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 99mTc-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 177Lu-labeled tetrazine
in vitro, and pretargeting experiments in mice, using 2 and the 177Lu-labeled tetrazine, yielded high activity
concentrations in shoulder and knee joints, with minimal uptake in
other tissues. Pretargeting experiments with 2 and a
novel 99mTc-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.

本研究成功合成了一种新型的小分子双磷酸酯修饰的环辛四烯（TCO-BP，2），该分子能够与活跃的骨代谢区域结合，并在体内通过生物正交逆电子需求Diels-Alder（IEDDA）环加成反应捕获功能化四嗪。2的99mTc标记衍生物在正常小鼠的生物分布研究中表现出对肩部和膝关节的选择性定位。体外实验中，化合物2与177Lu标记的四嗪迅速反应，使用2和177Lu标记的四嗪进行的小鼠预先靶向实验，在肩部和膝关节中产生了高活性浓度，而在其他组织中的摄取量最小。2与一种新型的99mTc标记四嗪的预先靶向实验同样在膝关节和肩部产生了高活性浓度。值得注意的是，当2不存在时，这两种放射性标记的四嗪在骨骼和关节中的摄取量均可忽略不计。化合物2可用于将功能化四嗪靶向至骨骼，并作为一种便捷的试剂来测试新型四嗪在体内生物正交预先靶向策略中的应用。