Spiropiperidyl rifabutins: expanded in vitro testing against ESKAPE pathogens and select bacterial biofilms

Abstract

The expanded microbiological evaluation of a series of rifastures, novel spiropiperidyl rifamycin derivatives, against clinically relevant ESKAPE bacteria has identified several analogs with promising in vitro bioactivities against antibiotic-resistant strains of Enterococcus faecium and Staphylococcus aureus. Thirteen of the rifastures displayed minimum inhibitory concentrations (MICs) below 1 µg/ml against the methicillin- and vancomycin-resistant forms of S. aureus and E. faecium (MRSA, VRSA, VRE). Aryl-substituted rifastures 1, 11, and 12 offered the greatest bioactivity, with MICs reaching ≤0.063 µg ml^-1 for these human pathogens. Further analysis indicates that diphenyl rifasture 1 had greater antibiofilm activity against S. aureus and lower cytotoxicity in mammalian HEK cells than rifabutin.

Fig. 1

Chemical structures of rifamycin S and rifabutin

Fig. 2

Spiropiperidyl rifastures (RFA) were synthesized by condensation of 3-amino-4-iminorifamycin S with substituted 4-piperidones [15]

Fig. 3

Inhibitory effects of rifastures (RFA) and rifamycin standards on S. aureus growth. a Comparison of growth inhibition by optical density measurements (600 nm). Samples were tested in duplicate at 1 × MIC against S. aureus COL. b Comparison of minimum bactericidal concentrations (MBCs) where MBC₅₀ and MBC₉₀ represents reduction in cfu/ml by 50% and 90%, respectively. Samples were tested in triplicate at 0.25 to 4 × MIC against S. aureus CBD-635. Percent recovery was calculated based on cfu ml⁻¹ in comparison to DMSO control. MBC₅₀ and MBC₉₀ values were calculated using linear regression of the percent recovery compared to no-treatment controls. c Comparison of minimum biofilm eradication concentrations (MBECs) where MBEC₅₀ and MBEC₉₀ corresponds to reduction in the viability of cells within the biofilm by 50% and 90%, respectively. Samples were tested in triplicate at 0.25 to 4 × MIC against S. aureus CBD-635. Percent recovery was calculated based on cfu ml⁻¹ in comparison to DMSO control

Fig. 4

Cytotoxicity of compounds against HEK293. Cells were seeded and allowed to grow for 24 h prior to incubation with compounds for 48 h. Results were calculated based on DMSO controls. Error bars represent ± SEM of three replicates