| Congresso Brasileiro de Microbiologia 2023 | Resumo: 359-1 | ||||
Resumo:Tuberculosis (TB), the infection disease caused mainly by Mycobacterium tuberculosis (M. tb), remains an important cause of death around of the world. In 2021, approximately 1.6 million people died and 10.6 million became ill from TB. The standard treatment scheme is long (minimum 6 months) and inactive against resistant strains of M. tb, which have developed multiple resistance mechanisms against these standard drugs. The development of new medicines for TB treatment is one of the goals of the WHO’s End TB Strategy and, focusing on this, a new benzofuroxan compound was analysed and showed great potential to inhibit the mycobacteria. The minimal inhibitory concentration (MIC90) of benzofuroxan TB10 was obtained using the Resazurin Microtiter Assay (REMA method) against standard (H37Rv ATCC 27294) and clinical strains (n = 8) of M. tb: briefly the inoculum was cultured until 1 MacFarland standard and diluted with the compound in a range of 0.09 to 25 ug/mL in a 96-wells plate; after 7 days of incubation at 37 °C, resazurin was added and the fluorescence was read after 24h. The MIC was considered the minor concentration able to inhibit 90% of the inoculum. The intramacrophagic activity was performed using J774A.1 cells infected with M. tb (MOI 2:1). The cells were cultured using RPMI medium and plated in a 24-wells plate. After adherence, the mycobacterium inoculum was added. The treatment was added at 1x, 5x and 10x MIC90 and after 72 h the cells were lysed, and the remaining bacteria was platted in agar medium. The units forming colony (UFC/mL) were counted and the inhibition activity of the compound is presented in a log10. In vivo efficacy was tested treating BALB/C mice infected with M. tb (intranasal infection) with daily dose of oral suspension of TB10 for 4 weeks. The mice were euthanized, the lungs were collected, processed, and platted in agar medium. The UFC/mL were counted and a log10 graphic represent the results obtained. The activity in the association with rifampicin was observed by Checkboarder methodology. The compounds were diluted in 96-well plates and mixed in different concentrations. M. tb inoculum was added and, like REMA method, the plate was incubated, and the fluorescence was read. The compound interaction could be classified as a synergistic, antagonistic, or neutral activity. The MIC90 obtained against standard strain of M. tb was 0.09 g/mL and against different resistant clinical strains (including multi and extensive-drug resistant strains) the MIC90 range of 0.26 to 4.71 ug/mL. Against intramacrophagic bacteria, the TB10 compound decreased 0.9 log10 of the inoculum at 10x MIC90 after 72 h of exposition. In vivo, more than 3.0 log10 of mycobacterial inoculum were inhibit after the treatment with TB10, and the in vitro interaction with rifampicin presents a synergistic effect, with a decreased MIC90 value to both compounds. The compound TB10 demonstrated a great potential in inhibit diverse strains of M. tb, with potent in vitro, intracellular, and in vivo activity. In association with a standard drug treatment, its synergistic effect demonstrates an advantage to treatment scheme. Palavras-chave: Mycobacterium tuberculosis, Drug Discovery, Benzofuroxan Agęncia de fomento:FAPESP (Process n. 2018/21778-3 and 2020/13497-4) | |||||