Molecular imaging of active mutant L858R EGF receptor (EGFR) kinase-expressing nonsmall cell lung carcinomas using PET/CT

Abstract

The importance of the EGF receptor (EGFR) signaling pathway in the development and progression of nonsmall cell lung carcinomas (NSCLC) is widely recognized. Gene sequencing studies revealed that a majority of tumors responding to EGFR kinase inhibitors harbor activating mutations in the EGFR kinase domain. This underscores the need for novel biomarkers and diagnostic imaging approaches to identify patients who may benefit from particular therapeutic agents and approaches with improved efficacy and safety profiles. To this goal, we developed 4-[(3-iodophenyl)amino]-7-{2-[2-{2-(2-[2-{2-([(18)F]fluoroethoxy)-ethoxy}-ethoxy]-ethoxy)-ethoxy}-ethoxy]-quinazoline-6-yl-acrylamide ([(18)F]F-PEG6-IPQA), a radiotracer with increased selectivity and irreversible binding to the active mutant L858R EGFR kinase. We show that PET with [(18)F]F-PEG6-IPQA in tumor-bearing mice discriminates H3255 NSCLC xenografts expressing L858R mutant EGFR from H441 and PC14 xenografts expressing EGFR or H1975 xenografts with L858R/T790M dual mutation in EGFR kinase domain, which confers resistance to EGFR inhibitors (i.e., gefitinib). The T790M mutation precludes the [(18)F]F-PEG6-IPQA from irreversible binding to EGFR. These results suggest that PET with [(18)F]F-PEG6-IPQA could be used for the selection of NSCLC patients for individualized therapy with small molecular inhibitors of EGFR kinase that are currently used in the clinic and have a similar structure (i.e., iressa, gefitinib, and erlotinib).

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Characterization of PC14, H441, H3255, and H1975 tumor cells lines in vitro. The level of (A) total EGFR and (B) phospho-EGFR. Time-dependent accumulation of [18F]F-PEG6-IPQA (C) at baseline and (D) in the presence of 100 μM/L iressa.

Fig. 2.

Assessment of irreversible binding of [18F]F-PEG6-IPQA to WT and mutant EGFR kinases. (A) The autoradiogram of a Western blot membrane and (B) chemiluminescent image of the same membrane stained with anti-EGFR kinase antibody. Individual lanes are labeled with corresponding cell lines and dilutions of protein extracts (the H1975 and PC14 extracts were undiluted). (C) Inhibition of substrate phosphorylation by recombinant WT (red squares), L858R active mutant (blue triangles), and L585R/T790M dual-mutant (green triangles) EGFR kinases in the presence of different concentrations of F-PEG6-IPQA; the IC50 concentrations are indicated next to the enzyme labels.

Fig. 3.

Representative coronal and axial PET/CT images obtained 120 min postinjection of [18F]F-PEG6-IPQA in two different mice bearing tumor xenografts in the opposite shoulders (arrows). Color coding of the images is set to maximize the visualization of tumors in each projection.

Fig. 4.

Time-activity curves of [18F]F-PEG6-IPQA–derived radioactivity (percent ID per gram) in different tumors (A) at baseline and (B) after pretreatment of animals with iressa. Quantification of [18F]F-PEG6-IPQA accumulation in different tumor xenografts at baseline (blue bars) and after pretreatment with iressa (red bars). (C) The rate of unidirectional accumulation (Ki) of [18F]F-PEG6-IPQA determined using Patlak graphical analysis and (D) binding potential (BP) of [18F]F-PEG6-IPQA determined by Logan model-independent graphical analysis. Bars are SEs; statistically significant differences (P < 0.05) are indicated by an asterisk.