ABSTRACT
Novel (non-fluoroquinolone) inhibitors of bacterial type II topoisomerases (NBTIs) are an emerging class of antibacterial agents. We report an optimized series of cyclobutylaryl-substituted NBTIs. Compound 14 demonstrated excellent activity both in vitro (S. aureus MIC90=0.125µg/mL) and in vivo (systemic and tissue infections). Enhanced inhibition of Topoisomerase IV correlated with improved activity in S. aureus strains with mutations conferring resistance to NBTIs. Compound 14 also displayed an improved hERG IC50 of 85.9µM and a favorable profile in the anesthetized guinea pig model.
Subject(s)
Anti-Bacterial Agents/pharmacology , DNA Gyrase/metabolism , DNA Topoisomerase IV/antagonists & inhibitors , Quinolines/pharmacology , Topoisomerase II Inhibitors/pharmacology , Topoisomerase Inhibitors/pharmacology , Animals , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , DNA Topoisomerase IV/metabolism , Dose-Response Relationship, Drug , Ether-A-Go-Go Potassium Channels/metabolism , Guinea Pigs , Humans , Microbial Sensitivity Tests , Molecular Structure , Quinolines/chemical synthesis , Quinolines/chemistry , Staphylococcus aureus/drug effects , Staphylococcus aureus/enzymology , Streptococcus pyogenes/drug effects , Streptococcus pyogenes/enzymology , Structure-Activity Relationship , Topoisomerase II Inhibitors/chemical synthesis , Topoisomerase II Inhibitors/chemistry , Topoisomerase Inhibitors/chemical synthesis , Topoisomerase Inhibitors/chemistryABSTRACT
A structurally novel set of inhibitors of bacterial type II topoisomerases with potent in vitro and in vivo antibacterial activity was developed. Dual-targeting ability, hERG inhibition, and pharmacokinetic properties were also assessed.
Subject(s)
Anti-Bacterial Agents/pharmacology , DNA Topoisomerase IV/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Quinolines/pharmacology , Staphylococcus aureus/drug effects , Streptococcus pneumoniae/drug effects , Topoisomerase II Inhibitors , Animals , Anti-Bacterial Agents/administration & dosage , Anti-Bacterial Agents/chemistry , DNA Gyrase/metabolism , DNA Topoisomerase IV/metabolism , DNA Topoisomerases, Type II/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/chemistry , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Humans , Microbial Sensitivity Tests , Molecular Structure , Quinolines/administration & dosage , Quinolines/chemistry , Rats , Staphylococcus aureus/enzymology , Streptococcus pneumoniae/enzymology , Structure-Activity RelationshipABSTRACT
Recent studies have demonstrated that bacteria possess an essential protein translocation system similar to mammalian signal recognition particle (SRP). Here we have identified the Ffh, a homologue of the mammalian SRP54 subunit from S. pneumoniae. Ffh is a 58-kDa protein with three distinct domains: an N-terminal hydrophilic domain (N-domain), a G-domain containing GTP/GDP binding motifs, and a C-terminal methionine-rich domain (M-domain). The full-length Ffh and a truncated protein containing N and G domains (Ffh-NG) were overexpressed in E. coli and purified to homogeneity. The full-length Ffh has an intrinsic GTPase activity with k(cat) of 0.144 min(-1), and the K(m) for GTP is 10.9 microM. It is able to bind to 4.5S RNA specifically as demonstrated by gel retardation assay. The truncated Ffh-NG has approximately the same intrinsic GTPase activity to the full-length Ffh, but is unable to bind to 4.5S RNA, indicating that the NG domain is sufficient for supporting intrinsic GTP hydrolysis, and that the M domain is required for RNA binding. The interaction of S. pneumoniae Ffh with its receptor, FtsY, resulted in a 20-fold stimulation in GTP hydrolysis. The stimulation was further demonstrated to be independent of the 4.5S RNA. In addition, a similar GTPase stimulation is also observed between Ffh-NG and FtsY, suggesting that the NG domain is sufficient and the M domain is not required for GTPase stimulation between Ffh and FtsY.
