Statin adjunctive therapy shortens the duration of TB treatment in mice

Abstract

Background: The repurposing of existing agents may accelerate TB drug development. Recently, we reported that the lipid-lowering drug simvastatin, when added to the first-line antitubercular regimen, reduces the lung bacillary burden in chronically infected mice.

Objectives: We investigated whether the addition of simvastatin to the first-line regimen (isoniazid/rifampicin/pyrazinamide) shortens the duration of curative TB treatment in mice.

Methods: Mycobacterium tuberculosis-infected THP-1 cells were exposed to simvastatin to determine the effect of statins on the activity of first-line anti-TB drug activity and intracellular rifampicin concentration. Single-dose and steady-state pharmacokinetic studies guided optimized simvastatin dosing in vivo. BALB/c mice were aerosol-infected with M. tuberculosis H37Rv and drug treatment was initiated 6 weeks post-infection. Separate groups of mice received standard TB treatment with or without simvastatin. Relapse rates were assessed 3 months after discontinuation of each treatment regimen. MALDI-MS imaging was used to image the cholesterol content of mouse lung lesions.

Results: Simvastatin significantly enhanced the bactericidal activity of first-line drugs against intracellular M. tuberculosis without altering intracellular rifampicin concentrations. Adjunctive treatment with 60 mg/kg simvastatin shortened the time required to achieve culture-negative lungs from 4.5 to 3.5 months. Following 2.5, 3.5 and 4.5 months of treatment, relapse rates were 100%, 50% and 0%, respectively, in the control group and 50% (P = 0.03), 20% and 0%, respectively, in the statin group. Simvastatin did not alter plasma or lung lesion cholesterol levels.

Conclusions: Statins are attractive candidates for host-directed, adjunctive TB therapy. Further preclinical studies are needed to define the optimal statin and dosing.

Figure 1.

Effects of simvastatin alone and in combination with anti-TB drugs against M. tuberculosis residing in THP-1 macrophages. Data are presented as inhibition of M. tuberculosis growth in macrophages at 6 days post-infection in the presence of the indicated drug treatment, relative to the corresponding solvent (no drug) control. Drug doses yielding 50% reduction of RLU were used: 0.1 μM simvastatin; 0.011 μM isoniazid; 0.012 μM rifampicin; and 162.5 μM pyrazinamide. In the combination drug studies, one-half of the above concentrations of antitubercular drugs were used to study the potential adjunctive activity of simvastatin. S, simvastatin; SA, simvastatin acid; H, isoniazid; R, rifampicin; Z, pyrazinamide.

Figure 2.

Intracellular accumulation of simvastatin with and without rifampicin exposure within THP-1 cells. THP-1 cells were treated with a range of simvastatin doses (0.1–1 μM) given alone or with 0.224 μM rifampicin. After a 30 min exposure at 37°C, the intracellular concentrations of simvastatin and rifampicin were measured using LC-MS/MS. Data represent the mean of three biological replicates with standard deviations.

Figure 3.

Pharmacokinetic profile of simvastatin acid (dosed as 60 mg/kg simvastatin) with and without the first-line antitubercular drugs at steady-state. Average concentrations are shown (n = 3 at each timepoint) with standard deviations (error bars). HRZ, 10 mg/kg isoniazid + 10 mg/kg rifampicin + 150 mg/kg pyrazinamide.

Figure 4.

Adjunctive treatment with simvastatin reduces the time required to achieve culture-negative lungs in M. tuberculosis-infected BALB/c mice. HRZ, 10 mg/kg isoniazid + 10 mg/kg rifampicin + 150 mg/kg pyrazinamide; Sim, 60 mg/kg simvastatin.

Figure 5.

Simvastatin adjunctive therapy does not alter cholesterol content of lungs or plasma of M. tuberculosis-infected mice. (a) MALDI-MS images showing the distribution of cholesterol ([M + H–H₂O]⁺) in control, isoniazid/rifampicin/pyrazinamide-treated and isoniazid/rifampicin/pyrazinamide + simvastatin-treated mouse lung sections. A haematoxylin/eosin (H&E)-stained reference tissue is shown for identification of tissue structures. Isoniazid/rifampicin/pyrazinamide- and isoniazid/rifampicin/pyrazinamide + simvastatin-treated lungs have smaller lesions containing concentrated levels of cholesterol. Lesions are outlined in black in the corresponding H&E image. Scale bar = 5 mm. (b) Comparative quantification of cholesterol ions in the lesions of control, isoniazid/rifampicin/pyrazinamide-treated and isoniazid/rifampicin/pyrazinamide + simvastatin-treated mice. Relative quantification of cholesterol ions in mouse lesion sections was performed by determining the mean dehydrated cholesterol signal in regions of interest using MSiReader software.^, (c) Plasma cholesterol levels. n = 3 mice; values represent median ± SD. Data are from tissue collected at 1.5 months post-treatment. HRZ, 10 mg/kg isoniazid + 10 mg/kg rifampicin + 150 mg/kg pyrazinamide; Sim, 60 mg/kg simvastatin. This figure appears in colour in the online version of JAC and in black and white in the print version of JAC.