Assesment of EGFR somatic kinase domain mutant tumor response to erlotinib using FDG-PET imaging

Abstract

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Somatic mutations in the epidermal growth factor receptor (EGFR) have been identified in non-small lung cancer patients and are correlated with enhanced sensitivity to the tyrosine kinase domain inhibitor erlotinib (OSI-774, Tarceva^R). Erlotinib is a selective, orally available competitive inhibitor for ATP binding to the EGFR tyrosine kinase domain and has been approved for use in the treatment of advanced non-small cell lung cancer for patients who have failed at least one prior chemotherapy regimen. To evaluate the biological role of EGFR somatic mutations and the influence of these mutations on erlotinib sensitivity, we have generated stable NR6 cell lines expressing equivalent levels of EGFR, and two clinically detected EGFR somatic mutations, EGFR-L858R and EGFR-Del(746-752). NR6 cells do not express EGFR or detectable levels of ErbB2, ErbB3, or ErbB4. Both the EGFR-L858R and EGFR-Del(746-752) mutations facilitate ligand-dependent anchorage-independent growth in soft agar, confer a higher proliferative rate, form tumors in immune compromised mice, and are differentially sensitive to erlotinib compared to wild-type EGFR both in vivo and in vitro. In this study, we have used positron emission tomography (PET) imaging of 2-fluoro-2-deoxy-D-glucose (FDG) uptake to evaluate the difference in metabolic activity of EGFR, EGFR-L858R, and EGFR-Del(746-750) expressing tumors in immune deficient mice. Tumors expressing wild-type and mutant EGFR have similar glucose uptake rates except after a period of overnight fasting where tumors expressing EGFR kinase domain mutations demonstrate a 2-fold increase in FDG uptake compared to wild-type EGFR tumors (p=0.001). ³H-FDG uptake in EGFR expressing cells is significantly higher compared to the NR6 parental cell line and erlotinib differentially inhibits glucose uptake in vitro for EGFR compared to EGFR-L858R and EGFR Del(746-752) (IC₅₀=175nM, 85nM, and 9nM respectively). We find that FDG-PET is useful for evaluating the sensitivity of these mutations and the EGFR Del(746-752) and L858R mutations are significantly more sensitive to erlotinib compared to wild-type EGFR tumors. At the maximally effective dose of erlotinib (150mg/kg), the normalized glucose uptake decreased by 33% at 24 hours post-treatment and 57% at 48 hours in the tumors expressing EGFR-Del(746-752) compared with no significant decrease in the EGFR tumors. Our data indicate that EGFR somatic mutations influence EGFR signaling pathways that modulate glucose metabolism. These findings suggest FDG-PET is a sensitive method to evaluate the effects of small molecule kinase inhibitors and may be useful in evaluating clinical responders to erlotinib therapy.