Tetrazine-induced activation of a trimethyl lock as a click-to-release system for protected doxorubicin

We herein report a novel chemically triggered click-to-release system, that combines the trimethyl lock (TML) lactonization with the bioorthogonal inverse electron demand Diels-Alder (IEDDA) reaction of a vinyl ether and a tetrazine. Kinetic studies were carried out on a vinyl phenol model system with six tetrazines using NMR and UV/Vis spectroscopy, revealing that within the three step sequence the IEDDA reaction was rate-limiting. The reaction rates were enhanced by increasing the electrophilicity of the tetrazine, while balancing reactivity and stability of the tetrazines. The anticancer drug doxorubicin was conjugated to a vinyl-modified TML. Its subsequent liberation from vinyl-TML was triggered by dimethyl 1,2,4,5-tetrazine-3,6-dicarboxylate and followed quantitatively by NMR, thereby providing a proof-of-concept for the tetrazine/TML click-to-release system. In addition the applicability of the reaction under physiolgoical conditions could be shown.