Characterization of a novel recombinant calcium-binding protein from Arca subcrenata and its anti-hepatoma activities in vitro and in vivo

Abstract

Previous studies demonstrated that ASP-3 was a novel calcium-binding protein from Arca subcrenata that effectively inhibited the proliferation of HepG2 cells. To further study the antitumor activity and mechanism of ASP-3, the cytotoxic effects of recombinant ASP-3 were evaluated in HepG2 cells. The results demonstrated that ASP-3 inhibited the proliferation of HepG2 cells by competitively binding to the EGF binding pocket of EGFR and inhibiting the JAK-STAT, RAS-RAF-MEK-ERK, and PI3K-Akt-mTOR signaling pathways mediated by EGFR. ASP-3 significantly inhibited tumor growth in a HepG2 cell subcutaneous xenograft nude mouse model, and its (25 mg/kg and 75 mg/kg) tumor inhibition rates were 46.92 % and 60.28 %, respectively. Furthermore, the crystal structure of ASP-3 was resolved at 1.4 Å. ASP-3 formed as a stable dimer and folded as an EF-Hand structure. ASP-3 stably bound to domain I and domain III of the EGFR extracellular region by using molecular docking and molecular dynamics simulation analysis. Compared with the endogenous ligand EGF, ASP-3 displayed a stronger interaction with EGFR. These experimental results indicated that recombinant ASP-3 possessed an effective anti-hepatoma effect. So, it might be a potential molecule for liver cancer therapy.