Pharmacologic properties and inhibitory activity of 6-azasteroids against <i>Mycobacterium leprae in vivo</i> and <i>in vitro</i>

Although the current multidrug therapy (MDT) for leprosy is very successful, the long treatment duration and the emergence of antibiotic-resistant strains demand for new alternative drugs. One potential target for drug development against pathogenic mycobacteria is their need to degrade host cholesterol during infection. <i>Mycobacterium leprae</i>, due to its degenerate genome, has preserved only the first step of cholesterol catabolism, in which cholesterol is oxidized to cholestenone by the enzyme 3β-hydroxysteroid dehydrogenase (3β-HSD). <i>M. leprae</i> avidly produces cholestenone <i>in vivo,</i> and this metabolic activity seems to play an important role in bacterial pathogenesis. In this study, six 6-azasteroid analogs were developed with the potential to inhibit 3β-HSD, and their metabolic stability and <i>in vitro</i> and <i>in vivo</i> inhibitory activity against <i>M. leprae</i> were investigated. Pharmacologic properties indicated lower metabolic liabilities for azasteroid <b>5</b> in comparison to its progenitors, azasteroids <b>1</b> and <b>2,</b> resulting in improved accumulation and extended <i>t</i>_1/2 values in mice. Azasteroid <b>5</b> also partially inhibits <i>M. leprae</i> 3β-HSD <i>in vitro</i>. When tested in the Shepard's mouse footpad model of leprosy, azasteroid <b>5</b> showed effective bacterial killing that was accelerated when combined with a subinhibitory dose of rifampicin and exhibited an absence of detectable hepatotoxic effects. We concluded that 6-azasteroids derivatives are promising new antimicrobial candidates for leprosy treatment.IMPORTANCELeprosy remains a significant global health challenge, particularly in underserved regions. While multidrug therapy (MDT) has been effective, its prolonged duration and the emergence of antibiotic-resistant strains emphasize the urgent need for novel therapeutic strategies. Recent advances in understanding <i>Mycobacterium leprae</i>'s unique biology have identified cholesterol metabolism as a critical pathway for bacterial survival and pathogenesis, offering a promising new target for drug development. Building on insights from tuberculosis research, azasteroids-compounds known for their potential to disrupt mycobacterial cholesterol metabolism-are now being explored as candidates for leprosy treatment. These molecules inhibit <i>M. leprae</i>'s cholesterol oxidation, impairing bacterial persistence within the host. This innovative approach could lead to more effective, faster-acting therapies, overcoming current treatment limitations and resistance. Such efforts represent a vital step forward in reducing the burden of leprosy and empowering affected communities worldwide.