Cadherin-17 as a target for the immunoPET of adenocarcinoma

Abstract

Purpose: Cadherin-17 (CDH17) is a calcium-dependent cell adhesion protein that is overexpressed in several adenocarcinomas, including gastric, colorectal, and pancreatic adenocarcinoma. High levels of CDH17 have been linked to metastatic disease and poor prognoses in patients with these malignancies, fueling interest in the protein as a target for diagnostics and therapeutics. Herein, we report the synthesis, in vitro validation, and in vivo evaluation of a CDH17-targeted ⁸⁹Zr-labeled immunoPET probe.

Methods: The CDH17-targeting mAb D2101 was modified with an isothiocyanate-bearing derivative of desferrioxamine (DFO) to produce a chelator-bearing immunoconjugate - DFO-D2101 - and flow cytometry and surface plasmon resonance (SPR) were used to interrogate its antigen-binding properties. The immunoconjugate was then radiolabeled with zirconium-89 (t_1/2 ~ 3.3 days), and the serum stability and immunoreactive fraction of [⁸⁹Zr]Zr-DFO-D2101 were determined. Finally, [⁸⁹Zr]Zr-DFO-D2101's performance was evaluated in a trio of murine models of pancreatic ductal adenocarcinoma (PDAC): subcutaneous, orthotopic, and patient-derived xenografts (PDX). PET images were acquired over the course of 5 days, and terminal biodistribution data were collected after the final imaging time point.

Results: DFO-D2101 was produced with a degree of labeling of ~ 1.1 DFO/mAb. Flow cytometry with CDH17-expressing AsPC-1 cells demonstrated that the immunoconjugate binds to its target in a manner similar to its parent mAb, while SPR with recombinant CDH17 revealed that D2101 and DFO-D2101 exhibit nearly identical K_D values: 8.2 × 10^-9 and 6.7 × 10^-9 M, respectively. [⁸⁹Zr]Zr-DFO-D2101 was produced with a specific activity of 185 MBq/mg (5.0 mCi/mg), remained >80% stable in human serum over the course of 5 days, and boasted an immunoreactive fraction of >0.85. In all three murine models of PDAC, the radioimmunoconjugate yielded high contrast images, with high activity concentrations in tumor tissue and low uptake in non-target organs. Tumoral activity concentrations reached as high as >60 %ID/g in two of the cohorts bearing PDXs.

Conclusion: Taken together, these data underscore that [⁸⁹Zr]Zr-DFO-D2101 is a highly promising probe for the non-invasive visualization of CDH17 expression in PDAC. We contend that this radioimmunoconjugate could have a significant impact on the clinical management of patients with both PDAC and gastrointestinal adenocarcinoma, most likely as a theranostic imaging tool in support of CDH17-targeted therapies.

Keywords: Adenocarcinoma; Cadherin-17; Pancreatic ductal adenocarcinoma; Positron emission tomography; Radioimmunoconjugate; Zirconium-89.

Fig. 1

A Binding parameters of D2101 and DFO-D2101 for CDH17 as determined via surface plasmon resonance. B Flow cytometry data for D2101 and DFO-D2101 with CDH17-expressing AsPC-1 cells. C Longitudinal stability study of [⁸⁹Zr]Zr-DFO-D2101 in human serum. D Immunoreactivity assay for [⁸⁹Zr]Zr-DFO-D2101 performed using CDH17-coated magnetic beads

Fig. 2

A Representative coronal PET images obtained following the administration of [⁸⁹Zr]Zr-DFO-D2101 to athymic nude mice bearing subcutaneous AsPC-1 PDAC xenografts (T, tumor). B Ex vivo biodistribution data collected 120 h after the administration of [⁸⁹Zr]Zr-DFO-D2101. [⁸⁹Zr]Zr-DFO-IgG was used as a control. Statistical significance was determined via a two-tailed t test with a Welch’s correction using GraphPad Prism: *p < 0.05

Fig. 3

A Representative coronal PET images and a PET-CT maximum intensity projection obtained following the administration of [⁸⁹Zr]Zr-DFO-D2101 to athymic nude mice bearing orthotopic AsPC-1_luc PDAC xenografts (T, tumor; S, spleen; L, liver). B Ex vivo biodistribution data collected 120 h after the administration of [⁸⁹Zr]Zr-DFO-D2101. C CDH17 immunohistochemical staining (top) and autoradiographic (bottom) images of pancreas tissue excised 120 h after the administration of [⁸⁹Zr]Zr-DFO-D2101 (arrows denote tumor lesions). D Transverse PET-CT image acquired 120 h after the administration of [⁸⁹Zr]Zr-DFO-D2101 (T, tumor; same mouse as in panel A)

Fig. 4

A Confocal microscopy images of a trio of patient-derived PDAC tissues — 38a, 46a, and 56b — following immunofluorescence staining for CDH17. Blue = DAPI; Green = staining for CDH17 visualized via goat anti-human IgG Alexa-488. A CDH17-expressing AsPC-1 tumor was also frozen, cut, stained for CDH17, and imaged. B Representative coronal PET images and a PET-CT maximum intensity projection obtained following the administration of [⁸⁹Zr]Zr-DFO-D2101 to NSG mice bearing patient-derived PDAC tumors (T, tumor). C Ex vivo biodistribution data collected 120 h after the administration of [⁸⁹Zr]Zr-DFO-D2101