Structure guided generation of thieno[3,2-d]pyrimidin-4-amine Mycobacterium tuberculosis bd oxidase inhibitors

Cytochrome bd oxidase (Cyt-bd) is an attractive drug target in Mycobacterium tuberculosis, especially in the context of developing a drug combination targeting energy metabolism. However, currently few synthetically assessable scaffolds target Cyt-bd. Herein, we report that thieno[3,2-d]pyrimidin-4-amines inhibit Cyt-bd, and report an initial structure-activity-relationship (SAR) of 13 compounds in three mycobacterial strains: Mycobacterium bovis BCG, Mycobacterium tuberculosis H37Rv and Mycobacterium tuberculosis clinical isolate N0145 in an established ATP depletion assay with or without the cytochrome bcc : aa ₃ (QcrB) inhibitor Q203. All compounds displayed activity against M. bovis BCG and the M. tuberculosis clinical isolate strain N0145 with ATP IC₅₀ values from 6 to 54 μM in the presence of Q203 only, as expected from a Cyt-bd inhibitor. All derivatives were much less potent against M. tuberculosis H37Rv compared to N0145 (IC₅₀'s from 24 to >100 μM and 9-52 μM, respectively), an observation that may be attributed to the higher expression of the Cyt-bd-encoding genes in the laboratory-adapted M. tuberculosis H37Rv strain. N-(4-(tert-butyl)phenethyl)thieno[3,2-d]pyrimidin-4-amine (19) was the most active compound with ATP IC₅₀ values from 6 to 18 μM against all strains in the presence of Q203, making it a good chemical probe for interrogation the function of the mycobacterial Cyt-bd under various physiological conditions.