# Pharmacokinetic Optimization of Radiocopper-Based Theranostic Pretargeting

## TL;DR

This study develops and tests radioligands for cancer imaging and therapy, showing improved tumor targeting and safety in mice.

## Contribution

The novel contribution is the pharmacokinetic optimization of radiocopper-based pretargeting agents for theranostic applications.

## Key findings

- Three tetrazine radioligands were synthesized and radiolabeled with copper-64 for immunoPET.
- [64Cu]Cu-SarAr-PEG10-Tz provided the best tumor-to-background contrast in a mouse model.
- Pretargeted radioimmunotherapy with [67Cu]Cu-SarAr-PEG10-Tz showed promising efficacy and safety in colorectal cancer.

## Abstract

In vivo pretargeting offers a strategy to improve nuclear
imaging
and radiopharmaceutical therapy by increasing tumor-to-background
activity concentration ratios and decreasing radiation burden to healthy
tissues. One particularly promising approach to in vivo pretargeting
is predicated on the inverse electron-demand Diels–Alder (IEDDA)
ligation between tetrazine (Tz)-based radioligands and trans-cyclooctene (TCO)-bearing immunoconjugates. Not surprisingly, the
performance of such systems is highly dependent upon the pharmacokinetic
profiles of the small molecule radioligands. Herein, we report the
synthesis and characterization of a trio of sarcophagine-bearing tetrazinesSarAr-Tz,
SarAr-PEG5-Tz, and SarAr-PEG10-Tzas
well as their radiolabeling with copper-64 (64Cu, t
1/2 ∼ 12.7 h), a positron-emitting radioisotope
of copper. These radioligands were paired with a TCO-bearing variant
of the A33 antigen-targeting antibody huA33 (i.e., huA33-TCO) for
pretargeted immunoPET in a murine model of colorectal cancer, revealing
that all three produced images with excellent tumor-to-background
contrast, but [64Cu]­Cu-SarAr-PEG10-Tz yielded
the best tumor-to-tissue activity concentration ratios. In light of
its superior performance, SarAr-PEG10-Tz was subsequently
radiolabeled with copper-67 (67Cu, t
1/2 ∼ 61.8 h), a β–-emitting
radioisotope of copper, to produce [67Cu]­Cu-SarAr-PEG10-Tz. This radioligand was then paired with huA33-TCO for
in vivo biodistribution and longitudinal therapy studies, ultimately
revealing that pretargeted radioimmunotherapy with [67Cu]­Cu-SarAr-PEG10-Tz exhibits promising efficacy and safety in a murine model
of colorectal cancer.