Abstract
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Objectives: Pancreatic cancer is one type of highly lethal diseases. While there is not a standard test for diagnosis, pancreatic cancer is generally found in the advanced stage with an extremely low survival rate. The trophoblast cell surface antigen 2 (Trop2), a transmembrane glycoprotein that is associated with cell proliferation and is overexpressed in most of the solid epithelial cancers has shown great potential as an effective marker for diagnosis. Thus, we aim to evaluate the expression level of Trop2 among various pancreatic cancer cell lines and employ the isotope-labeled anti-Trop2 antibody as PET tracer for detecting the Trop2-positive tumors in both subcutaneous and orthotopic models of pancreatic cancer.
Methods: The anti-Trop2 antibody (AF650, Research and Diagnostic Systems, Inc.) was conjugated with desferrioxamine (DFO) for radiolabeling of 89Zr (half-life 78.41 h). The relative expression level of Trop2 among BxPC-3, MIA PaCa and AsPC-1 pancreatic cancer cell lines was investigated via western blot and flow cytometry. And the competitive binding assay of AF650 was performed in vitro (BxPC-3 cell) as well. To evaluate the targeting capacity and specificity of the PET tracer, 89Zr-DFO-AF650 has been employed for the PET imaging and bio-distribution studies in subcutaneous models of pancreatic cancer. The expression of Trop2 was evaluated via the immunofluorescence staining of tumor tissue sections as well. Moreover, the targeting efficiency of PET-tracer was further examined in a Trop2 positive orthotopic cancer model.
Results: The Trop2 expression has been found in BxPC-3 cell, and the MIA PaCa and AsPC-1 rarely expressed the Trop2. The competitive binding assay indicated the Kd and Bmax values of AF650 are 22.34 ± 2.509 and 0.1163 ± 0.005 respectively. According to the PET imaging, around 19.3 ± 2.1 % tracer (89Zr-DFO-AF650) had been uptaken in BxPC-3 tumors, while the values of MIA PaCa and AsPC-1 were about 4.7 ± 1.0 and 2.6 ± 0.4 %ID/g at 120 h post-injection in subcutaneous cancer models (n=4), respectively. Correspondingly, the bio-distribution showed significantly higher accumulation of tracer in BxPC-3 (28.8 ± 7.6 %ID/g) tumor than that in MIA PaCa (6.8 ± 2.0 %ID/g) and AsPC-1 (3.5 ± 0.5 %ID/g) tumors (n=4). In the BxPC-3 tumor model, the accumulation of non-specific IgG in the tumor was about 6.9 ± 1.3 and 8.7 ± 4.9 %ID/g via PET-imaging and ex-vivo assays (n=4) respectively, which was much lower than the values of the tracer. Furthermore, the tracer could effectively identify the BxPC-3 (Trop2 positive) tumor in the othotopic model according to the results of PET-imaging (18.1 ± 3.9 %ID/g) and bio-distribution (26.0 ± 6.3 %ID/g) (n=4). Conclusion: This work demonstrated the differentiated expression of Trop2 among three pancreatic cancer cell lines. The 89Zr-DFO-AF650 could be employed as a specific tracer for imaging Trop2 positive pancreatic cancer. Overall, this targeted antibody holds great promise for PET imaging of pancreatic cancer.
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