Effect of sorafenib on des-γ-carboxyprothrombin secretion by a human hepatocellular carcinoma cell line

Patients with hepatocellular carcinoma (HCC) who respond to sorafenib have been reported to exhibit an increase in the level of des-γ-carboxyprothrombin (DCP) in the blood, subsequent to the initiation of sorafenib treatment. In the present study, the levels of secretion of DCP and DCP with more γ-carboxyglutamic residues (NX-DCP) and the effects of hypoxic conditions were examined in 13 liver cancer cell lines, and the presence of vitamin K and sorafenib, in the KYN-2 cell line, which resulted in confirmed DCP and NX-DCP secretion. DCP, NX-DCP and prothrombin secretion were confirmed in 2/13 cell lines, KYN-2 and KIM-1. The level of secretions increased under hypoxic conditions. The addition of vitamin K suppressed cell proliferation, and DCP expression decreased to below detectable levels, however the level of prothrombin expression increased. Sorafenib treatment increased the level of apoptosis and suppressed cell proliferation, and decreased DCP and NX-DCP. In contrast, levels of prothrombin and vascular endothelial growth factor (VEGF) expression exhibited a slight increase. When the same experiment was conducted under hypoxic conditions, DCP secretion significantly decreased in the presence of sorafenib. The level of DCP secretion increased by several fold in the sorafenib-treated and non-treated cells compared with the normoxic conditions. Prothrombin and VEGF values with normoxic conditions remained almost similar with hypoxic conditions. Under hypoxic conditions, NX-DCP significantly decreased below the control values for the first 48 h subsequent to sorafenib treatment, but significantly increased at 72 h. In vivo experiments demonstrated that sorafenib inhibited angiogenesis and tumor proliferation, but the levels of DCP and NX-DCP did not differ significantly from the controls. These findings indicate that the suppression of neovascularization by sorafenib promotes blood vessel ischemia, producing hypoxic conditions whereby vitamin K uptake and utilization efficiency is reduced.