MYC-driven accumulation of 2-hydroxyglutarate is associated with breast cancer prognosis
Atsushi Terunuma, … , Arun Sreekumar, Stefan Ambs
Atsushi Terunuma, … , Arun Sreekumar, Stefan Ambs
Published December 9, 2013
Citation Information: J Clin Invest. 2014;124(1):398-412. https://doi.org/10.1172/JCI71180.
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Research Article Oncology

MYC-driven accumulation of 2-hydroxyglutarate is associated with breast cancer prognosis

Abstract

Metabolic profiling of cancer cells has recently been established as a promising tool for the development of therapies and identification of cancer biomarkers. Here we characterized the metabolomic profile of human breast tumors and uncovered intrinsic metabolite signatures in these tumors using an untargeted discovery approach and validation of key metabolites. The oncometabolite 2-hydroxyglutarate (2HG) accumulated at high levels in a subset of tumors and human breast cancer cell lines. We discovered an association between increased 2HG levels and MYC pathway activation in breast cancer, and further corroborated this relationship using MYC overexpression and knockdown in human mammary epithelial and breast cancer cells. Further analyses revealed globally increased DNA methylation in 2HG-high tumors and identified a tumor subtype with high tissue 2HG and a distinct DNA methylation pattern that was associated with poor prognosis and occurred with higher frequency in African-American patients. Tumors of this subtype had a stem cell–like transcriptional signature and tended to overexpress glutaminase, suggestive of a functional relationship between glutamine and 2HG metabolism in breast cancer. Accordingly, 13C-labeled glutamine was incorporated into 2HG in cells with aberrant 2HG accumulation, whereas pharmacologic and siRNA-mediated glutaminase inhibition reduced 2HG levels. Our findings implicate 2HG as a candidate breast cancer oncometabolite associated with MYC activation and poor prognosis.

Authors

Atsushi Terunuma, Nagireddy Putluri, Prachi Mishra, Ewy A. Mathé, Tiffany H. Dorsey, Ming Yi, Tiffany A. Wallace, Haleem J. Issaq, Ming Zhou, J. Keith Killian, Holly S. Stevenson, Edward D. Karoly, King Chan, Susmita Samanta, DaRue Prieto, Tiffany Y.T. Hsu, Sarah J. Kurley, Vasanta Putluri, Rajni Sonavane, Daniel C. Edelman, Jacob Wulff, Adrienne M. Starks, Yinmeng Yang, Rick A. Kittles, Harry G. Yfantis, Dong H. Lee, Olga B. Ioffe, Rachel Schiff, Robert M. Stephens, Paul S. Meltzer, Timothy D. Veenstra, Thomas F. Westbrook, Arun Sreekumar, Stefan Ambs

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Figure 8

Glutamine metabolism is linked to aberrant accumulation of 2HG in breast cancer cells.

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Glutamine metabolism is linked to aberrant accumulation of 2HG in breast...
(A) Incorporation of C13-labeled glutamine into 2HG in MDA-MB-231 (2HG-high) and MCF7 (2HG-low) cells 10 seconds and 3 hours after adding C13-glutamine to the culture medium (n = 3 each). MDA-MB-231 cells, with aberrant 2HG accumulation, incorporated the C13 label into 2HG. (B) Reduced 2HG in breast cancer cells treated with GLS1 siRNA (n = 4 per group; t test). Cell pellets were harvested 48 hours after transfection. (C) Reduced 2HG and glutamate in breast cancer cells treated with the mitochondrial GLS1 inhibitor compound 968 (n = 3 per group; *P < 0.05 vs. control, t test). Cells were treated with 10 μM inhibitor for 48 hours. (D) Increased intracellular 2HG after c-Myc induction (+MYC) in human mammary epithelial cells with an inducible MYC-ER fusion transgene. Cell pellets were harvested 48 hours after induction of c-Myc (3 independent experiments; t test). (E) Knockdown of c-Myc with 2 different doxycycline-inducible shRNA expression constructs (right) caused a significant reduction of aberrantly accumulated 2HG (left) in SUM159T cells (4 independent experiments; t test). Cell pellets were harvested 3 days after shRNA induction. Values are shown normalized to internal standard. All graphs show mean ± SD. See complete unedited blots in the supplemental material.