A bioorthogonal strategy for the radioiodination of antibodies

Objectives To develop a methodology employing bioorthogonal click chemistry for the radioiodination of antibodies for PET imaging and radioimmunotherapy

Methods A modular approach to the synthesis of radioiodinated antibodies was developed based on the rapid and bioorthogonal inverse electron demand Diels-Alder cycloaddition. A tyrosine-modified tetrazine precursor (Tz-Tyr) was synthesized via standard organic chemistry techniques, characterized via UV-Vis, ¹H-NMR, ¹³C-NMR, ESI-MS, and LC-MS, and radiolabeled with ¹³¹I. As a model system, the HER2-targeting antibody trastuzumab (Tras) was modified with trans-cyclooctenes (TCO) via the conjugation of TCO-NHS to the lysines of the antibody.

Results The radioiodinated precursor ¹³¹I-Tz-Tyr was synthesized using chloramine-T and purified via HPLC in high purity and specific activity. The antibody trastuzumab was conjugated with TCO-NHS under basic conditions and purified to yield a Tras-TCO immunoconjugate with 2.0 ± 0.1 TCO/mAb. ¹³¹I-Tz-Tyr was incubated with Tras-TCO in PBS (pH 7.4) for 2 hours at room temperature, and the resulting radioimmunoconjugate was purified to give ¹³¹I-Tras with a specific activity of 1-2 mCi/mg. In vivo experiments in athymic nude mice bearing BT474 human breast cancer xenografts are forthcoming.

Conclusions A bioorthogonal strategy for labeling antibodies with the positron-emitting and therapeutic radioisotopes ¹²⁴I and ¹³¹I has been developed and effectively applied to the synthesis of a radioiodinated trastuzumab immunoconjugate in high specific activity and purity.