Highly efficient click labeling using 2-[¹⁸F]fluoroethyl azide and synthesis of an ¹⁸FN-hydroxysuccinimide ester as conjugation agent

Introduction: Click labeling using 2-[¹⁸F]fluoroethyl azide has been proven to be promising methods of radiolabeling small molecules and peptides, some of which are undergoing clinical evaluations. However, the previously reported method afforded low yield, poor purities and under desirable reproducibility.

Methods: A vacuum distillation method was used to isolate 2-[¹⁸F]fluoroethyl azide, and the solvent effect of acetonitrile and dimethylformamide (DMF) on the click labeling using Cu(I) from copper sulfate/sodium ascorbate was studied. The labeling conditions were optimized to radiosynthesize a hydroxysuccinimide ester (N-hydroxysuccinimide, or NHS).

Results: 2-[¹⁸F]fluoroethyl azide was isolated by the vacuum distillation method with >80% yield within 10min in a "pure" and click-ready form. It was found that the amount of DMF was critical for maintaining high levels of Cu(I) from copper sulfate/sodium ascorbate in order to rapidly complete the click labeling reaction. The addition of bathophenanthrolinedisulfonic acid disodium salt to the mixture of copper sulfate/sodium ascorbate also greatly improved the click labeling efficiency. Through exploiting these optimizations, a base-labile NHS ester was rapidly radiosynthesized in 90% isolated yield with good chemical and radiochemical purities.

Conclusions: We have developed a general method to click-label small molecules efficiently using [¹⁸F]2 for research and clinical use. This NHS ester can be used for conjugation chemistry to label antibodies, peptides and small molecules as positron emission tomography tracers.