title: Antimicrobial activity of antisense peptide-peptide nucleic acid conjugates against non-typeable Haemophilus influenzae in planktonic and biofilm forms.
abstract: Antisense peptide nucleic acids (PNAs) are synthetic polymers that mimic DNA/RNA and inhibit bacterial gene expression in a sequence-specific manner.
  
  
  
  To assess activity against non-typeable Haemophilus influenzae (NTHi), we designed six PNA-peptides that target acpP, encoding an acyl carrier protein. MICs and minimum biofilm eradication concentrations (MBECs) were determined. Resistant strains were selected by serial passages on media with a sub-MIC concentration of acpP-PNA.
  
  
  
  The MICs of six acpP-PNA-peptides were 2.9-11 mg/L (0.63-2.5 μmol/L) for 20 clinical isolates, indicating susceptibility of planktonic NTHi. By contrast, MBECs were up to 179 mg/L (40 μmol/L). Compared with one original PNA-peptide (acpP-PNA1-3'N), an optimized PNA-peptide (acpP-PNA14-5'L) differs in PNA sequence and has a 5' membrane-penetrating peptide with a linker between the PNA and peptide. The optimized PNA-peptide had an MBEC ranging from 11 to 23 mg/L (2.5-5 μmol/L), indicating susceptibility. A resistant strain that was selected by the original acpP-PNA1-3'N had an SNP that introduced a stop codon in NTHI0044, which is predicted to encode an ATP-binding protein of a conserved ABC transporter. Deletion of NTHI0044 caused resistance to the original acpP-PNA1-3'N, but showed no effect on susceptibility to the optimized acpP-PNA14-5'L. The WT strain remained susceptible to the optimized PNA-peptide after 30 serial passages on media containing the optimized PNA-peptide.
  
  
  
  A PNA-peptide that targets acpP, has a 5' membrane-penetrating peptide and has a linker shows excellent activity against planktonic and biofilm NTHi and is associated with a low risk for induction of resistance.
authors.affiliation: Division of Infectious Diseases, Department of Medicine, University at Buffalo, State University of New York, Buffalo, NY, USA.|Clinical and Translational Research Center, University at Buffalo, State University of New York, Buffalo, NY, USA.
authors.forename: Taketo
authors.initials: T
authors.lastname: Otsuka
authors.affiliation: Division of Infectious Diseases, Department of Medicine, University at Buffalo, State University of New York, Buffalo, NY, USA.|Clinical and Translational Research Center, University at Buffalo, State University of New York, Buffalo, NY, USA.
authors.forename: Aimee L
authors.initials: AL
authors.lastname: Brauer
authors.affiliation: Division of Infectious Diseases, Department of Medicine, University at Buffalo, State University of New York, Buffalo, NY, USA.|Clinical and Translational Research Center, University at Buffalo, State University of New York, Buffalo, NY, USA.
authors.forename: Charmaine
authors.initials: C
authors.lastname: Kirkham
authors.affiliation: Department of Microbiology, Oregon State University, Corvallis, OR, USA.
authors.forename: Erin K
authors.initials: EK
authors.lastname: Sully
authors.affiliation: Department of Epidemiology of Microbial Diseases, Yale School of Public Health, Yale University, New Haven, CT, USA.
authors.forename: Melinda M
authors.initials: MM
authors.lastname: Pettigrew
authors.affiliation: Department of Molecular Biophysics and Biochemistry, W.M. Keck Foundation Biotechnology Resource Laboratory, Yale University, New Haven, CT, USA.
authors.forename: Yong
authors.initials: Y
authors.lastname: Kong
authors.affiliation: Department of Microbiology, Oregon State University, Corvallis, OR, USA.
authors.forename: Bruce L
authors.initials: BL
authors.lastname: Geller
authors.affiliation: Division of Infectious Diseases, Department of Medicine, University at Buffalo, State University of New York, Buffalo, NY, USA murphyt@buffalo.edu.|Clinical and Translational Research Center, University at Buffalo, State University of New York, Buffalo, NY, USA.|Department of Microbiology and Immunology, University at Buffalo, State University of New York, Buffalo, NY, USA.
authors.forename: Timothy F
authors.initials: TF
authors.lastname: Murphy
mesh_terms: D000213:Acyl Carrier Protein / Q000037:antagonists & inhibitors*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors; D018441:Biofilms / Q000187:drug effects*; D024881:Drug Resistance, Bacterial; D006193:Haemophilus influenzae / Q000187:drug effects* / Q000502:physiology; D008826:Microbial Sensitivity Tests; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology*; D020135:Peptide Nucleic Acids / Q000494:pharmacology*; D012692:Serial Passage
chemical_list: D000213:Acyl Carrier Protein; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D020319:Oligodeoxyribonucleotides, Antisense; D020135:Peptide Nucleic Acids