Biomarkers of Response to the Glutaminase Inhibitor CB-839 in Multiple Myeloma Cells

• Published in: Blood 2014-12, Vol.124 (21), p.3429-3429
• Main Authors: Mackinnon, Andrew, Bennett, Mark, Rodriguez, Mirna, Parlati, Francesco
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V124.21.3429.3429

CB-839 is a potent, orally bioavailable small-molecule inhibitor of the tumor metabolism target glutaminase (GLS) that is currently in Phase 1 clinical trials for the treatment of solid and hematological malignancies. GLS is a mitochondrial enzyme that converts Gln to glutamate (Glu) to support several metabolic processes including amino acid synthesis, maintenance of cellular redox homeostasis, and the replacement of TCA cycle intermediates. CB-839 has in vitro antiproliferative activity across a sub-set of cell lines from diverse hematological tumor types including multiple myeloma (MM), acute lymphocytic leukemia, and non–Hodgkin’s lymphoma [Parlati et al. Blood 2013 122:4226]. To identify biomarkers that would predict sensitivity to CB-839 in MM, we profiled cellular metabolites, mRNA transcripts, and oncogenic signaling pathways in eight MM cell lines representing four CB-839-sensitive cell lines (RPMI8226, MM.1S, KMS-11, and IM-9) and four CB-839-resistant cell lines (AMO-1, L-363, KMS-28PE, and OPM-2). CB-839 treatment for 4 hours significantly decreased the levels of amino acids (Glu, aspartate, proline) as well as TCA cycle intermediates (fumarate, malate, succinate) across all cell lines. However, prior to treatment, CB-839-sensitive cells had significantly lower baseline levels of pyruvate-, fumarate-, and succinyl-CoA-derived amino acids compared to CB-839-resistant cells. In addition, both the adenylate and guanylate energy charges, a measure of cellular metabolic activity [Atkinson and Walton, J. Biol. Chem. 1967 242: 3239-41], were significantly lower in CB-839 sensitive cells. These observations suggest that cells with low levels of amino acids and/or low cellular energy charge are more susceptible to the pharmacological effects of CB-839. Reverse phase protein array and immunoblot analysis were used to evaluate the impact of CB-839 on signaling pathways across the panel of cell lines. Consistent with the observed decreases in amino acid pools, CB-839 treatment for 4 hours led to an acute inhibition of the amino-acid sensing kinase mTORC1 across all cell lines, as evidenced by decreased phosphorylation of p70S6K and S6. CB-839 treatment also reduced the levels of c-Myc across all cell lines, consistent with a block in five-prime cap-dependent translation as a likely consequence of mTORC1 inhibition. However, untreated sensitive cells had lower baseline levels of both 4E-BP1 and phospho-4E-BP1 in comparison to resistant cells, and at 24 hour