Abstract P2-09-03: Antitumor activity of the glutaminase inhibitor, CB-839, in triple-negative breast cancer
Authors: Gross M , Budczies J , Demo S , Janes J , Lewis E , Parlati F , MacKinnon A , Rodriguez M , Yang J , Zhao F , Richardson A , Denkert C , Fiehn O , Bennett M 
Published: August 2014

Glutamine (Gln) serves as an important source of energy and building blocks for many tumor cells. The first step in Gln utilization is its conversion to glutamate (Glu) by the mitochondrial enzyme glutaminase. CB-839 is a potent, selective, and orally bioavailable inhibitor of both splice variants of glutaminase (KGA and GAC). CB-839 inhibits recombinant GAC with an IC50 of &lt;15 nM. Treatment of the triple-negative breast cancer (TNBC) cell line MDA-MB-231 in vitro with CB-839 reduced cell viability and suppressed cellular Glu pools with comparable potency (IC50 ∼15 nM). TNBC breast cancer cell lines have been reported to have a heightened dependence on exogenous Gln for growth relative to estrogen receptor (ER) positive cells (Kung et al, 2011). Consistent with this observation, an analysis of 58 breast cancer cell lines in the Cancer Cell Line Encyclopedia revealed that low ER and/or Her2 expression is correlated with high glutaminase expression (particularly the GAC isoform) and low expression of glutamine synthetase (GLUL), an enzyme that converts Glu to Gln. This reciprocal pattern of GAC and GLUL expression suggests that TNBC may be particularly sensitive to glutaminase inhibition. To test this hypothesis, the anti-tumor activity of CB-839 was evaluated on a panel of breast tumor cell lines (n&gt;25) that included a mixture of TNBC and ER/Her2-positive subtypes. The TNBC subtype displayed the greatest sensitivity to CB-839 treatment (IC50s ranging from 5-100 nM) and this sensitivity was correlated with: i) dependence on extracellular Gln for growth (r = 0.83, p&lt;0.0001); ii) GAC (but not KGA) expression (r = -0.76, p&lt;0.0001); iii) glutaminase activity in cell lysates (r = -0.75, p = 0.0002); and iv) the ratio of cellular Glu to Gln (r = 0.53, p = 0.014) indicative of efficient Gln to Glu conversion. CB-839 also displayed significant anti-tumor activity in two TNBC xenograft models: 1) as a single agent in a patient-derived tumor chosen based on Gln utilization biomarkers and 2) in a cell line model (JIMT1) both as a single agent and in combination with paclitaxel. To determine if potential markers of Gln utilization and CB-839 sensitivity are reflected in primary tumors, an analysis of large mRNA expression and metabolomic datasets was undertaken. Consistent with the observations in cell lines, there was significantly elevated expression of GAC and reduced expression of GLUL in TNBC tumors relative to the ER+ population in The Cancer Genome Atlas mRNA expression dataset (n = 756). Likewise, analysis of the metabolite levels in 262 primary breast tumors showed that the Glu to Gln ratio in the ER- populations was significantly elevated relative to the ER+ population (fold increase = 3.2, p = 8.0E-09). This elevated metabolite ratio in the ER- population was also significantly correlated with the mRNA expression level of GLUL (r = -0.27, p = 0.001). These observations suggest that biomarkers of Gln utilization and CB-839 sensitivity extend to primary TNBC tumors. Together with the in vitro and in vivo preclinical anti-tumor activity of CB-839 in TNBC models, this primary tumor biomarker evaluation provides a strong rationale for the clinical investigation of CB-839 as a targeted therapeutic in TNBC patients.

Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-09-03.