TY - JOUR
T1 - Lipophilicity and Click Reactivity Determine the Performance of Bioorthogonal Tetrazine Tools in Pretargeted In Vivo Chemistry
AU - Steen, E. Johanna L.
AU - Jorgensen, Jesper T.
AU - Denk, Christoph
AU - Battisti, Umberto M.
AU - Norregaard, Kamilla
AU - Edem, Patricia E.
AU - Bratteby, Klas
AU - Shalgunov, Vladimir
AU - Wilkovitsch, Martin
AU - Svatunek, Dennis
AU - Poulie, Christian B. M.
AU - Hvass, Lars
AU - Simon, Marina
AU - Wanek, Thomas
AU - Rossin, Raffaella
AU - Robillard, Marc
AU - Kristensen, Jesper L.
AU - Mikula, Hannes
AU - Kjaer, Andreas
AU - Herth, Matthias M.
PY - 2021
Y1 - 2021
N2 - The development of highly selective and fast biocompatible reactions for ligation and cleavage has paved the way for new diagnostic and therapeutic applications of pretargeted in vivo chemistry. The concept of bioorthogonal pretargeting has attracted considerable interest, in particular for the targeted delivery of radionuclides and drugs. In nuclear medicine, pretargeting can provide increased target-to-background ratios at early time-points compared to traditional approaches. This reduces the radiation burden to healthy tissue and, depending on the selected radionuclide, enables better imaging contrast or higher therapeutic efficiency. Moreover, bioorthogonally triggered cleavage of pretargeted antibody-drug conjugates represents an emerging strategy to achieve controlled release and locally increased drug concentrations. The toolbox of bioorthogonal reactions has significantly expanded in the past decade, with the tetrazine ligation being the fastest and one of the most versatile in vivo chemistries. Progress in the field, however, relies heavily on the development and evaluation of (radio)labeled compounds, preventing the use of compound libraries for systematic studies. The rational design of tetrazine probes and triggers has thus been impeded by the limited understanding of the impact of structural parameters on the in vivo ligation performance. In this work, we describe the development of a pretargeted blocking assay that allows for the investigation of the in vivo fate of a structurally diverse library of 45 unlabeled tetrazines and their capability to reach and react with pretargeted trans-cyclooctene (TCO)-modified antibodies in tumor-bearing mice. This study enabled us to assess the correlation of click reactivity and lipophilicity of tetrazines with their in vivo performance. In particular, high rate constants (>50 000 M-1 s(-1)) for the reaction with TCO and low calculated logD(7.4) values (below -3) of the tetrazine were identified as strong indicators for successful pretargeting. Radiolabeling gave access to a set of selected F-18-labeled tetrazines, including highly reactive scaffolds, which were used in pretargeted PET imaging studies to confirm the results from the blocking study. These insights thus enable the rational design of tetrazine probes for in vivo application and will thereby assist the clinical translation of bioorthogonal pretargeting.
AB - The development of highly selective and fast biocompatible reactions for ligation and cleavage has paved the way for new diagnostic and therapeutic applications of pretargeted in vivo chemistry. The concept of bioorthogonal pretargeting has attracted considerable interest, in particular for the targeted delivery of radionuclides and drugs. In nuclear medicine, pretargeting can provide increased target-to-background ratios at early time-points compared to traditional approaches. This reduces the radiation burden to healthy tissue and, depending on the selected radionuclide, enables better imaging contrast or higher therapeutic efficiency. Moreover, bioorthogonally triggered cleavage of pretargeted antibody-drug conjugates represents an emerging strategy to achieve controlled release and locally increased drug concentrations. The toolbox of bioorthogonal reactions has significantly expanded in the past decade, with the tetrazine ligation being the fastest and one of the most versatile in vivo chemistries. Progress in the field, however, relies heavily on the development and evaluation of (radio)labeled compounds, preventing the use of compound libraries for systematic studies. The rational design of tetrazine probes and triggers has thus been impeded by the limited understanding of the impact of structural parameters on the in vivo ligation performance. In this work, we describe the development of a pretargeted blocking assay that allows for the investigation of the in vivo fate of a structurally diverse library of 45 unlabeled tetrazines and their capability to reach and react with pretargeted trans-cyclooctene (TCO)-modified antibodies in tumor-bearing mice. This study enabled us to assess the correlation of click reactivity and lipophilicity of tetrazines with their in vivo performance. In particular, high rate constants (>50 000 M-1 s(-1)) for the reaction with TCO and low calculated logD(7.4) values (below -3) of the tetrazine were identified as strong indicators for successful pretargeting. Radiolabeling gave access to a set of selected F-18-labeled tetrazines, including highly reactive scaffolds, which were used in pretargeted PET imaging studies to confirm the results from the blocking study. These insights thus enable the rational design of tetrazine probes for in vivo application and will thereby assist the clinical translation of bioorthogonal pretargeting.
KW - bioorthogonal chemistry
KW - tetrazine ligation
KW - pretargeted imaging
KW - PET
KW - fluorine-18
KW - molecular imaging
KW - TRIGGERED DRUG-RELEASE
KW - DIELS-ALDER REACTION
KW - TRANS-CYCLOOCTENE
KW - C-11-LABELED TETRAZINE
KW - IMPROVED STABILITY
KW - BIODISTRIBUTION
KW - RADIOSYNTHESIS
KW - MODULATION
KW - ANTIBODIES
U2 - 10.1021/acsptsci.1c00007
DO - 10.1021/acsptsci.1c00007
M3 - Journal article
C2 - 33860205
VL - 4
SP - 824
EP - 833
JO - ACS Pharmacology and Translational Science
JF - ACS Pharmacology and Translational Science
SN - 2575-9108
IS - 2
ER -