Abstract
Abstract Background: Most HCC presents at an advanced stage, for which there are few effective treatments. Sorafenib enhanced survival by 10 weeks in a randomized trial and was approved by FDA. Sorafenib is a base for building better treatments. Regorafenib (Fluoro-Sorafenib, BAY 73-4506 or Stivarga) is a more potent analog in current clinical trials. Methods: HCC cell lines were grown in culture. Growth was assessed using MTT, migration by scratch assay, medium AFP by clinical chemoluminescent immunometric method, cell AFP and kinases by western blot and cell AFP mRNA by real-time PCR. Results: HCC growth in different cell lines was inhibited by Regorafenib 1-5 μM IC50. Removal of drug from the medium resulted in resumption of normal cell growth even after subsequent sub-culture. This recovery could be partially blocked by low concentrations of vitamin K1 or doxorubicin that were not growth inhibitory when used alone. Regorafenib 1 μM pre-treatment for 2 weeks resulted in very slow recovery. Cell treatment in culture with Regorafenib resulted in early decreases in phospho-ERK levels, followed by return to baseline levels within hours, despite continuous drug exposure. Cell growth recovery after Regorafenib was associated with induction of high phospho-JNK levels and the recovery was blocked by the actions of a JNK inhibitor. Regorafenib also inhibited cell migration and both AFP mRNA by PCR and cellular AFP on WB, and cell culture medium levels of AFP, but at much lower concentrations than needed (0.1-1 μM) for growth inhibition. Removal of drug also resulted in recovery of cell migration, but this recovery could be partially blocked by addition of vitamin K1 or doxorubicin, at drug concentrations that were not themselves inhibitory of cell migration. Even at 1 μM, Regorafenib slowed cell growth and migration after prolonged cell exposure and sub-culture. Pre-treated and recovering cells were much more sensitive to inhibition of migration by low concentration vitamin K1 6.25 μM or doxorubicin 0.01 μM. Conclusions: Regorafenib is a potent inhibitor of HCC cell line growth and works even at very low drug concentrations when cells are exposed for prolonged times. The effects are reversible and JNK could be a critical player in this process. Very low concentrations inhibit HCC cell migration and AFP. Regorafenib appears to be a promising new agent. Citation Format: Brian I. Carr, Rosalba D'Alessandro, Maria Grazie Refolo, Minna Iacovazzi, Catia Lippolis, Caterina Messa, Aldo Cavallini. Regorafenib-mediated potent and reversible inhibition of human hepatocellular carcinoma (HCC) cell growth, migration and AFP. [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 1055. doi:10.1158/1538-7445.AM2013-1055
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