Abstract 4771: PET imaging of tissue factor using 64Cu-labeled active site-inhibited factor VII: A potential companion diagnostic for tissue factor targeted cancer therapies
Authors: Kristensen Lotte K. , Nielsen Carsten H. , Jeppesen Troels E. , Jensen Mette M. , Madsen Jacob , Wiinberg Bo , Petersen Lars C. , Kjaer Andreas 
Published: July 2016

Abstract
Introduction: The transmembrane glycoprotein tissue factor (TF) is the primary initiator of coagulation. In addition to its physiological role, TF is associated with a variety of pathophysiological processes including tumor growth, tumor angiogenesis and metastasis. Increased TF expression has been reported in 53-89% of all pancreatic adenocarcinomas and clinically correlates with advanced stage and poor survival. Non-invasive imaging of tumor TF status holds great clinical potential as a companion diagnostic for anti-cancer therapy targeting TF. The goal of our study was to develop and evaluate a PET tracer for imaging of TF expression in pancreatic cancer by utilizing the natural ligand for TF, coagulation factor VII.
Experimental procedures: Active site-inhibited factor VII (FVIIai) was conjugated to 2-S-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p-SCN-Bn-NOTA) followed by radiolabeling with 64Cu (64Cu-NOTA-FVIIai). Mice bearing subcutaneous pancreatic adenocarcinoma tumors (BxPC-3) were subjected to PET imaging 1, 4, 15 and 36 hours post-injection of 64Cu-NOTA-FVIIai for longitudinal assessment. The in vivo specificity of 64Cu-NOTA-FVIIai towards TF was evaluated in a competition blocking experiment and in a panel of subcutaneous pancreatic tumor models exhibiting low, medium and high TF expression. Additionally, the ability of 64Cu-NOTA-FVIIai to visualize TF expression in orthotopically implanted BxPC-3 tumors was evaluated by PET/MRI. Ex vivo biodistribution, immunohistochemistry and flow cytometry was performed to verify the in vivo imaging data.
Results: PET imaging at 1, 4, 15 and 36 hours after injection of 64Cu-NOTA-FVIIai revealed a tumor uptake of 2.3 ± 0.2, 3.7 ± 0.3, 3.4 ± 0.3 and 2.4 ± 0.3% injected dose per gram (%ID/g), respectively. A graduate increase in image contrast (tumor to background ratio) was observed over the imaging time course with no further increase beyond 15 hours. Competition with excess unlabeled FVIIai significantly reduced tumor uptake of 64Cu-NOTA-FVIIai (p&lt; 0.001) in subcutaneous BxPC-3 tumors, while the uptake was unaffected in other organs. The ex vivo biodistribution confirmed the data obtained by in vivo PET imaging. In pancreatic cancer models with different levels of TF expression, the tumor uptake was found to be significantly different from each other (p &lt; 0.001). This was in agreement with the TF level evaluated by immunohistochemistry and flow cytometry. Furthermore orthotopic BxPC-3 tumors were visualized and distinguishable by PET/MRI using 64Cu-NOTA-FVIIai as the PET tracer.
Conclusions: 64Cu-NOTA-FVIIai is well suited for specific PET imaging of TF in pancreatic cancer and its uptake is related to the tumor TF expression level. The data supports further development of 64Cu-NOTA-FVIIai as a companion diagnostic and theranostic agent.
Citation Format: Lotte K. Kristensen, Carsten H. Nielsen, Troels E. Jeppesen, Mette M. Jensen, Jacob Madsen, Bo Wiinberg, Lars C. Petersen, Andreas Kjaer. PET imaging of tissue factor using 64Cu-labeled active site-inhibited factor VII: A potential companion diagnostic for tissue factor targeted cancer therapies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4771.