Bactericidal activity of PA-824 against Mycobacterium tuberculosis under anaerobic conditions and computational analysis of its novel analogues against mutant Ddn receptor.

BACKGROUND: The resurgence of multi-drug resistant tuberculosis (MDR-TB) and HIV associated tuberculosis (TB) are of serious global concern. To contain this situation, new anti-tuberculosis drugs and reduced treatment regimens are imperative. Recently, a nitroimidazole, PA-824, has been shown to be active against both replicating and non-replicating bacteria. It is activated by the enzyme Deazaflavin-dependent nitroreductase (Ddn) present in Mycobacterium tuberculosis which catalyzes the reduction of PA-824, resulting in the release of lethal reactive nitrogen species (RNS) within the bacteria. In this context, PA-824 was analyzed for its activity against latent tuberculosis under anaerobic conditions and compared with rifampicin (RIF) and pyrazinamide (PZA). Recent mutagenesis studies have identified A76E mutation which affects the above mentioned catalysis and leads to PA-824 resistance. Hence, novel analogues which could cope up with their binding to mutant Ddn receptor were also identified through this study. RESULTS: PA-824 at an optimum concentration of 12.5 µg/ml showed enhanced bactericidal activity, resulting in 0 CFU/ml growth when compared to RIF and PZA at normal pH and anaerobic condition. Further docking studies revealed that a combinatorial structure of PA-824 conjugated with moxifloxacin (ligand 8) has the highest binding affinity with the wild type and mutant Ddn receptor. CONCLUSIONS: PA-824 has been demonstrated to have better activity under anaerobic condition at 12.5 µg/ml, indicating an optimized dose that is required for overcoming the detoxifying mechanisms of M. tuberculosis and inducing its death. Further, the development of resistance through A76E mutation could be overcome through the in silico evolved ligand 8.

Astrocytes resolve ER stress through mitochondrial fusion facilitated by biotin availability.

Structures of cellular organelles are intertwined with their functions that undergo alterations once the organelles are stressed. Since organelle functions are dependent on each other, an organelle-specific stress possibly influences the structure and function of its associated organelles. In this perspective, our study demonstrated that endoplasmic reticulum (ER)-specific stress induced by tunicamycin in primary astroglial culture is associated with altered mitochondrial dynamics and matched with the changes as observed in the aging rat brain. However, the exogenous addition of biotin, a highly lipogenic and mitochondrial vitamin, ameliorates ER stress even though its direct targets are not known within ER. Alternatively, the increased biotinylation of mitochondrial carboxylases preserves its basal respiratory capacity by upregulating mitofusin 2 (Mfn2) and, possibly, its associated role on mitochondrial fusion. Furthermore, the Mfn2 increase by biotin augments physical interaction between ER and functional mitochondria to exchange biomolecules as a part of ER stress resolution. This suggests an increased demand for micronutrient biotin under ER stress resolves the same by undergoing appropriate structural and metabolic contacts between ER and mitochondria. These findings provide a paradigm to resolve stress in one organelle by sustaining the metabolic commitments of another interdependent organelle. The findings also highlight a novel role of biotin in inducing Mfn2 expression and localization under ER stress in addition to its known role as a co-enzyme.

Distinct utilization of biotin in and between adipose and brain during aging is associated with a lipogenic shift in Wistar rat brain.

Tissue-specific metabolism determines their functions that collectively sense and respond to numerous stress cues to achieve systemic homeostasis. Chronic stress skews such metabolic profiles and leads to failure of organs as evidenced by a bias towards lipid synthesis and storage in the aging brain, muscle, and liver under Alzheimer's disease, sarcopenia, and non-alcoholic fatty liver disease, respectively. In contrast, the tissue destined for lipid synthesis and storage, such as adipose, limits its threshold and develops diabetes mellitus. However, the underlying factors that contribute to this lipogenic shift between organs are unknown. From this perspective, differential biotin utilization between lipid-rich tissues such as adipose and brain during aging was hypothesized owing to the established role of biotin in lipogenesis. The same was tested using young and aged Wistar rats. We found that adipose-specific biotin content was much higher than the brain irrespective of aging status, as well as its associated cues. However, within tissues, the adipose fails to maintain its biotinylation levels during aging whereas the brain seizes more biotin and exhibits lipid accumulation. Furthermore, mimicking the age-related stress cues in vitro such as high glucose and endoplasmic reticulum stress deprive the astroglial biotin content, but not that of adipocytes. Lipid accumulation in the aging brain was also correlated with increased S-adenosylmethionine levels and biotin utilization by astrocytes. In summary, differential biotin utilization between adipose and brain under aging and their respective cell types like adipocytes and astrocytes under age-associated stress cues connects well with the lipogenic shift in rat brain.