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Imaging is essential in accurately detecting, staging, and treating primary liver cancer (hepatocellular carcinoma [HCC]), one of the most prevalent and lethal malignancies. We developed a novel multifunctional nanoparticle (NP) specifically targeting glypican-3 (GPC3), a proteoglycan implicated in promotion of cell growth that is overexpressed in most HCCs. Quantitative real-time polymerase chain reaction was performed to confirm the differential GPC3 expression in two human HCC cells, Hep G2 (high) and HLF (negligible). These cells were treated with biotin-conjugated GPC3 monoclonal antibody (αGPC3) and subsequently targeted using superparamagnetic iron oxide NPs conjugated to streptavidin and Alexa Fluor 647. Flow cytometry demonstrated that only GPC3-expressing Hep G2 cells were specifically targeted using this αGPC3-NP conjugate (fourfold mean fluorescence over nontargeted NP), and magnetic resonance imaging (MRI) experiments showed similar findings (threefold R2 relaxivity). Confocal fluorescence microscopy localized the αGPC3 NPs only to the cell surface of GPC3-expressing Hep G2 cells. Further characterization of this construct demonstrated a negatively charged, monodisperse, 50 nm NP, ideally suited for tumor targeting. This GPC3-specific NP system, with dual-modality imaging capability, may enhance pretreatment MRI, enable refined intraoperative HCC visualization by near-infrared fluorescence, and be potentially used as a carrier for delivery of tumor-targeted therapies, improving patient outcomes

Glypican-3 Targeting of Liver Cancer Cells Using Multifunctional Nanoparticles