Structural Modifications of 3-Triazeneindoles and Their Increased Activity Against Mycobacterium tuberculosis.

We synthesized 100 novel indole-based compounds with polyaza-functionalities, including 3-triazeneindoles, and tested their activity in vitro against laboratory M. tuberculosis H37Rv and clinical izoniazid-resistant CN-40 isolates, using gross and fine titration approaches. Here we present a few 3-triazeneindoles with the highest anti-mycobacterial activity. Introduction of short lipid tails into the 3-triazeneindole core additionally increased their activity against mycobacteria engulfed by murine macrophages. We also demonstrate that the compound TU112, one of the most active in our previous study, being not bioavailable after administration in mice per os, manifests prominent anti-mycobacterial activity after intravenous or aerosol delivery, as assessed by the mouse serum and lung supernatant titration assays.

A new model for chronic and reactivation tuberculosis: Infection with genetically attenuated Mycobacterium tuberculosis in mice with polar susceptibility.

TB infection in mice develops relatively rapidly which interferes with experimental dissection of immune responses and lung pathology features that differ between genetically susceptible and resistant hosts. Earlier we have shown that the M. tuberculosis strain lacking four of five Rpf genes (ΔACDE) is seriously attenuated for growth in vivo. Using this strain, we assessed key parameters of lung pathology, immune and inflammatory responses in chronic and reactivation TB infections in highly susceptible I/St and more resistant B6 mice. ΔACDE mycobacteria progressively multiplied only in I/St lungs, whilst in B6 lung CFU counts decreased with time. Condensed TB foci apeared in B6 lungs at week 4 of infection, whilst in I/St their formation was delayed. At the late phase of infection, in I/St lungs TB foci fused resulting in extensive pneumonia, whereas in B6 lungs pathology was limited to condensed foci. Macrophage and neutrophil populations characteristically differed between I/St and B6 mice at early and late stages of infection: more neutrophils accumulated in I/St and more macrophages in B6 lungs. The expression level of chemokine genes involved in neutrophil influx was higher in I/St compared to B6 lungs. B6 lung cells produced more IFN-γ, IL-6 and IL-11 at the early and late phases of infection. Overall, using a new mouse model of slow TB progression, we demonstrate two important features of ineffective infection control underlined by shifts in lung inflammation: delay in early granuloma formation and fusion of granulomas resulting in consolidated pneumonia late in the infectious course.

In Vitro Activity of 3-Triazeneindoles against Mycobacterium tuberculosis and Mycobacterium avium.

Among 230 target-synthesized indole-based compounds, seven 3-triazenoindoles showed MICs of 0.2 to 0.5 µg/ml against Mycobacterium tuberculosis strain H37Rv and isoniazid-resistant human isolate CN-40. The TU112 compound was active also against a dormant form of M. tuberculosis Some of these triazenoindoles were active against Mycobacterium avium, with MICs of 0.05 to 0.5 µg/ml. The selectivity indices (SI) for M. tuberculosis and M. avium were significantly higher than 10, making these compounds acceptable for the next testing step.

Drug testing in mouse models of tuberculosis and nontuberculous mycobacterial infections.

Mice as a species are susceptible to tuberculosis infection while mouse inbred strains present wide spectrum of susceptibility/resistance to this infection. However, non-tuberculosis Mycobacterial infections usually cannot be modeled in mice of common inbred strains. Introduction of specific properties, such as gene mutations, recombinants, targeted gene knockouts significantly extended the use of mice to mimic human Mycobacterial infections, including non-tuberculosis ones. This review describes the available mouse models of tuberculosis and non-tuberculosis infections and drug therapy in these models. Mouse models of non-tuberculosis infections are significantly less developed than tuberculosis models, hampering the development of therapies.

Anti-tuberculosis drug therapy in mice of different inbred strains.

Standard anti-tuberculosis (TB) drug therapy had distinct effects on the bacilli burden in mice of DBA/2, C3H, SWR/J, and C57BL/6 inbred strains. To standardize the TB infection process, susceptible DBA/2 mice were infected with 1/10 of the dose used for relatively resistant C57BL/6 mice, such that the lung CFUs were roughly identical 3 weeks after infection when therapy was initiated. We found that TB treatment was more effective in the susceptible DBA/2 mice than in the relatively resistant C57BL/6 mice.

Activity of SQ641, a capuramycin analog, in a murine model of tuberculosis.

New delivery vehicles and routes of delivery were developed for the capuramycin analogue SQ641. While this compound has remarkable in vitro potency against Mycobacterium tuberculosis, it has low solubility in water and poor intracellular activity. We demonstrate here that SQ641 dissolved in the water-soluble vitamin E analogue alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) or incorporated into TPGS-micelles has significant activity in a mouse model of tuberculosis.

Preclinical study of new TB drugs and drug combinations in mouse models.

Tuberculosis (TB) remains one of the leading infectious killers in the world. New anti-TB drugs and more effective drug combinations are urgently needed, particularly given the increasing incidence of drug-resistant TB and HIV-TB co-infection. This review describes the available mouse models of TB and describes their utility in the evaluation of new TB drug candidates and in the evaluation of the efficacy of new TB drug combinations. Some of the most recent patents on promising TB drug-candidates are also mentioned here.

Drug therapy of experimental tuberculosis (TB): improved outcome by combining SQ109, a new diamine antibiotic, with existing TB drugs.

Substitution of the new diamine antibiotic SQ109 for ethambutol in a mouse model of chronic tuberculosis (TB) improved efficacy of combination drug therapy with first-line TB drugs rifampin and isoniazid, with or without pyrazinamide: at 8 weeks, lung bacteria were 1.5 log10 lower in SQ109-containing regimens.

Rapid, simple in vivo screen for new drugs active against Mycobacterium tuberculosis.

We evaluated the use of a simple and easy-to-obtain potential marker of tuberculosis (TB) drug efficacy, body weight, and correlated weight loss or gain with the number of CFU of Mycobacterium tuberculosis in lungs and spleens of infected mice. C3H mice were infected intravenously with 10(6) CFU of virulent M. tuberculosis H37Rv, and body weight was evaluated for several weeks after infection. At day 20, infected untreated mice consistently lost more than 25% of their body weight. Chemotherapy with selected orally active anti-TB drugs was initiated 7 days following infection and continued for 13 days. Drugs that were administered daily by gavage included isoniazid (INH), ethambutol (EMB), rifampin (RIF), and moxifloxacin (MXF). At the most effective doses, each of these drugs inhibited bacterial growth and abolished infection-induced body weight loss. Chemotherapy with 1/10 the standard dose of INH determined in accepted long-term murine models of TB also prevented body weight loss, while chemotherapy with 1/10 the standard dose of RIF did not. With only 2 weeks of chemotherapy, we observed a good reverse correlation between CFU in lung or spleen and body weight of mice. The simple measurement of weight in TB-infected drug-treated mice required only a weight balance, and go/no-go drug efficacy data was available on day 20 without the necessity of prolonged drug treatment and long (3 weeks or more) in vitro culture times to obtain organ CFU values.

Multigenic control of disease severity after virulent Mycobacterium tuberculosis infection in mice.

Following challenge with virulent Mycobacterium tuberculosis, mice of the I/St inbred strain exhibit shorter survival time, more rapid body weight loss, higher mycobacterial loads in organs, and more severe lung histopathology than mice of the A/Sn strain. We previously performed a genome-wide scan for quantitative trait loci (QTLs) that control the severity of M. tuberculosis-triggered disease in [(A/Sn x I/St) F1 x I/St] backcross-1 (BC1) mice and described several QTLs that are significantly or suggestively linked to body weight loss. In the present study we expanded our analysis by including the survival time phenotype and by genotyping 406 (A/Sn x I/St) F2 mice for the previously identified chromosomal regions of interest. The previously identified 12-cM-wide QTL on distal mouse chromosome 3 was designated tbs1 (tuberculosis severity 1); the location of the QTL on proximal chromosome 9 was narrowed to a 9-cM interval, and this QTL was designated tbs2. Allelic variants of the tbs2 locus appeared to be involved in control of both body weight loss and survival time. Also, the data strongly suggested that a QTL located in the vicinity of the H-2 complex on chromosome 17 is involved in control of tuberculosis in mice of both genders, whereas the tbs1 locus seemed to have an effect on postinfection body weight loss in female mice. Interestingly, these loci appeared to interact with each other, which suggests that there might be a basic genetic network for the control of intracellular parasites. Overall, linkage data reported here for F2 mice are in agreement with, and add to, our previous findings concerning the control of M. tuberculosis-triggered disease in the BC1 segregation.

Murine Model of Tuberculosis. In vitro and in vivo Study.

Animal models that mimic the whole spectrum of susceptibility and resistance of human populations to tuberculosis would be useful for studying pathogenesis of tuberculosis, mechanisms of the host defense, and for testing prospective prophylactic and therapeutic tuberculosis vaccines and drugs. Inbred mouse strains are broadly and successfully used to model human tuberculosis and study mechanisms of host-mycobacteria interaction. Here we consider numerous aspects of murine models of tuberculosis. We analyze the course of tuberculosis depending on route and dosage of mycobacterial infection, and discuss the conformity of these approaches to the natural route of human infection. The advantage of the mouse model is provided by the availability of hundreds of inbred strains with different genetic background. It allows a genetic analysis of the mouse susceptibility/resistance to tuberculosis. Mouse strains with specific properties such as congenic, recombinant, recombinant inbred, recombinant congenic, mutant and usage of whole genome screening led to mapping loci controlling these traits. Transgenic mouse strains with gene insertion or targeted mutation (knock-out of a gene) are very effective tools for studying the role of specific genes controlling the anti-tuberculosis immunity. Mice with knock-out genes for cytokines, chemokines, cell subpopulations, cell receptors, and enzymes are broadly used to identify the mechanisms of tuberculosis control. In this review we examine a mouse model for anti-tuberculosis vaccine and drug testing. In addition, we present some important phenomena of tuberculosis immunity - latent tuberculosis/reactivation and immunological memory to tuberculosis.