of mouse plasma and liver enzymes, making it suitable for studies to investigate an inhibitor combination targeting DXPS and CoA synthesis in UPEC by the in vivo ascending UTI assay.IMPORTANCENew approaches are needed to control the emergence of drug resistance in bacterial pathogens that cause life-threatening infections. Targeting DXPS-dependent synthesis of vitamins is a promising approach to prevent pathogen metabolic adaptation. Metabolic processes requiring DXPS-dependent synthesis of pyridoxal phosphate (PLP) or thiamin diphosphate (ThDP) can become particularly vulnerable in a specific pathogen and/or host environment, under conditions of DXPS inhibition. We previously observed that UPEC grown in urine is particularly sensitive to an inhibitor combination targeting DXPS and CoA synthesis; however, the CoA inhibitor is readily hydrolyzed by a host pantetheinase. This study is significant, as it identifies a pantothenamide inhibitor of CoA synthesis, N5-α-Pan (6), that is stable to mouse plasma and liver enzymes and whose activity is enhanced in the presence of a DXPS inhibitor. Thus, 6 is suitable for studies to explore how a pathogen can become sensitized in vivo under conditions of DXPS inhibition.