title: 13C isotope-assisted methods for quantifying glutamine metabolism in cancer cells.
abstract: Glutamine has recently emerged as a key substrate to support cancer cell proliferation, and the quantification of its metabolic flux is essential to understand the mechanisms by which this amino acid participates in the metabolic rewiring that sustains the survival and growth of neoplastic cells. Glutamine metabolism involves two major routes, glutaminolysis and reductive carboxylation, both of which begin with the deamination of glutamine to glutamate and the conversion of glutamate into α-ketoglutarate. In glutaminolysis, α-ketoglutarate is oxidized via the tricarboxylic acid cycle and decarboxylated to pyruvate. In reductive carboxylation, α-ketoglutarate is reductively converted into isocitrate, which is isomerized to citrate to supply acetyl-CoA for de novo lipogenesis. Here, we describe methods to quantify the metabolic flux of glutamine through these two routes, as well as the contribution of glutamine to lipid synthesis. Examples of how these methods can be applied to study metabolic pathways of oncological relevance are provided.
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Woo Suk
authors.initials: WS
authors.lastname: Ahn
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Paulo A
authors.initials: PA
authors.lastname: Gameiro
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Keibler
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Zhe
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA. Electronic address: gregstep@mit.edu.
authors.forename: Gregory
authors.initials: G
authors.lastname: Stephanopoulos
mesh_terms: D001671:Biochemistry / Q000379:methods*; D002247:Carbon Isotopes; D015687:Cell Hypoxia; D002478:Cells, Cultured; D008401:Gas Chromatography-Mass Spectrometry; D018698:Glutamic Acid / Q000032:analysis* / Q000378:metabolism; D005973:Glutamine / Q000032:analysis* / Q000378:metabolism*; D006801:Humans; D008055:Lipids / Q000096:biosynthesis; D009369:Neoplasms / Q000378:metabolism*
chemical_list: D002247:Carbon Isotopes; D008055:Lipids; D005973:Glutamine; D018698:Glutamic Acid
keywords: Cancer cell; Glutamine metabolism; Metabolic flux; Reductive carboxylation; Stable isotopic tracer; Tricarboxylic acid cycle