Multimodal Targeted Nanoparticle-Based Delivery System for Pancreatic Tumor Imaging in Cellular and Animal Models
Authors: Medina Oula Penate , Tower Robert J. , Medina Tuula Penate , Ashkenani Fatma , Appold Lia , Bötcher Marcus , Huber Lukas , Will Olga , Ling Qi , Hauser Charlotte , Rohwedder Arndt , Heneweer Carola , Peschke Eva , Hövener Jan-Bernd , Lüdtke-Buzug Kerstin , Boretius Susann , Mentlein Rolf , Kairemo Kalevi , Glüer Claus C. , Sebens Susanne , Kalthoff Holger 
Published: July 2020

Background:
Pancreatic ductal adenocarcinoma (PDAC), which ranks forth on the cancer-related death statistics
still is both a diagnostic and a therapeutic challenge. Adenocarcinoma of the exocrine human pancreas originates in most
instances from malignant transformation of ductal epithelial cells, alternatively by Acinar-Ductal Metaplasia (ADM). RA96 antibody targets to a mucin M1, according to the more recent nomenclature MUC5AC, an extracellular matrix component
excreted by PDAC cells. In this study, we tested the usability of multimodal nanoparticle carrying covalently coupled RA96 Fab fragments for pancreatic tumor imaging.

Methods:
In order to make and evaluate a novel, better targeting, theranostic nanoparticle, iron nanoparticles and the optical
dye indocyanin green (ICG) were encapsulated into the cationic sphingomyelin (SM) consisting liposomes. RA-96 Fab
fragment was conjugated to the liposomal surface of the nanoparticle to increase tumor homing ability. ICG and iron
nanoparticle-encapsulated liposomes were studied in vitro with cells and (i) their visibility in magnetic resonance imaging
(MRI), (ii) optical, (iii) Magnetic particle spectroscopy (MPS) and (iv) photoacoustic settings was tested in vitro and also
in in vivo models. The targeting ability and MRI and photoacoustic visibility of the RA-96-nanoparticles were first tested
in vitro cell models where cell binding and internalization was studied. In in vivo experiments liposomal nanoparticles were injected into a tail vain using an orthotopic pancreatic tumor xenograft model and subcutaneous pancreas cancer cell
xenografts bearing mice to determine in vivo targeting abilities of RA-96-conjugated liposomes.

Results:
Multimodal liposomes could be detected by MRI, MPS and by photoacoustic imaging in addition to optical imaging
showing a wide range of imaging utility. The fluorescent imaging of ICG in pancreatic tumor cells Panc89 and Capan-2
revealed increased association of ICG-encapsulated liposomes carrying RA-96 Fab fragments in vitro compared to the
control liposomes without covalently linked RA-96. Fluorescent molecular tomography (FMT) studies showed increased
accumulation of the RA96-targeted nanoparticles in the tumor area compared to non-targeted controls in vivo. Similar
accumulation in the tumor sites could be seen with liposomal ferric particles in MRI. Fluorescent tumor signal was
confirmed by using an intraoperative fluorescent imaging system which showed fluorescent labeling of pancreatic tumors.

Conclusion:

These results suggest that RA-96-targeted liposomes encapsulating ICG and iron nanoparticles can be used to
image pancreatic tumors with a variety of optical and magnetic imaging techniques. Additionally, they might be a suitable
drug delivery tool to improve treatment of PDAC patients.