A comparative PET imaging study with the reversible and irreversible EGFR tyrosine kinase inhibitors [(11)C]erlotinib and [(18)F]afatinib in lung cancer-bearing mice

Abstract

Background: Tyrosine kinase inhibitors (TKIs) have experienced a tremendous boost in the last decade, where more than 15 small molecule TKIs have been approved by the FDA. Unfortunately, despite their promising clinical successes, a large portion of patients remain unresponsive to these targeted drugs. For non-small cell lung cancer (NSCLC), the effectiveness of TKIs is dependent on the mutational status of epidermal growth factor receptor (EGFR). The exon 19 deletion as well as the L858R point mutation lead to excellent sensitivity to TKIs such as erlotinib and gefitinib; however, despite initial good response, most patients invariably develop resistance against these first-generation reversible TKIs, e.g., via T790M point mutation. Second-generation TKIs that irreversibly bind to EGFR wild-type and mutant isoforms have therefore been developed and one of these candidates, afatinib, has now reached the market. Whether irreversible TKIs differ from reversible TKIs in their in vivo tumor-targeting properties is, however, not known and is the subject of the present study.

Methods: Erlotinib was labeled with carbon-11 and afatinib with fluorine-18 without modifying the structure of these compounds. A preclinical positron emission tomography (PET) study was performed in mice bearing NSCLC xenografts with a representative panel of mutations: an EGFR-WT xenograft cell line (A549), an acquired treatment-resistant L858R/T790M mutant (H1975), and a treatment-sensitive exon 19 deleted mutant (HCC827). PET imaging was performed in these xenografts with both tracers. Additionally, the effect of drug efflux transporter permeability glycoprotein (P-gp) on the tumor uptake of tracers was explored by therapeutic blocking with tariquidar.

Results: Both tracers only demonstrated selective tumor uptake in the HCC827 xenograft line (tumor-to-background ratio, [(11)C]erlotinib 1.9 ± 0.5 and [(18)F]afatinib 2.3 ± 0.4), thereby showing the ability to distinguish sensitizing mutations in vivo. No major differences were observed in the kinetics of the reversible and the irreversible tracers in each of the xenograft models. Under P-gp blocking conditions, no significant changes in tumor-to-background ratio were observed; however, [(18)F]afatinib demonstrated better tumor retention in all xenograft models.

Conclusions: TKI-PET provides a method to image sensitizing mutations and can be a valuable tool to compare the distinguished targeting properties of TKIs in vivo.

Keywords: Activated EGFR; Afatinib; Erlotinib; PET; Personalized medicine; Radiochemistry.

Figures

Figure 1

Chemical structures of erlotinib and afatinib.

Scheme 1

Radiosynthesis of [11C]erlotinib. TBAOH, tetrabutylammonium hydroxide; DMF, dimethylformamide.

Scheme 2

Radiosynthesis of [18F]6 and subsequent condensation towards [18F]afatinib. MeCN, acetonitrile; MeOH, methanol; DBU, 1,8-diazabicyclo[5.4.0]undec-7-een; NMP, N-methylpyrrolidone.

Figure 2

Immunohistochemical staining of NSCLC xenograft lines as used in PET studies. Images depicted at 5× magnification. Mutations: A549 (wild type), H1975 (L858R/T790M), and HCC827 (exon 19 deletion).

Figure 3

PET images and TACs of [11C]erlotinib (top) and [18F]afatinib (bottom). Circle indicates tumor position and arrow indicates reference tissue. [11C]erlotinib images summed from 60 to 90 min, [18F]afatinib images from 90 to 120 min. TACs are averaged over three animals. Mutations: A549 (wild type), H1975 (L858R/T790M), and HCC827 (exon 19 deletion). Tumor and background were manually outlined on the basis of an additional [18F]FDG scan, which was performed directly after the TKI-PET scan was finished.

Figure 4

Whole brain TACs of [11C]erlotinib (left) and [18F]afatinib (right) before and after tariquidar pre-treatment. TACs are averaged over three animals.

Figure 5

TACs of [11C]erlotinib (top) and [18F]afatinib (bottom) after tariquidar treatment. TACs are averaged over three animals. Mutations: A549 (wild type), H1975 (L858R/T790M), and HCC827 (exon 19 deletion). Tumor and background were manually outlined on the basis of an additional [18F]FDG scan, which was performed directly after the TKI-PET scan was finished.