title: Target- and resistance-based mechanistic studies with TP-434, a novel fluorocycline antibiotic.
abstract: TP-434 is a novel, broad-spectrum fluorocycline antibiotic with activity against bacteria expressing major antibiotic resistance mechanisms, including tetracycline-specific efflux and ribosomal protection. The mechanism of action of TP-434 was assessed using both cell-based and in vitro assays. In Escherichia coli cells expressing recombinant tetracycline resistance genes, the MIC of TP-434 (0.063 μg/ml) was unaffected by tet(M), tet(K), and tet(B) and increased to 0.25 and 4 μg/ml in the presence of tet(A) and tet(X), respectively. Tetracycline, in contrast, was significantly less potent (MIC ≥ 128 μg/ml) against E. coli cells when any of these resistance mechanisms were present. TP-434 showed potent inhibition in E. coli in vitro transcription/translation (50% inhibitory concentration [IC(50)] = 0.29 ± 0.09 μg/ml) and [(3)H]tetracycline ribosome-binding competition (IC(50) = 0.22 ± 0.07 μM) assays. The antibacterial potencies of TP-434 and all other tetracycline class antibiotics tested were reduced by 4- to 16-fold, compared to that of the wild-type control strain, against Propionibacterium acnes strains carrying a 16S rRNA mutation, G1058C, a modification that changes the conformation of the primary binding site of tetracycline in the ribosome. Taken together, the findings support the idea that TP-434, like other tetracyclines, binds the ribosome and inhibits protein synthesis and that this activity is largely unaffected by the common tetracycline resistance mechanisms.
authors.affiliation: Tetraphase Pharmaceuticals, Inc, Watertown, Massachusetts, USA. tgrossman@tphase.com
authors.forename: Trudy H
authors.initials: TH
authors.lastname: Grossman
authors.forename: Agata L
authors.initials: AL
authors.lastname: Starosta
authors.forename: Corey
authors.initials: C
authors.lastname: Fyfe
authors.forename: William
authors.initials: W
authors.lastname: O'Brien
authors.forename: David M
authors.initials: DM
authors.lastname: Rothstein
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Mikolajka
authors.forename: Daniel N
authors.initials: DN
authors.lastname: Wilson
authors.forename: Joyce A
authors.initials: JA
authors.lastname: Sutcliffe
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001667:Binding, Competitive; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D008826:Microbial Sensitivity Tests; D009154:Mutation; D011425:Propionibacterium acnes / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis / Q000187:drug effects; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology*; D013753:Tetracycline Resistance / Q000235:genetics; D013754:Tetracyclines / Q000494:pharmacology*; D014169:Transformation, Bacterial
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D012336:RNA, Ribosomal, 16S; D013754:Tetracyclines; C571179:eravacycline; D013752:Tetracycline