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The need to develop new, improved treatments for tuberculosis (TB) remains urgent, and the repurposing of existing drugs represents a possible shortcut to market. Recently, there has been significant interest in host-directed adjuvant therapy to enhance bacillary killing. HMG-CoA reductase inhibitors (statins), which are among the most commonly prescribed drugs, have immunomodulatory properties and improve the clinical outcomes of bacterial infections.We studied the tuberculocidal activity of simvastatin alone and in combination with first-line anti-TB drugs in J774 macrophages and during chronic TB infection.Exposure to 5 μM simvastatin significantly increased the tuberculocidal activity of isoniazid in J774 macrophages at Day 3 after infection versus isoniazid alone (P=0.02). Similarly, relative to the standard oral regimen of rifampicin (10 mg/kg), isoniazid (10 mg/kg) and pyrazinamide (150 mg/kg) given five times weekly, the addition of 25 mg/kg simvastatin enhanced bacillary killing, reducing the number of lung cfu by an additional 1 log10 at Day 28 (P0.01) and by a further 1.25 log10 at Day 56 (P0.01).The potential additive activity of simvastatin to first-line TB treatment holds promise. However, further studies to identify the optimal statin and dosing are required. In addition the ability of combination treatment with statins to accelerate the time required to achieve a stable cure remains to be explored.

Objectives The antipsychotic drug thioridazine is active in the murine model of tuberculosis infection, which is predominantly intracellular in nature. Recent clinical reports suggest that thioridazine may play a role in the treatment of drug-resistant tuberculosis. We studied the tuberculocidal activity of thioridazine in guinea pigs, which develop necrotic lung granulomas histologically resembling their human counterparts. Methods Pharmacokinetic studies were performed in guinea pigs to establish human-equivalent doses of thioridazine. Guinea pigs were aerosol-infected with ∼100 bacilli of Mycobacterium tuberculosis and single-drug treatment was started 4 weeks later with a range of thioridazine doses daily (5 days/week) for up to 4 weeks. Control animals received no treatment or 60 mg/kg isoniazid. Results The human-equivalent dose of thioridazine was determined to be 5 mg/kg with saturable absorption noted above 50 mg/kg. At the start of treatment, the lung bacterial burden was ∼6.2 log10 cfu. Although isoniazid reduced bacillary counts more than 10-fold, thioridazine monotherapy showed limited killing over the range of doses tested, reducing lung bacillary counts by 0.3-0.5 log10 following 1 month of treatment. Thioridazine was tolerated up to 40 mg/kg. Conclusions Thioridazine has limited bactericidal activity against extracellular bacilli within necrotic granulomas. Its contribution to the sterilizing activity of combination regimens against drug-susceptible and drug-resistant tuberculosis remains to be determined.

Objectives: It is widely believed that persistent Mycobacterium tuberculosis inhabits necrotic lung granulomas in humans and that the microenvironmental conditions encountered therein render the bacilli phenotypically tolerant to antibiotics, accounting for the long duration required for successful treatment of tuberculosis (TB). To validate this belief, we directly compared the activity of rifampicin/isoniazid/pyrazinamide (RHZ) against chronic TB infection in guinea pigs, which exhibit caseous granulomas histologically resembling human caseous foci, and in mice, which lack necrotic granulomas. Methods: Guinea pigs and mice were aerosol-infected with M. tuberculosis CDC1551 and twice weekly treatment with RHZ was started 4 weeks later. Culture-positive relapse was assessed in subgroups of guinea pigs after 3 months and 4 months of treatment. Results: All guinea pig lungs exhibited histological evidence of granulomas with central caseation, while mouse lungs exhibited cellular lesions at the initiation of antibiotic treatment. Guinea pig lungs became culturenegative after 2 months of RHZ given twice weekly at human-equivalent doses. Relapse rates in guinea pigs were 0% (0/10) both after 3 months and 4 months of treatment. In contrast, all mouse lungs remained culture-positive after 4 months of equivalent RHZ exposures. Conclusions: Caseous necrosis does not reduce the sterilizing activity of the standard antituberculosis regimen of RHZ. Our findings have important implications for the use of alternative animal models in testing novel TB drug regimens and for modelling M. tuberculosis persistence.

Objectives In this study, we sought to compare the sterilizing activity of human-equivalent doses of the ‘Denver regimen’ against acute tuberculosis (TB) infection in the standard mouse model and in the guinea pig. Methods Pharmacokinetic studies in guinea pigs were used to establish human-equivalent doses for rifampicin, isoniazid and pyrazinamide. Guinea pigs and mice were aerosol-infected with Mycobacterium tuberculosis CDC1551 and treatment was started 2 weeks later with rifampicin/isoniazid/pyrazinamide for up to 6 months. For the first 2 weeks of therapy, the dosing frequency was 5 days/week, and for the remaining period, twice weekly. Treatment was discontinued in groups of 30 mice and 10 guinea pigs at 5 months and at 6 months, and these animals were held for a further 3 months in order to assess relapse rates. Results Guinea pig lungs became culture-negative after 3 months of predominantly twice-weekly treatment and relapse rates were 0% (0/10) both after 5 months and after 6 months of treatment. In contrast, all mice remained culture-positive despite 6 months of the same treatment, and 93% (28/30) and 69% (20/29) of mice relapsed after treatment for 5 and 6 months, respectively. Conclusions Treatment with rifampicin/isoniazid/pyrazinamide administered at human-equivalent doses is much more potent against acute TB infection in guinea pigs than in mice. Our findings have important implications for the use of alternative animal models in testing novel TB drug regimens and for modelling M. tuberculosis persistence.

Despite having potent activity against actively replicating Mycobacterium tuberculosis, isoniazid has very limited activity against dormant bacilli. In order to investigate the lack of bactericidal activity of this drug under conditions leading to mycobacterial dormancy, we studied the transcriptional pattern of M. tuberculosis in different physiological states following exposure to isoniazid.Global gene expression analysis was used to study M. tuberculosis treated with isoniazid in dormancy models of nutrient depletion and progressive hypoxia in vitro, as well as in an in vivo hollow fibre model of dormancy. Mycobacterial expression of the drug's putative transcriptional signature was investigated by RT-PCR in each dormancy model, and during the early and chronic phases of infection in the mouse aerosol model. Transcriptional responses were correlated with the bactericidal activity of isoniazid in the respective models.A small group of genes directly relevant to the mechanism of action of isoniazid was confirmed to constitute a transcriptional signature of the drug, as differential regulation of these genes was abrogated in an isoniazid-resistant, katG-deficient M. tuberculosis strain following isoniazid exposure. Isoniazid-induced expression of this transcriptional signature was abolished in dormant bacilli which had acquired phenotypic tolerance to isoniazid, regardless of the specific conditions responsible for the induction of the dormancy phenotype. Quantitative RT-PCR revealed that expression of isoniazid-regulated genes (IRGs) is dramatically altered under conditions of nutrient depletion and progressive hypoxia in vitro. Although these IRGs are highly induced following drug exposure early in infection in the mouse hollow fibre and aerosol models, correlating with potent bactericidal activity of the drug, their expression levels are markedly diminished during late-stage infection in these two models, coinciding with the greatly reduced bactericidal activity of isoniazid against these organisms.The reduced susceptibility of bacilli to the bactericidal drug isoniazid, as well as lack of expression of IRGs upon exposure to the drug, may be defining features of M. tuberculosis dormancy.