title: LpxC inhibitors as new antibacterial agents and tools for studying regulation of lipid A biosynthesis in Gram-negative pathogens.
abstract: The problem of multidrug resistance in serious Gram-negative bacterial pathogens has escalated so severely that new cellular targets and pathways need to be exploited to avoid many of the preexisting antibiotic resistance mechanisms that are rapidly disseminating to new strains. The discovery of small-molecule inhibitors of LpxC, the enzyme responsible for the first committed step in the biosynthesis of lipid A, represents a clinically unprecedented strategy to specifically act against Gram-negative organisms such as Pseudomonas aeruginosa and members of the Enterobacteriaceae. In this report, we describe the microbiological characterization of LpxC-4, a recently disclosed inhibitor of this bacterial target, and demonstrate that its spectrum of activity extends to several of the pathogenic species that are most threatening to human health today. We also show that spontaneous generation of LpxC-4 resistance occurs at frequencies comparable to those seen with marketed antibiotics, and we provide an in-depth analysis of the mechanisms of resistance utilized by target pathogens. Interestingly, these isolates also served as tools to further our understanding of the regulation of lipid A biosynthesis and enabled the discovery that this process occurs very distinctly between P. aeruginosa and members of the Enterobacteriaceae. Finally, we demonstrate that LpxC-4 is efficacious in vivo against multiple strains in different models of bacterial infection and that the major first-step resistance mechanisms employed by the intended target organisms can still be effectively treated with this new inhibitor.
  
  
  OBJECTIVE
  New antibiotics are needed for the effective treatment of serious infections caused by Gram-negative pathogens, and the responsibility of identifying new drug candidates rests squarely on the shoulders of the infectious disease community. The limited number of validated cellular targets and approaches, along with the increasing amount of antibiotic resistance that is spreading throughout the clinical environment, has prompted us to explore the utility of inhibitors of novel targets and pathways in these resistant organisms, since preexisting target-based resistance should be negligible. Lipid A biosynthesis is an essential process for the formation of lipopolysaccharide, which is a critical component of the Gram-negative outer membrane. In this report, we describe the in vitro and in vivo characterization of novel inhibitors of LpxC, an enzyme whose activity is required for proper lipid A biosynthesis, and demonstrate that our lead compound has the requisite attributes to warrant further consideration as a novel antibiotic.
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA andrew.tomaras@pfizer.com.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: Tomaras
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Craig J
authors.initials: CJ
authors.lastname: McPherson
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Kuhn
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Arlene
authors.initials: A
authors.lastname: Carifa
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Mullins
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: David
authors.initials: D
authors.lastname: George
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Charlene
authors.initials: C
authors.lastname: Desbonnet
authors.affiliation: Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, New York, USA.
authors.forename: Tess M
authors.initials: TM
authors.lastname: Eidem
authors.affiliation: Worldwide Medicinal Chemistry, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Justin I
authors.initials: JI
authors.lastname: Montgomery
authors.affiliation: Worldwide Medicinal Chemistry, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Matthew F
authors.initials: MF
authors.lastname: Brown
authors.affiliation: Worldwide Medicinal Chemistry, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Usa
authors.initials: U
authors.lastname: Reilly
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: Alita A
authors.initials: AA
authors.lastname: Miller
authors.affiliation: Discovery Biology, Antibacterials Research Unit, Pfizer Worldwide Research and Development, Groton, Connecticut, USA.
authors.forename: John P
authors.initials: JP
authors.lastname: O'Donnell
mesh_terms: D000581:Amidohydrolases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D017353:Gene Deletion; D020128:Inhibitory Concentration 50; D007711:Klebsiella pneumoniae / Q000187:drug effects / Q000378:metabolism; D008050:Lipid A / Q000096:biosynthesis*; D008070:Lipopolysaccharides / Q000378:metabolism; D051379:Mice; D008809:Mice, Inbred C3H; D008826:Microbial Sensitivity Tests; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; C490371:FabZ protein, Enterococcus faecalis; D008050:Lipid A; D008070:Lipopolysaccharides; D009097:Multienzyme Complexes; C105708:OprN protein, Pseudomonas aeruginosa; D000581:Amidohydrolases; C514484:LpxC deacetylase, Pseudomonas