MmpL3-IN-1, also known as compound 32, is a highly effective inhibitor of Mycobacterial membrane protein large 3 (MmpL3). With an anti-tuberculosis activity exhibiting a minimum inhibitory concentration (MIC) of less than 0.016 μg/mL in M. tuberculosis, MmpL3-IN-1 is a valuable compound for research on drug-resistant tuberculosis [1].

MmpL3-IN-1 (compound 32) demonstrated potent anti-M. tuberculosis activity with a minimal inhibitory concentration (MIC) below 0.016 μg/mL over a treatment duration of 2-7 days at dosages of 0.26-64 μg/mL, showing minimal toxicity towards Vero cells. This compound exhibited good microsomal stability and negligible inhibition of hERG K+ channels with an IC50 exceeding 30 μM. At concentrations of 0.0625-1 μg/mL over 16 hours, MmpL3-IN-1 specifically inhibited the transport of trehalose monomycolate (TMM), disrupting M. tuberculosis cell wall formation by targeting MmpL3. Assays revealed that, in Vero cells treated for 48 hours with 0.26-64 μg/mL, cell viability was inhibited, showing an IC50 value of 35.3 μg/mL. A western blot analysis on M. tuberculosis H37Rv mc^26206 exposed to 0.0625-1 μg/mL for 16 hours highlighted a dose-dependent increase in TMM and a decrease in cell wall-bound mycolic acid methyl esters (MAMEs), indicating that high concentrations of MmpL3-IN-1 reduce the synthesis of trehalose dimycolate (TDM) and accumulate free mycolate.

MmpL3-IN-1 (compound 32), when administered orally at a dosage of 100 mg/kg for 5 days a week over a period of 30 days, demonstrated potent anti-tuberculosis efficacy in SPF BALB/c female mice infected with H37Rv, achieving a significant 2.0 log reduction in lung colony-forming units of H37Rv. In addition, when administered either orally (p.o.) at 100 mg/kg or intravenously (i.v.) at 10 mg/kg, MmpL3-IN-1 exhibited notable pharmacokinetic properties, including half-life (t 1/2), maximum concentration (Cmax), time to reach maximum concentration (Tmax), area under the plasma drug concentration-time curve from 0 to the last measurable concentration (AUC 0-t), mean residence time (MRT 0-t), volume of distribution (V), clearance (CL), and oral bioavailability (F%). Specifically, oral administration showed a half-life of about 7.48 hours with peak concentrations reached within 0.5 hours, while intravenous administration demonstrated a quicker Tmax at 0.03 hours, reflecting its rapid absorption and distribution within the body.