24-Methylenecyclopropane steroidal inhibitors: A Trojan horse in ergosterol biosynthesis that prevents growth of Trypanosoma brucei

Matthew B. Miller, Presheet Patkar, Ujjal K. Singha, Minu Chaudhuri, W. David Nes

Research output: Contribution to journalArticlepeer-review

6 Scopus citations


A new class of steroidal therapeutics based on phylogenetic-guided design of covalent inhibitors that target parasite-specific enzymes of ergosterol biosynthesis is shown to prevent growth of the protozoan-Trypanosoma brucei, responsible for sleeping sickness. In the presence of approximately 15 ± 5 μM 26,27-dehydrolanosterol, T. brucei procyclic or blood stream form growth is inhibited by 50%. This compound is actively converted by the parasite to an acceptable substrate of sterol C24-methyl transferase (SMT) that upon position-specific side chain methylation at C26 inactivates the enzyme. Treated cells show dose-dependent depletion of ergosterol and other 24β-methyl sterols with no accumulation of intermediates in contradistinction to profiles typical of tight binding inhibitor treatments to azoles showing loss of ergosterol accompanied by accumulation of toxic 14-methyl sterols. HEK cells accumulate 26,27-dehydrolanosterol without effect on cholesterol biosynthesis. During exposure of cloned TbSMT to 26,27-dehydrozymosterol, the enzyme is gradually inactivated (kcat/kinact = 0.13 min− 1/0.08 min− 1; partition ratio of 1.6) while 26,27-dehydrolanosterol binds nonproductively. GC–MS analysis of the turnover product and bound intermediate released as a C26-methylated diol (C3-OH and C24-OH) confirmed substrate recognition and covalent binding to TbSMT. This study has potential implications for design of a novel class of chemotherapeutic leads functioning as mechanism-based inhibitors of ergosterol biosynthesis to treat neglected tropical diseases.

Original languageEnglish
Pages (from-to)305-313
Number of pages9
JournalBiochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Issue number3
StatePublished - Mar 1 2017


  • Ergosterol biosynthesis inhibitor
  • Human African trypanosomiasis
  • Sleeping sickness
  • Sterol C24-methyltransferase
  • Suicide substrate


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