Deciphering the mechanism of action of cholesterol analogs as inhibitors of Mycobacterium tuberculosis

Drug‐resistance continues to be a public health problem, with nearly half million people infected by multidrug‐resistant strain of Mycobacterium tuberculosis (Mtb) worldwide, urging the need to develop new antitubercular drugs. Cholesterol plays a critical role in phagocytosis by host macrophages. Mtb also utilizes cholesterol as a source of carbon and energy during infection and degrades it through three catabolic pathways acting on AB‐ and CD‐rings, and on the aliphatic side‐chain. These pathways produce acetyl‐ and propionyl‐CoA intermediates, which are then incorporated into cellular lipids and virulence factors. Cholesterol analogs with degradation‐resistant alkyl side‐chains, including 3β,16β,26‐trihydroxy‐cholesterol (16OHC), have been recently shown to effectively inhibit Mtb growth, but the mechanism of action remains unclear. In this study, we combined the use of synthetic cholesterol analogs with genetic and mass spectrometry approaches to demonstrate whether cholesterol side‐chain catabolic activities could be used as potential drug targets. Metabolomic profiling of Mtb wild‐type and mutant strains with decreased cholesterol side‐chain degrading activities fed with 16OHC revealed that this cholesterol analog is metabolized by all strains, except by Dcyp125 cells, to produce a metabolite that we identified as 3‐oxo‐chola‐1,4‐dien‐16β‐ol‐24‐oate methyl ester. Unexpectedly, our data showed that 16OHC is not an inhibitor of the cholesterol side‐chain degradation pathway, but rather an inhibitor of the AB‐ring degradation pathway. The inhibitory effect of 16OHC on selected recombinant Mtb AB‐ring degradation enzymes is being tested and data will be presented. Support or Funding Information Financial support from the Ralph & Kathleen Ponce de Leon Endowment at UTEP, NIH BBRC 5G12MD007592 and SC1 AI116567‐01A1 (NIH/NIAID/NIGMS) are gratefully acknowledged. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .