Fig. 1. Resistance to EGFR TKIs in preclinical models and GBM patients treated with an EGFR TKI is associated with a decreasing ratio of EGFRvIII^High/EGFRvIII^Low tumor cells.

(A) FACS-sorted EGFRvIII^High and EGFRvIII^Low cells obtained from GBM39 differ in their PI3K-Akt-mTOR activity as determined by immunoblotting. (B) Immunofluorescence (IF) for EGFRvIII and K_i-67 on isolated GBM39 tumor cells shows differences in basal proliferative rate between EGFRvIII^High and EGFRvIII^Low tumor cells. *P < 0.005. (C) Terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick end labeling (TUNEL) stain and EGFRvIII IF indicate a higher basal apoptosis in EGFRvIII^Low tumor cells. *P < 0.005. (D and E) Radiopharmaceutical imaging chip analysis of ¹⁸F-fluorodeoxyglucose from FACS-sorted EGFRvIII^High and EGFRvIII^Low cells indicates higher glucose uptake in EGFRvIII^High cells. **P < 0.05. (F) FACS-sorted EGFRvIII^High and EGFRvIII^Low were treated with erlotinib (5 μM) for 24 hours, and cell viability was determined by trypan blue exclusion assay. **P < 0.05. (G and H) Resistance to erlotinib in GBM39 xenografts (n = 4 mice per group). During initial response (blue curve) and at the time of resistance (red curve), there is a relative loss of EGFRvIII-expressing tumor cells. (I and J) In GBM patients, 10 days of treatment with the EGFR tyrosine kinase inhibitor lapatinib reduces EGFRvIII expression relative to pretreatment levels. *P < 0.01; **P < 0.0001; ^#P < 0.001. All values are mean ± SEM. P values were obtained from unpaired t test.