Despite being crucial for energy generation in most forms of life, few if any microbial antibiotics specifically inhibit glycolysis. To develop a specific inhibitor of the glycolytic enzyme enolase 2 (ENO2) for the treatment of cancers with deletion of ENO1 (encoding enolase 1), we modeled the synthetic tool compound inhibitor phosphonoacetohydroxamate (PhAH) into the active site of human ENO2. A ring-stabilized analog of PhAH, in which the hydroxamic nitrogen is linked to Cα by an ethylene bridge, was predicted to increase binding affinity by stabilizing the inhibitor in a bound conformation. Unexpectedly, a structure-based search revealed that our hypothesized backbone-stabilized PhAH bears strong similarity to SF2312, a phosphonate antibiotic of unknown mode of action produced by the actinomycete Micromonospora, which is active under anaerobic conditions. Here, we present multiple lines of evidence, including a novel X-ray structure, that SF2312 is a highly potent, low-nanomolar inhibitor of enolase.
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We thank J. Holton and G. Meigs for their assistance with X-ray diffraction data collection at the Advanced Light Source, beamline 8.3.1. We thank K. Kalhurachi and J. McMurray for assistance with 13C-NMR measurements. We thank S. Millward, J. Marszalek and Y. Sun for critical comments and suggestions. We thank M. Yuan and S. Breitkopf for help with MS experiments. We thank M. Protopopova for assistance with Operetta cell imaging. We thank L. Lebioda, G. Reed and R. Poyner for useful discussions about the catalytic mechanism of enolase. We thank R. Zielinski for assistance with hypoxia experiments. The cell line D423-MG (D423) was kindly shared by D. Bigner (Duke University Medical Center) while the Gli56 cell line was shared by D. Louis (Massachusetts General Hospital) and TS neurospheres were shared by C. Brennan (Memorial Sloan Kettering Cancer Center). Inhibitors were provided by the Pharmaceutical Chemistry Facility at MD Anderson, supported by the US National Institutes of Health (NIH) National Cancer Institute under award number P30CA016672. Financial support was provided by NIH-NCI grant 7P01CA095616-10 (R.A.D.); Cancer Prevention Research Institute of Texas (CPRIT) grant RP140612 (R.A.D.); NCI grants 5P01CA120964 (J.M.A.) and 5P30CA006516 (J.M.A.) and NIH Career Development Project through the Specialized Programs of Research Excellence (SPORE) P50CA127001-07 (F.L.M.). F.L.M. was supported by Research Scholar Grant RSG-15-145-01-CDD from the American Cancer Society.
The authors declare no competing financial interests.
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Leonard, P., Satani, N., Maxwell, D. et al. SF2312 is a natural phosphonate inhibitor of enolase. Nat Chem Biol 12, 1053–1058 (2016) doi:10.1038/nchembio.2195
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