Site-specifically labeled CA19.9-targeted immunoconjugates for the PET, NIRF, and multimodal PET/NIRF imaging of pancreatic cancer

Abstract

Molecular imaging agents for preoperative positron emission tomography (PET) and near-infrared fluorescent (NIRF)-guided delineation of surgical margins could greatly enhance the diagnosis, staging, and resection of pancreatic cancer. PET and NIRF optical imaging offer complementary clinical applications, enabling the noninvasive whole-body imaging to localize disease and identification of tumor margins during surgery, respectively. We report the development of PET, NIRF, and dual-modal (PET/NIRF) imaging agents, using 5B1, a fully human monoclonal antibody that targets CA19.9, a well-established pancreatic cancer biomarker. Desferrioxamine (DFO) and/or a NIRF dye (FL) were conjugated to the heavy-chain glycans of 5B1, using a robust and reproducible site-specific (ss) labeling methodology to generate three constructs ( ss DFO-5B1, ss FL-5B1, and ss dual-5B1) in which the immunoreactivity was not affected by the conjugation of either label. Each construct was evaluated in a s.c. xenograft model, using CA19.9-positive (BxPC3) and -negative (MIAPaCa-2) human pancreatic cancer cell lines. Each construct showed exceptional uptake and contrast in antigen-positive tumors with negligible nonspecific uptake in antigen-negative tumors. Additionally, the dual-modal construct was evaluated in an orthotopic murine pancreatic cancer model, using the human pancreatic cancer cell line, Suit-2. The ss dual-5B1 demonstrated a remarkable capacity to delineate metastases and to map the sentinel lymph nodes via tandem PET-computed tomography (PET/CT) and NIRF imaging. Fluorescence microscopy, histopathology, and autoradiography were performed on representative sections of excised tumors to visualize the distribution of the constructs within the tumors. These imaging tools have tremendous potential for further preclinical research and for clinical translation.pancreatic cancer | CA19.9 | molecular imaging | PET imaging P ancreatic ductal adenocarcinoma (PDAC) is currently the fourth leading cause of cancer mortality and is expected to surpass both colorectal and breast cancer in total annual deaths by 2030 (1, 2). Surgical resection of the pancreas is the only curative treatment, but the presence of metastases precludes over 80% of patients from resection ab initio (3). The overall 5-y survival rate is ∼5%, and for those who qualify for surgical resection, the 5-y survival rate is only 25% due to the high incidence of undiscovered metastases (4, 5). Further complicating this dire situation, patients with PDAC are regularly misdiagnosed or understaged, confounding treatment strategies and preventing proper enrollment in clinical trials. Many of these problems could be avoided and outcomes improved if adequate clinical tools for diagnosing, staging, and treating PDAC were available.Positron emission tomography (PET) is a promising technological platform for detecting, staging, and monitoring the progression or regression of many solid tumors, including PDAC. Optical imaging is a complementary platform that makes possible the accurate ...

Establishment of the In Vivo Efficacy of Pretargeted Radioimmunotherapy Utilizing Inverse Electron Demand Diels-Alder Click Chemistry

“…1C) and a TCO-modified immunoconjugate of 5B1, a fully human antibody that targets the PDAC biomarker carbohydrate antigen 19.9 (CA19.9). (23, 24, 29–31) We demonstrate that this approach elicits a dose-dependent therapeutic response in mice bearing CA19.9-expressing BxPC3 human PDAC xenografts. Moreover, this strategy has proven effective while delivering only a small fraction of the radiation dose to healthy tissues delivered by traditional RIT with a directly-labeled 177 Lu-5B1 construct.…”

“…All in vivo studies were carried out in female, athymic nude mice [Crl:NU(NCr)-Foxn1 nu , 6–8 weeks) bearing subcutaneous BxPC3 xenografts that were inoculated on the right flank, as previously described. (29, 31) BxPC3 cells obtained from ATCC (Manassas, VA) were obtained in November 2014, validated via STR analysis, and used within 4 weeks of resuscitation. BxPC3 cells were grown in RPMI medium modified to contain 4.5 g/L glucose, 1.5g/L sodium bicarbonate and supplemented with 10% (vol/vol) heat-inactivated fetal calf serum, 100 IU penicillin, 100 µg/mL streptomycin, 10mM HEPES and 10 cc/L non-essential amino acids.…”

Investigation into metastatic processes and the therapeutic effects of gemcitabine on human pancreatic cancer using an orthotopic SUIT‑2 pancreatic cancer mouse model

“…The SUIT-2 orthotopic pancreatic cancer mouse model has been used in other studies for evaluating promising new anticancer agents (14,(17)(18)(19)(20) evaluated the tumor reduction effects by measuring tumor weight in a mouse model. SUIT-2 is reported not only in an orthotopic but also in a subcutaneous model, a peritoneal dissemination model, and a lung metastasis model.…”

Pretargeted PET Imaging Using a Site-Specifically Labeled Immunoconjugate

“…In three different murine model systems, we have illustrated that this approach produces well-defined and homogeneous immunoconjugates with in vivo behavior comparable—and often superior—to immunoconjugates synthesized using traditional, non-site-specific methods. 26,28,29 …”