A One-Pot Three-Component Double-Click Method for Synthesis of [67Cu]-Labeled Biomolecular Radiotherapeutics

Abstract

A one-pot three-component double-click process for preparing tumor-targeting agents for cancer radiotherapy is described here. By utilizing DOTA (or NOTA) containing tetrazines and the TCO-substituted aldehyde, the two click reactions, the tetrazine ligation (an inverse electron-demand Diels-Alder cycloaddition) and the RIKEN click (a rapid 6π-azaelectrocyclization), could simultaneously proceed under mild conditions to afford covalent attachment of the metal chelator DOTA or NOTA to biomolecules such as to albumin and anti-IGSF4 antibody without altering their activities. Subsequently, radiolabeling of DOTA- or NOTA-attached albumin and anti-IGSF4 antibody (an anti-tumor-targeting antibody) with [67Cu], a β−-emitting radionuclide, could be achieved in a highly efficient manner via a simple chelation with DOTA proving to be a more superior chelator than NOTA. Our work provides a new and operationally simple method for introducing the [67Cu] isotope even in large quantities to biomolecules, thereby representing an important process for preparations of clinically relevant tumor-targeting agents for radiotherapy.

Evaluation of a 68Ga-Labeled DOTA-Tetrazine as a PET Alternative to 111In-SPECT Pretargeted Imaging

“…Apart from this, great effort has been put into the development of additional radiolabeled tetrazines as potential pretargeting imaging agents. However, many reports of these new radiolabeled tetrazines lack in vivo evaluation of their use as pretargeted imaging agents, likely due to the lack of alternative pretargeting screening models to mAbs [29][30][31][32][33][34]. Although TCO-CC49 mAb serves as a positive control for pretargeting experiments because of its extensive optimization and evaluation, antibodies as a screening model is challenged by the need of meticulous pretargeting procedures, disease models to test them, and the fact that they and their TCO modification can be expensive to produce.…”

Multifunctional Clickable Reagents for Rapid Bioorthogonal Astatination and Radio‐Crosslinking

“…In particular, tetrazine ligations – the fastest bioorthogonal reactions described so far – have had a significant impact by enabling strategies for (i) rapid radiolabeling using prosthetic groups, (ii) site‐specific radiolabeling, and (iii) pretargeting approaches based on in vivo chemistry . To this end, various radiolabeled 1,2,4,5‐tetrazines (Tz) have been developed as clickable tools for PET and SPECT imaging ( 18 F, 11 C, 111 In, 64 Cu, 123 I, 125 I), and targeted radiotherapy ( 177 Lu, 67 Cu) making use of the fast bioorthogonal ligation with trans ‐cyclooctenes (TCO).…”

Harnessing the Coordination Chemistry of 1,4,7‐Triazacyclononane for Biomimicry and Radiopharmaceutical Applications

“…Examples of these include ligands 55 , 56 and 60 for rapid conjugation to specific peptides by Cu I ‐catalysed click reaction, and strained cyclooctynes 57 – 59 for copper‐free attachment to azide‐functionalised peptides, proteins and nanoparticles. Also, tetrazine‐bearing NOTA derivatives 61 and 62 have been used for linking trans ‐cyclooctene‐functionalised antibodies using inverse‐electron‐demand Diels–Alder reactions …”