Evaluation of an integrin αvβ6-specific peptide labeled with [18F]fluorine by copper-free, strain-promoted click chemistry

Introduction: Click chemistry, particularly the Huisgen 1,3-dipolar cycloaddition of an alkyne with an azide, has quickly become popular for site-specific radiolabeling. Recently, strain-promoted click chemistries have been developed, eliminating the need for potentially toxic copper catalysts. This study presents radiolabeling of an α(v)β(6) integrin targeting peptide (A20FMDV2) via strain-promoted click using a fluorine-18 prosthetic group, and in vitro and in vivo evaluation.

Methods: The radiotracer was prepared from and N(3)-PEG(7)-A20FMDV2 (ethanol; 10 min; 35-45 °C). HPLC-purified and formulated radiotracer 1 was evaluated in vitro by cell binding (DX3puroβ6, α(v)β(6)-positive; and DX3puro, α(v)β(6)-negative control) and serum stability, and in vivo using PET/CT imaging and biodistribution studies in mice.

Results: The radiotracer 1 was readily prepared and purified (from 2: 40±4 min including HPLC, 11.9±3.2% decay corrected isolated radiochemical yield, >99% radiochemical purity, n=4) and displayed good stability (1 h: >99%, saline; 94.6%, serum). Strong α(v)β(6)-targeted binding was observed in vitro (DX3puroβ6 cells, 15 min: 43.2% binding, >6:1 for DX3puroβ6:DX3puro). In the mouse model DX3puroβ6-tumor binding was low (1 h: 0.47±0.28% ID/g, 4h: 0.14±0.09% ID/g) and clearing from the bloodstream was via the renal and hepatobiliary routes (urine: 167±84% ID/g at 1 h, 10.3±4.8% ID/g at 4 h; gall bladder: 95±33% ID/g at 1 h, 63±11% ID/g at 4 h).

Conclusion: Copper-free, strain-promoted click chemistry is an attractive, straightforward approach to radiolabeling. Although the [(18)F]FBA-C(6)-ADBIO-based prosthetic group did not interfere with α(v)β(6)-targeted binding in vitro, it did influence the pharmacokinetics, possibly due to its size and lipophilic nature.