6-methylflavone exerts selective antimycobacterial activity with membrane energetics disruption and demonstrates therapeutic potential against Mycobacterium abscessus infection

ABSTRACT Treatment of nontuberculous mycobacteria (NTM) diseases, especially Mycobacterium abscessus pulmonary disease, remains a major clinical challenge due to its broad antibiotic resistance and limited therapeutic options. To search for new candidates, we established a synthetic flavonoid library and screened it using a bioluminescent M. abscessus reporter strain. Among the compounds tested, 6-methylflavone (6-MF) was identified as a selective inhibitor of mycobacterial growth. 6-MF showed consistent activity against multiple clinical isolates of M. abscessus with minimum inhibitory concentration (MIC) values of 8–16 μg/mL and was also effective against intracellular mycobacteria without cytotoxicity. The antimicrobial activity was largely restricted to mycobacteria, indicating its narrow spectrum. Checkerboard assays demonstrated strong synergy with clarithromycin, while the effect with amikacin was limited to weak synergy or indifference. Resistance to 6-MF emerged at a much lower frequency than to amikacin in vitro, underlining its clinical advantage. Notably, 6-MF increased mycobacterial respiratory flux and reduced membrane potential, phenotypes associated with disruption of membrane energetics. In a mouse model of pulmonary infection, 6-MF reduced bacterial burden and lung inflammation to a level comparable to amikacin. These results indicate that 6-MF is a promising lead compound with the potential for further development as a therapeutic option for M. abscessus infection. IMPORTANCE Despite the increasing global incidence of nontuberculous mycobacterial (NTM) diseases, effective treatment regimens remain limited. In particular, Mycobacterium abscessus exhibits notoriously multidrug resistance, making its treatment extremely challenging. In this study, we identified 6-methylflavone (6-MF) as a selective antimycobacterial compound through screening of synthetic flavonoid library. 6-MF specifically inhibited mycobacterial growth and disrupted membrane energetics while showing negligible effects on other bacterial genera. Mycobacteria have unique energy metabolic systems distinct to other bacteria and have been reported to be particularly susceptible to energy metabolism disruptors, such as bedaquiline, clofazimine, and telacebec (Q203). We hope this research will inspire further studies aimed at developing novel antimycobacterial strategies based on energetics disruption.