作者: Eva-Anne Feuerbaum ; Philip W. Howard ; Denise E. Waldron ; Colin H. James ; Peter W. Taylor ; David E. Thurston ; Joao B. Moreira ; Keith R. Fox ; Helena Rosado ; Eva Stecher ; Maria de la Fuente ; Khondaker M. Rahman ; Stephen J. Gregson

OBJECTIVESPyrrolobenzodiazepine (PBD) dimers, tethered through inert propyldioxy or pentyldioxy linkers, possess potent bactericidal activity against a range of Gram-positive bacteria by virtue of their capacity to cross-link duplex DNA in sequence-selective fashion. Here we attempt to improve the antibacterial activity and cytotoxicity profile of PBD-containing conjugates by extension of dimer linkers and replacement of one PBD unit with phenyl-substituted or benzo-fused heterocycles that facilitate non-covalent interactions with duplex DNA.METHODSDNase I footprinting was used to identify high-affinity DNA binding sites. A staphylococcal gene microarray was used to assess epidemic methicillin-resistant Staphylococcus aureus 16 phenotypes induced by PBD conjugates. Molecular dynamics simulations were employed to investigate the accommodation of compounds within the DNA helix.RESULTSIncreasing the length of the linker in PBD dimers led to a progressive reduction in antibacterial activity, but not in their cytotoxic capacity. Complex patterns of DNA binding were noted for extended PBD dimers. Modelling of DNA strand cross-linking by PBD dimers indicated distortion of the helix. A majority (26 of 43) of PBD-biaryl conjugates possessed potent antibacterial activity with little or no helical distortion and a more favourable cytotoxicity profile. Bactericidal activity of PBD-biaryl conjugates was determined by inability to excise covalently bound drug molecules from bacterial duplex DNA.CONCLUSIONSPBD-biaryl conjugates have a superior antibacterial profile compared with PBD dimers such as ELB-21. We have identified six PBD-biaryl conjugates as potential drug development candidates.

2011-05-01·Journal of Antimicrobial Chemotherapy2区 · 医学

The minor groove-binding agent ELB-21 forms multiple interstrand and intrastrand covalent cross-links with duplex DNA and displays potent bactericidal activity against methicillin-resistant Staphylococcus aureus

2区 · 医学

Article

作者: Jason Hinds ; Eva-Anne Feuerbaum ; Khondaker M. Rahman ; David E. Thurston ; Helena Rosado ; Peter W. Taylor

OBJECTIVESThe antistaphylococcal pyrrolobenzodiazepine dimer ELB-21 forms multiple adducts with duplex DNA through covalent interactions with appropriately spaced guanine residues; it is now known to form interstrand and intrastrand adducts with oligonucleotide sequences of variable length. We determined the DNA sequence preferences of ELB-21 in relation to its capacity to exert a bactericidal effect by damaging DNA.METHODSFormation of adducts by ELB-21 and 12- to 14-mer DNA duplexes was investigated using ion-pair reversed phase liquid chromatography and mass spectrometry. Drug-induced changes in gene expression were measured in prophage-free Staphylococcus aureus RN4220 by microarray analysis.RESULTSELB-21 preferentially formed intrastrand adducts with guanines separated by three nucleotide base pairs. Interstrand and intrastrand adducts were formed with duplexes both longer and shorter than the preferred target sequences. ELB-21 elicited rapid bactericidal effects against prophage-carrying and prophage-free S. aureus strains; cell lysis occurred following activation and release of resident prophages. Killing appeared to be due to irreparable damage to bacterial DNA and susceptibility to ELB-21 was governed by the capacity of staphylococci to repair DNA lesions through induction of the SOS DNA damage response mediated by the RecA-LexA pathway.CONCLUSIONSThe data support the contention that ELB-21 arrests DNA replication, eliciting formation of ssDNA-RecA filaments that inactivate LexA, the SOS repressor, and phage repressors such as Cl, resulting in activation of the DNA damage response and de-repression of resident prophages. Above the MIC threshold, DNA repair is ineffective.

2009-11-01·Journal of Antimicrobial Chemotherapy2区 · 医学

Response of Staphylococcus aureus to subinhibitory concentrations of a sequence-selective, DNA minor groove cross-linking pyrrolobenzodiazepine dimer

2区 · 医学

Article

作者: Jason Hinds ; Eva-Anne Feuerbaum ; Marie Doyle ; David E. Thurston ; Keith R. Fox ; Peter W. Taylor

OBJECTIVESELB-21 is a pyrrolo[2,1-c][1,4]benzodiazepine dimer with potent antistaphylococcal activity; it binds covalently to guanine residues on opposing strands of duplex DNA, interfering with regulatory proteins and transcription elongation in a sequence-selective manner. Transcriptional and proteomic alterations induced by exposure of Staphylococcus aureus clinical isolate EMRSA-16 to ELB-21 were determined in order to define more precisely the bactericidal mechanism of the drug.METHODSDNase I footprinting was used to identify high-affinity DNA binding sites. Microarrays and gel electrophoresis were used to assess the ELB-21-induced phenotype.RESULTSHigh-affinity interstrand binding sites in which guanine residues were separated by 4 bp, and also some intrastrand cross-linking sites of variable length were identified. Exposure of EMRSA-16 to 0.015 mg/L ELB-21 elicited a 2-fold or greater up-regulation of 168 genes in logarithmic phase and 181 genes in stationary phase; the majority of genes affected were associated with resident prophages Sa2 and Sa3, pathogenicity island SaPI4 and DNA damage repair. ELB-21 induced a marked increase in the number of viable phage particles in culture supernatants. The expression of only a limited number of genes showed a >50% reduction. Sixteen extracellular and four intracellular proteins were differentially expressed during logarithmic and stationary phases, including RecA, proteins associated with staphylococcal pathogenesis (IsaA, CspA), cell division and wall synthesis.CONCLUSIONSELB-21 kills S. aureus by forming multiple interstrand and intrastrand DNA cross-links, resulting in induction of the DNA damage response, derepression of resident prophages and modulation of a limited number of genes involved with cell wall synthesis.

100 项与 ELB-21 相关的药物交易

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