Exploring pitfalls of 64Cu-labeled EGFR-targeting peptide GE11 as a potential PET tracer

The epidermal growth factor receptor (EGFR) represents an important molecular target for both radiotracer-based diagnostic imaging and radionuclide therapy of various cancer entities. For the delivery of radionuclides to the tumor, peptides hold great potential as a transport vehicle. With respect to EGFR, the peptide YHWYGYTPQNVI (GE11) has been reported to bind the receptor with high specificity and affinity. In the present study, GE11 with β-alanine (β-Ala-GE11) was conjugated to the chelating agent p -SCN-Bn-NOTA and radiolabeled with 64 Cu for the first radio pharmacological evaluation as a potential probe for positron emission tomography (PET)-based cancer imaging. For better water solubility, an ethylene glycol-based linker was introduced between the peptide’s N terminus and the radionuclide chelator. The stability of the 64 Cu-labeled peptide conjugate and its binding to EGFR-expressing tumor cells was investigated in vitro and in vivo, and then compared with the 64 Cu-labeled EGFR-targeting antibody conjugate NOTA-cetuximab. The GE11 peptide conjugate [ 64 Cu]Cu-NOTA-linker-β-Ala-GE11 ([ 64 Cu]Cu- 1 ) was stable in a buffer solution for at least 24 h but only 50% of the original compound was detected after 24 h of incubation in human serum. Stability could be improved by amidation of the peptide’s C terminus (β-Ala-GE11-NH 2 ( 2 )). Binding assays with both conjugates, [ 64 Cu]Cu- 1 and [ 64 Cu]Cu- 2 , using the EGFR-expressing tumor cell lines A431 and FaDu showed no specific binding. A pilot small animal PET investigation in FaDu tumor-bearing mice revealed only low tumor uptake (standard uptake value (SUV)