Cancer cells exhibit increased use of nutrients, including glucose and glutamine, to support the bioenergetic and biosynthetic demands of proliferation. We tested the small-molecule inhibitor of glutaminase CB-839 in combination with erlotinib on epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) as a therapeutic strategy to simultaneously impair cancer glucose and glutamine utilization and thereby suppress tumor growth. Here, we show that CB-839 cooperates with erlotinib to drive energetic stress and activate the AMP-activated protein kinase (AMPK) pathway in EGFR (del19) lung tumors. Tumor cells undergo metabolic crisis and cell death, resulting in rapid tumor regression in vivo in mouse NSCLC xenografts. Consistently, positron emission tomography (PET) imaging with ¹⁸F-fluoro-2-deoxyglucose (¹⁸F-FDG) and ¹¹C-glutamine (¹¹C-Gln) of xenografts indicated reduced glucose and glutamine uptake in tumors following treatment with CB-839 + erlotinib. Therefore, PET imaging with ¹⁸F-FDG and ¹¹C-Gln tracers can be used to non-invasively measure metabolic response to CB-839 and erlotinib combination therapy.