Abstract 715: Molecular imaging of hepatocellular carcinoma using radiolabeled monoclonal antibody PET probes targeting glypican-3.
Authors: Yang Xiaoyang , Hoppmann Susan , Liu Hongguang , Allegretta Mark , Chua Mei-Sze , Cheng Zhen , So Samuel 
Published: August 2014

Abstract
Background: Glypican-3 (GPC3) is a cell-surface protein which is over-expressed in more than 50% of hepatocellular carcinoma (HCC) patients, but is negligibly expressed in normal livers, benign liver lesions, and tissues adjacent to HCC. GPC3 is therefore a potential molecular target for HCC diagnosis.
Methods: Two radiotracers, 64Cu-DOTA-GPC3 and 89Zr-DFO-GPC3, were synthesized based on the human anti-GPC3 monoclonal antibody (MAb clone 1G12). These radiotracers were used for positron emission tomography (PET) imaging of subcutaneous or orthotopic HCC xenografts in nude mice, generated from HCC cell lines expressing high (HepG2), moderate (Hep3B), or low (PLC/PRF/5) levels of GPC3. Biodistribution analysis and tumor-to-liver ratios were calculated to assess the performance and specificity of these radiotracers.
Results: The anti-GPC3 MAb showed high binding affinity to recombinant human GPC3 protein in vitro (KD = 0.40 ± 0.04 nM), and specifically identified HCC cell lines and human specimens with high GPC3 expression. In vivo, 64Cu-DOTA-GPC3 showed significantly higher uptake in HepG2 xenografts (17.05 ± 1.31% ID/g) compared with non-specific 18F-fluoro-deoxy-glucose (18F-FDG) (3.17 ± 0.59% ID/g), and the non-targeting 64Cu-DOTA-IgG (5.10 ± 1.27% ID/g) at 72 h post-injection (p.i.) (p &lt; 0.005). However, the tumor-to-liver ratio was only 0.46 ± 0.32 at 72 h p.i., due to high uptake of 64Cu-DOTA-GPC3 in the liver. To increase tumor-to-liver ratio, we synthesized the 89Zr-DFO-GPC3 radiotracer, which displayed specific cellular uptake in GPC3-positive cells in vitro, and also internalized over time in these cells. In vivo, 89Zr-DFO-GPC3 showed high and specific uptake in the subcutaneous HepG2 xenografts, but not in the non-HCC, GPC3-negative ones (PC3 and A375M). Consistently, in vivo biodistribution analysis demonstrated significantly higher uptake of 89Zr-DFO-GPC3 in HepG2 xenografts at 48 h p.i. (10.80 ± 1.69% ID/g, n=4) and 192 h p.i. (8.76 ± 1.12% ID/g, n=4), compared to 1.19 ± 0.27% ID/g in PC3 xenografts and 2.02 ± 0.58% ID/g in A375M xenografts at 192 h time point (p &lt; 0.005) The tumor-to-liver ratios for HepG2 xenografts at 48 h and 192 h were 2.64 ± 0.20, and 4.10 ± 0.17, respectively. More importantly, 89Zr-DFO-GPC3 was able to delineate orthotopic HCC xenografts from the surrounding normal liver. Biodistribution analysis in these orthotopic xenografts showed uptake of 14.0 ± 2.50% ID/g for HepG2; 7.71 ± 1.64% ID/g for Hep3B; and 5.54 ± 1.00% ID/g for PLC/PRF/5 at 168 h p.i. The corresponding tumor-to-liver ratios were 6.65 ± 1.33 for HepG2, 6.15 ± 1.75 for PLC/PRF/5, and 4.29 ± 0.52 for Hep3B.
Conclusion: We demonstrate that anti-GPC3 MAb-based PET probes can detect GPC3-positive HCCs in subcutaneous and orthotopic animal models. Especially, 89Zr-DFO-GPC3 is a promising probe and clinically useful probe for the specific imaging of GPC3-positive HCCs.
Citation Format: Xiaoyang Yang, Susan Hoppmann, Hongguang Liu, Mark Allegretta, Mei-Sze Chua, Zhen Cheng, Samuel So. Molecular imaging of hepatocellular carcinoma using radiolabeled monoclonal antibody PET probes targeting glypican-3. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 715. doi:10.1158/1538-7445.AM2013-715