High-dose rifampicin kills persisters, shortens treatment duration, and reduces relapse rate in vitro and in vivo

Anthony Coates,Alexander Liu,Denis Mitchison,Fatima Ortega-Muro,Laura Alameda-Martin,Yanmin Hu

doi:10.3389/fmicb.2015.00641

Abstract

Although high-dose rifampicin holds promise for improving tuberculosis control by potentially shortening treatment duration, these effects attributed to eradication of persistent bacteria are unclear. The presence of persistent Mycobacterium tuberculosis was examined using resuscitation promoting factors (RPFs) in both in vitro hypoxia and in vivo murine tuberculosis models before and after treatment with incremental doses of rifampicin. Pharmacokinetic parameters and dose-dependent profile of rifampicin in the murine model were determined. The Cornell mouse model was used to test efficacy of high-dose rifampicin in combination with isoniazid and pyrazinamide and to measure relapse rate. There were large numbers of RPF-dependent persisters in vitro and in vivo. Stationary phase cultures were tolerant to rifampicin while higher concentrations of rifampicin eradicated plate count positive but not RPF-dependent persistent bacteria. In murine infection model, incremental doses of rifampicin exhibited a dose-dependent eradication of RPF-dependent persisters. Increasing the dose of rifampicin significantly reduced the risk of antibiotic resistance emergence. In Cornell model, mice treated with high-dose rifampicin regimen resulted in faster visceral clearance; organs were M. tuberculosis free 8 weeks post-treatment compared to 14 weeks with standard-dose rifampicin regimen. Organ sterility, plate count and RPF-dependent persister negative, was achieved. There was no disease relapse compared to the standard dose regimen (87.5%). High-dose rifampicin therapy results in eradication of RPF-dependent persisters, allowing shorter treatment duration without disease relapse. Optimizing rifampicin to its maximal efficacy with acceptable side-effect profiles will provide valuable information in human studies and can potentially improve current tuberculosis chemotherapy.

Highlights

Tuberculosis remains a major cause of mortality worldwide (World Health Organization [WHO], 2010)
Materials and Methods In Vitro Hypoxia Model M. tuberculosis strain H37Rv was grown in 7H9 medium containing 0.05% Tween 80 supplemented with 10% albumin dextrose complex (ADC; Becton and Dickinson, UK) at 37◦C without disturbance for 200 days (Hu et al, 2000)
The susceptibility patterns of the strain to anti-TB drugs defined as minimum inhibitory concentration (MIC) are listed as following: rifampicin 0.5 mg/L, rifabutin 0.0625 mg/L, rifapentine 0.25 mg/L, isoniazid 0.25 mg/L, ethambutol 2 mg/L, Pharmacokinetic of Rifampicin in BALB/S Mice Pharmacokinetic (PK) of rifampicin in non-infected mice was determined by dosing-ranging studies

Summary

Introduction

Tuberculosis remains a major cause of mortality worldwide (World Health Organization [WHO], 2010). Similar observations were made in mice (Jayaram et al, 2003; Rosenthal et al, 2012; de Steenwinkel et al, 2013), with a maximum tolerable dose of 160 mg/kg per day (de Steenwinkel et al, 2013). Whether these effects are attributed to eradication of persistent bacteria, the fundamental problem preventing effective disease control, is unclear

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Conclusion