A Bone-Seeking trans-Cyclooctene for Pretargeting and Bioorthogonal Chemistry: A Proof of Concept Study Using 99mTc- and 177Lu-Labeled Tetrazines

A high yield synthesis of a novel, small molecule, bisphosphonate-modified trans -cyclooctene (TCO-BP, 2 ) that binds to regions of active bone metabolism and captures functionalized tetrazines in vivo, via the bioorthogonal inverse electron demand Diels–Alder (IEDDA) cycloaddition, was developed. A 99m Tc-labeled derivative of 2 demonstrated selective localization to shoulder and knee joints in a biodistribution study in normal mice. Compound 2 reacted rapidly with a 177 Lu-labeled tetrazine in vitro, and pretargeting experiments in mice, using 2 and the 177 Lu-labeled tetrazine, yielded high activity concentrations in shoulder and knee joints, with minimal uptake in other tissues. Pretargeting experiments with 2 and a novel 99m Tc-labeled tetrazine also produced high activity concentrations in the knees and shoulders. Critically, both radiolabeled tetrazines showed negligible uptake in the skeleton and joints when administered in the absence of 2 . Compound 2 can be utilized to target functionalized tetrazines to bone and represents a convenient reagent to test novel tetrazines for use with in vivo bioorthogonal pretargeting strategies.