Dual-ligand-modified cantharidin nanoparticles for the treatment of hepatocellular carcinoma via the inhibition of Ephb4

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality, yet conventional chemotherapy is limited by poor tumor specificity and severe toxicity. Cantharidin (CTD), a natural antitumor agent, is hindered by nephrotoxicity and hepatotoxicity. This study aimed to construct a dual-ligand-modified CTD-loaded solid lipid nanoparticle (GA-FA-CSLNs) using 18-glycyrrhetinic acid (GA) for hepatocyte targeting and folate-PEG3500-DSPE (FA) for long circulation, and to evaluate its antitumor efficacy, mechanism, and safety. GA-FA-CSLNs were prepared by emulsification-ultrasonication and optimized via Box-Behnken design. The nanoparticles were characterized. In vitro antitumor activity was assessed in Huh-7 cells using CCK-8, flow cytometry, and Western blotting. In vivo efficacy and safety were evaluated in Huh-7 tumor-bearing nude mice, and pharmacokinetics in SD rats. GA-FA-CSLNs exhibited favorable physicochemical properties. In vitro studies showed enhanced cellular uptake, the lowest IC₅₀, and 43.2% Apoptosis in Huh-7 cells. Mechanistically, GA-FA-CSLNs downregulated Eph receptor B4(EphB4) and Bcl-2, while upregulating Caspase-3 and Bax. In vivo, the tumor inhibition rate reached 58.67%, superior to free CTD and sorafenib, with 100% 14-day survival and no significant abnormalities in serum biochemistry or histopathology. Pharmacokinetic analysis showed prolonged elimination half-life (2.54 h) and a 3.2-fold increase in area under the blood concentration-time curve versus unmodified CSLNs. GA-FA-CSLNs represent a promising actively targeted nanoplatform that enhances CTD antitumor efficacy against HCC via EphB4 pathway inhibition while reducing systemic toxicity, offering a potential strategy for targeted HCC therapy.