Discovery of novel 5-(ethyl or hydroxymethyl) analogs of 2'-'up' fluoro (or hydroxyl) pyrimidine nucleosides as a new class of Mycobacterium tuberculosis, Mycobacterium bovis and Mycobacterium avium inhibitors.

Discovery of novel antimycobacterial compounds that work on distinctive targets and by diverse mechanisms of action is urgently required for the treatment of mycobacterial infections due to the emerging global health threat of tuberculosis. We have identified a new class of 5-ethyl or hydroxy (or methoxy) methyl-substituted pyrimidine nucleosides as potent inhibitors of Mycobacterium bovis, Mycobacterium tuberculosis (H37Ra, H37Rv) and Mycobacterium avium. A series of 2'-'up' fluoro (or hydroxy) nucleosides (1, 2, 4-6, 9, 10, 13, 16, 18, 21, 24) was synthesized and evaluated for antimycobacterial activity. Among 2'-fluorinated compounds, 1-(3-bromo-2,3-dideoxy-2-fluoro-ß-d-arabinofuranosyl)-5-ethyluracil (13) exhibited promising activity against M. bovis and Mtb alone, and showed synergism when combined with isoniazid. The most active compound emerging from these studies, 1-(ß-d-arabinofuranosyl)-4-thio-5-hydroxymethyluracil (21) inhibited Mtb (H37Ra) (MIC(50)=0.5 µg/mL) and M. bovis (MIC(50)=0.5 µg/mL) at low concentrations, and was ten times more potent against Mtb (H37Ra) than cycloserine (MIC(50)=5.0 µg/mL), a second line drug. It also showed an additive effect when combined with isoniazid. Compound 21 retained sensitivity against a rifampicin-resistant (H37Rv) strain of Mtb (MIC(50)=1 µg/mL) at concentrations similar to that for a rifampicin-sensitive (H37Rv) strain, suggesting that it has no cross-resistance to a first-line anti-TB drug. In addition, the replication of M. avium was also inhibited by 21 (MIC(50)=10 µg/mL). No cellular toxicity of 13 or 21 was observed up to the highest concentration tested (CC(50)>100 µg/mL). These observations offer promise for a new drug treatment regimen to augment and complement the current chemotherapy of TB.

Antimycobacterial activities of 5-alkyl (or halo)-3'-substituted pyrimidine nucleoside analogs.

Several 5-alkyl (or halo)-3'-azido (amino or halo) analogs of pyrimidine nucleosides have been synthesized and evaluated against Mycobacterium bovis, Mycobacterium tuberculosis and Mycobacterium avium. Among these compounds, 3'-azido-5-ethyl-2',3'-dideoxyuridine (3) was found to have significant antimycobacterial activities against M. bovis (MIC(50)=1µg/mL), M. tuberculosis (MIC(50)=10µg/mL) and M. avium (MIC(50)=10µg/mL).

3'-bromo analogues of pyrimidine nucleosides as a new class of potent inhibitors of Mycobacterium tuberculosis.

Tuberculosis (TB) is a major health problem worldwide. We herein report a new class of pyrimidine nucleosides as potent inhibitors of Mycobacterium tuberculosis (M. tuberculosis). Various 2'- or 3'-halogeno derivatives of pyrimidine nucleosides containing uracil, 5-fluorouracil, and thymine bases were synthesized and evaluated for antimycobacterial activities. Among the compounds tested, 3'-bromo-3'-deoxy-arabinofuranosylthymine (33) was the most effective antituberculosis agent in the in vitro assays against wild-type M. tuberculosis strain (H37Ra) (MIC(50) = 1 microg/mL) as well as drug-resistant (H37Rv) (rifampicin-resistant and isoniazid-resistant) strains of M. tuberculosis (MIC(50) = 1-2 microg/mL). Compound 33 also inhibited intracellular M. tuberculosis in a human monocytic cell line infected with H37Ra, demonstrating higher activity against intramacrophagic mycobacteria (80% reduction at 10 microg/mL concentration) than extracellular mycobacteria (75% reduction at 10 microg/mL concentration). In contrast, pyrimidine nucleosides possessing 5-fluorouracil base were weak inhibitors of M. tuberculosis. No cytotoxicity was found up to the highest concentration of compounds tested (CC(50) > 100-200 microg/mL) against a human cell line. Overall, these encouraging results substantiate the potential of this new class of compounds as promising antituberculosis agents.

Growth inhibition of Mycobacterium bovis, Mycobacterium tuberculosis and Mycobacterium avium in vitro: effect of 1-beta-D-2'-arabinofuranosyl and 1-(2'-deoxy-2'-fluoro-beta-D-2'-ribofuranosyl) pyrimidine nucleoside analogs.

The resurgence of tuberculosis and the emergence of multiple-drug-resistant strains of Mycobacteria necessitate the search for new classes of antimycobacterial agents. We synthesized a series of 1-beta-D-2'-arabinofuranosyl and 1-(2'-deoxy-2'-fluoro-beta-D-ribofuranosyl) pyrimidine nucleosides possessing diverse sets of alkynyl, alkenyl, alkyl, and halo substituents at the C-5 position of the uracil and investigated their effect on activity against M. tuberculosis, M. bovis, and M. avium. Among these molecules, 5-alkynyl-substituted derivatives emerged as potent inhibitors of M. bovis, M. tuberculosis, and M. avium. Nucleosides 1-beta-D-2'-arabinofuranosyl-5-dodecynyluracil (5), 1-(2'-deoxy-2'-fluoro-beta-D-ribofuranosyl)-5-dodecynyluracil (24), and 1-(2'-deoxy-2'-fluoro-beta-D-ribofuranosyl)-5-tetradecynyluracil (25) showed the highest antimycobacterial potency against M. bovis and M. tuberculosis. The MIC90 exhibited by compounds 5, 24, and 25 was similar or close to that of the reference drug rifampicin. The most active compounds 5, 24, and 25 were also found to retain sensitivity against a rifampicin-resistant strain of M. tuberculosis H37Rv at similar concentrations. Some of these analogs also revealed in vitro antimicrobial effect against several other gram-positive pathogens.