Novel nucleoside analogs exhibit potent intracellular andactivities against.

Abstract

is a major causative agent of nontuberculous mycobacterial pulmonary disease, which poses therapeutic challenges owing to its intrinsic drug resistance and the need for prolonged multidrug regimens. In this study, we identified two novel nucleoside analogs, MCCB-04-35 and MCCB-04-37, as potential therapeutic candidates againstinfection. Both compounds exhibited significant bacteriostatic activityand in infected macrophages, with minimal cytotoxicity. Time-kill kinetics and MIC assays confirmed their potent inhibitory effects, particularly against slow-growing mycobacteria. Checkerboard synergy testing revealed additive to synergistic interactions with clinically used antibiotics such as clarithromycin and ciprofloxacin. In a mouse model of chronic lung infection, both compounds significantly reduced pulmonary bacterial burden, inflammatory cytokine levels, and histopathological damage. Transcriptomic analysis of treatedrevealed the downregulation of key metabolic pathways, including oxidative phosphorylation and nitrogen metabolism, indicating disruption of intracellular energy homeostasis. These findings suggest that MCCB-04-35 and MCCB-04-37 exert antimicrobial effects through metabolic interference and may serve as effective therapeutic agents either alone or in combination for treatinginfections.IMPORTANCEPulmonary disease caused by(MAC) is notoriously difficult to treat due to intrinsic antibiotic resistance and the need for prolonged multidrug therapy, often poorly tolerated with suboptimal outcomes. The identification of new therapeutic candidates with novel mechanisms of action is urgently needed. Here, we report two novel nucleoside analogs, MCCB-04-35 and MCCB-04-37, exhibiting strong anti-mycobacterial activity againstbothand, with minimal cytotoxicity. These compounds showed additive to synergistic effects when combined with existing antibiotics such as clarithromycin. In a mouse model of chronic lung infection, they significantly reduced bacterial burden, inflammation, and tissue damage. Transcriptomic profiling revealed downregulation of metabolic pathways essential for bacterial energy production, suggesting a unique mechanism of antimicrobial action. Our findings provide promising leads for the development of more effective treatments for MAC pulmonary disease, either as monotherapy or in combination with current drugs.