Preclinical evaluation of zirconium-89 labeled anti-Trop2 antibody-drug conjugate (Trodelvy) for imaging in gastric cancer and triple-negative breast cancer.

[PURPOSE] Trophoblast cell-surface antigen 2 (Trop2) is overexpressed in various solid tumors and contributes to tumor progression, while its expression remains low in normal tissues. Trop2-targeting antibody-drug conjugate (ADC), sacituzumab govitecan-hziy (Trodelvy), has shown efficacy in targeting this antigen. Leveraging the enhanced specificity of ADCs, we conducted the first immunoPET imaging study of Trop2 expression in gastric cancer (GC) and triple-negative breast cancer (TNBC) models using Zr-labeled Trodelvy ([Zr]Zr-DFO-Trodelvy). This approach enables preclinical screening to identify patients who may benefit from targeted therapies.

[MATERIALS AND METHODS] Trop2 expression levels in GC and TNBC cell lines (NCI-N87, HGC-27, MDST8, and MDA-MB-468) were assessed via flow cytometry and immunofluorescence staining. Labeling of DFO-Trodelvy with Zr was performed in NaCO buffer at pH 7 (37 °C, 1.5 h). In vitro stability was analyzed using radio-thin layer chromatography. Biological properties were evaluated through cell uptake, saturation binding assays, and biodistribution experiments. ImmunoPET imaging with [Zr]Zr-DFO-Trodelvy was performed at various time points to confirm its in vivo targeting. Immunohistochemical and immunofluorescence analyses were conducted on tumor tissues from tumor-bearing mice.

[RESULTS] The radiochemical yield of [Zr]Zr-DFO-Trodelvy exceeded 90%, with a radiochemical purity (RCP) greater than 99%. Trop2 expression was high in MDA-MB-468 and NCI-N87 cells, while it was low in MDST8 and HGC-27 cells. The KD values of [Zr]Zr-DFO-Trodelvy were 9.44 nM for MDA-MB-468 and 3.51 nM for NCI-N87 cells. ImmunoPET imaging with [Zr]Zr-DFO-Trodelvy provided clear visualization of tumor morphology in MDA-MB-468 and NCI-N87 models (n = 3) as early as 6 h post-injection. Tumor uptake of [Zr]Zr-DFO-Trodelvy increased over time, peaking at 48 h (MDA-MB-468: 10.03 ± 1.26%ID/g; NCI-N87: 14.30 ± 2.09%ID/g), and was significantly higher than in the MDST8 (5.27 ± 0.71%ID/g) and HGC-27 (4.37 ± 0.54%ID/g) models. Co-injection with 2 mg of unlabeled Trodelvy significantly reduced uptake in NCI-N87 and MDA-MB-468 tumors (P < 0.001). A high target-to-non-target ratio was observed at 48 h, showing specific tumor uptake and minimal off-target accumulation. Fluorescence imaging further confirmed higher tumor uptake in the IRDye800CW-Trodelvy group compared to the IRDye800CW-Trodelvy-blocking group (P < 0.001).

[CONCLUSIONS] [Zr]Zr-DFO-Trodelvy for immunoPET imaging in TNBC and GC tumor models demonstrated specific, rapid, and sustained accumulation in tumors with high Trop2 expression, allowing for noninvasive monitoring of Trop2 status. The increased tumor-to-background ratio observed in immunoPET imaging suggests strong potential for clinical translation. Additionally, optical imaging, with its superior spatial resolution compared to PET, was employed to aid in precise probe localization and potentially enhancing differentiation between healthy and malignant tissues during surgical procedures.