Preclinical Evaluation of a Novel 18F-Labeled dTCO-Amide Derivative for Bioorthogonal Pretargeted Positron Emission Tomography Imaging

Abstract

Pretargeted positron emission tomography (PET) imaging based on the bioorthogonal inverse-electron-demand Diels-Alder reaction between tetrazines (Tz) and trans-cyclooctenes (TCO) has emerged as a promising tool for solid tumor imaging, allowing the use of short-lived radionuclides in immune-PET applications. With this strategy, it became possible to achieve desirable target-to-background ratios and at the same time to decrease the radiation burden to nontargeted tissues because of the fast clearance of small PET probes. Here, we show the synthesis of novel ¹⁸F-labeled dTCO-amide probes for pretargeted immuno-PET imaging. The PET probes were evaluated regarding their stability, reactivity toward tetrazine, and pharmacokinetic profile. [ ¹⁸ F]MICA-213 showed an extremely fast kinetic rate (10,553 M^-1 s^-1 in 50:50 MeOH/water), good stability in saline and plasma up to 4 h at 37 °C with no isomerization observed, and the biodistribution in healthy mice revealed a mixed hepatobiliary and renal clearance with no defluorination and low background in other tissues. [ ¹⁸ F]MICA-213 was further used for in vivo pretargeted immune-PET imaging carried out in nude mice bearing LS174T colorectal tumors that were previously treated with a tetrazine-modified anti-TAG-72 monoclonal antibody (CC49). Pretargeted μPET imaging results showed clear visualization of the tumor tissue with a significantly higher uptake when compared to the control.

Figure 1

Chemical structures of the new dTCO amine scaffold and its ¹⁸F-labeled derivative PET probes.

Scheme 1. Synthesis of dTCO-amide Derivatives

Scheme 2. Radiosynthesis of [¹⁸F]MICA-212 and [¹⁸F]MICA-213 by ¹⁸F-Labeling

Scheme 3. Example of the Kinetic Reaction between MICA-213 Cold Reference (12) and Methyl-Tetrazine-NHS (Scheme S2)

Figure 2

Biodistribution of [¹⁸F]MICA-212 and [¹⁸F]MICA-213 in healthy BALB/c mice [mean ± standard deviation (SD), n = 3].

Figure 3

Time-activity curves from 0 to 60 min after iv administration of [¹⁸F]MICA-213 (mean ± SD, n = 4) in healthy BALB/c mice.

Figure 4

(A) Procedure followed for the in vivo pretargeted PET imaging study based on the bioorthogonal chemistry. (B) In vivo representative μPET/CT image of the LS174T tumor-bearing mice injected with [¹⁸F]MICA-213 24 h after injection of CC49 or CC49-Tz. Static images were acquired 60 min post tracer injection. The white dashed line encircles the tumor region. (C) Ex vivo biodistribution of LS174T tumor-bearing mice 60 min post tracer injection (mean ± SD, n = 5/group, p < 0.054).