Specific immunotherapeutic targeting of malignant hepatocytes enabled by transient and controlled ATP depletion
Final Report Abstract
TNF was the first cytokine employed for cancer therapy, but its use was limited due to insufficient selectivity towards malignant cells. Fructose induces hepatic ATP depletion in humans and rodents due to the liver-specific fructose metabolism via fructokinase, while other cells metabolize fructose via hexokinase. Under ATP depleted conditions hepatocytes are protected against TNF-induced apoptosis. Our aim was to identify metabolic differences between normal and malignant liver cells that can be exploited for selective immunotherapy. We analyzed the expression and activities of enzymes involved in fructose metabolism in primary rodent and human hepatocytes and hepatoma cell lines. Furthermore, we studied the influence of hexokinase II (HKII) on fructose-mediated ATP depletion and cytoprotection in murine hepatocytes by overexpression and hypoxia-mediated upregulation of HKII. Primary mouse, rat and human hepatocytes depleted of ATP by fructose were fully protected against TNF-induced cytotoxicity. By contrast, hepatic tumor cell lines lacked the fructose-mediated ATP depletion mechanism and, therefore, remained susceptible to TNF/ActD-induced apoptosis. The TNF-sensitive hepatoma cells showed increased expression of HKII. Inhibition of hexokinases by 3-bromopyruvate restored fructose-induced ATP depletion in HepG2 cells, and stabilization of endogenous hypoxia-inducible factor1 (HIF1) increased expression of HKII in primary murine hepatocytes, which in turn prevented fructose-induced ATP depletion. Finally, inhibition of TNF-induced apoptosis by fructose was prevented by overexpression of HKII in primary murine hepatocytes. We conclude that increased expression of HKII in malignant cells of hepatic origin shifts the liver-type to the muscle-type fructose metabolism, thereby preventing ATP depletion and subsequent cytoprotection of the target cells. Therefore, healthy liver cells are transiently protected from TNF-mediated cell death by fructose-induced ATP depletion, while malignant cells can be selectively eliminated through TNF-induced apoptosis.
