A High-Throughput Fluorimetric Assay for 2-Hydroxyglutarate Identifies Zaprinast as a Glutaminase Inhibitor

Recently identified IDH mutations lead to the production of 2-hydroxyglutarate (2HG), an onco-metabolite aberrantly elevated in selected cancers. We developed a facile and inexpensive fluorimetric microplate assay for quantitation of 2HG and performed an unbiased small molecule screen in live cells to identify compounds capable of perturbing 2HG production. Zaprinast, a PDE5 inhibitor, was identified as an efficacious modulator of 2HG production and confirmed to lower 2HG levels in vivo. The mechanism of action was not due to cGMP stabilization, but rather, profiling of metabolites upstream of mutant IDH1 pointed to targeted inhibition of the enzyme glutaminase (GLS). Zaprinast treatment reversed histone hypermethylation and soft agar growth of IDH1 mutant cells, and treatment of glutamine-addicted pancreatic cancer cells reduced growth and sensitized cells to oxidative damage. Thus, Zaprinast is efficacious against glutamine metabolism and further establishes the therapeutic linkages between GLS and 2HG-mediated oncogenesis.