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1.
Microbiol Spectr ; 10(2): e0243021, 2022 04 27.
Article in English | MEDLINE | ID: mdl-35297652

ABSTRACT

CpxRA is an envelope stress response system that is highly conserved in the Enterobacteriaceae. CpxA has kinase activity for CpxR and phosphatase activity for phospho-CpxR (CpxR-P), a transcription factor. In response to membrane stress, CpxR-P is produced and upregulates genes involved in membrane repair and downregulates genes that encode virulence factors that are trafficked across the cell membrane. Mutants that constitutively activate CpxRA in Salmonella enterica serovar Typhimurium and in uropathogenic Escherichia coli (UPEC) are attenuated in murine models. We hypothesized that pharmacologic activation of CpxR could serve as an antimicrobial/antivirulence strategy and recently showed that 2,3,4,9-tetrahydro-1H-carbazol-1-amines activate the CpxRA system by inhibiting CpxA phosphatase activity. Here, we tested the ability of a series of three CpxRA-activating compounds with increasing potency to clear UPEC stain CFT073 in a murine urinary tract infection model. We show that these compounds are well tolerated and achieve sufficient levels to activate CpxR in the kidneys, bladder, and urine. Although the first two compounds were ineffective in promoting clearance of CFT073 in the murine model, the most potent derivative, compound 26, significantly reduced bacterial recovery in the urine and trended toward reducing bacterial recovery in the bladder and kidneys, with efficacy similar to ciprofloxacin. Treatment of CFT073 cultured in human urine with compound 26 fostered accumulation of CpxR-P and decreased the expression of proteins involved in siderophore biosynthesis and binding, heme degradation, and flagellar movement. These studies suggest that chemical activation of CpxRA may present a viable strategy for treating infections due to UPEC. IMPORTANCE The increasing prevalence of urinary tract infections (UTIs) due to antibiotic-resistant uropathogenic Escherichia coli (UPEC) is a major public health concern. Bacteria contain proteins that sense their environment and have no human homologs and, thus, are attractive drug targets. CpxRA is a conserved sensing system whose function is to reduce stress in the bacterial cell membrane; activation of CpxRA reduces the expression of virulence determinants, which must cross the cell membrane to reach the bacterial surface. We previously identified a class of compounds that activate CpxRA. We show in a mouse UTI model that our most potent compound significantly reduced recovery of UPEC in the urine, trended toward reducing bacterial recovery in the bladder and kidneys, did not kill UPEC, and downregulated multiple proteins involved in UPEC virulence. Since these compounds do not act by a killing mechanism, they have potential to treat UTIs caused by antibiotic-resistant bacteria.


Subject(s)
Escherichia coli Infections , Escherichia coli Proteins , Urinary Tract Infections , Uropathogenic Escherichia coli , Animals , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Bacterial Proteins/genetics , Disease Models, Animal , Escherichia coli Infections/drug therapy , Escherichia coli Infections/microbiology , Escherichia coli Proteins/metabolism , Female , Gene Expression Regulation, Bacterial , Humans , Male , Mice , Phosphoric Monoester Hydrolases/genetics , Phosphoric Monoester Hydrolases/metabolism , Protein Kinases/genetics , Protein Kinases/metabolism , Salmonella typhimurium/metabolism , Urinary Tract Infections/drug therapy , Urinary Tract Infections/microbiology , Virulence Factors/genetics
2.
Bioorg Med Chem Lett ; 29(14): 1836-1841, 2019 07 15.
Article in English | MEDLINE | ID: mdl-31104993

ABSTRACT

Genetic activation of the bacterial two-component signal transduction system, CpxRA, abolishes the virulence of a number of pathogens in human and murine infection models. Recently, 2,3,4,9-tetrahydro-1H-carbazol-1-amines were shown to activate the CpxRA system by inhibiting the phosphatase activity of CpxA. Herein we report the initial structure-activity relationships of this scaffold by focusing on three approaches 1) A-ring substitution, 2) B-ring deconstruction to provide N-arylated amino acid derivatives, and 3) C-ring elimination to give 2-ethylamino substituted indoles. These studies demonstrate that the A-ring is amenable to functionalization and provides a promising avenue for continued optimization of this chemotype. Further investigations revealed that the C-ring is not necessary for activity, although it likely provides conformational constraint that is beneficial to potency, and that the (R) stereochemistry is required at the primary amine. Simplification of the scaffold through deconstruction of the B-ring led to inactive compounds, highlighting the importance of the indole core. A new lead compound 26 was identified, which manifests a ∼30-fold improvement in CpxA phosphatase inhibition over the initial hit. Comparison of amino and des-amino derivatives in bacterial strains differing in membrane permeability and efflux capabilities demonstrate that the amine is required not only for target engagement but also for permeation and accumulation in Escherichia coli.


Subject(s)
Carbazoles/therapeutic use , Animals , Carbazoles/pharmacology , Humans , Mice , Structure-Activity Relationship
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