Inhibition of glycoprotein synthesis in the endoplasmic reticulum as a novel anticancer mechanism of (-)-epigallocatechin-3-gallate.

(-)-Epigallocatechin-3-gallate (EGCG) has been found to trigger the unfolded protein response (UPR) likely due to the inhibition of glucosidase II, a key enzyme of glycoprotein processing and quality control in the endoplasmic reticulum (ER). These findings strongly suggest that EGCG interferes with glycoprotein maturation and sorting in the ER. This hypothesis was tested in SK-Mel28 human melanoma cells by assessing the effect of EGCG and deoxynojirimycin (DNJ) on the synthesis of two endogenous glycoproteins. Both tyrosinase and vascular endothelial growth factor (VEGF) protein levels were remarkably reduced despite unaltered mRNA expression in EGCG- or DNJ-treated cells compared to control. The hindrance of tyrosinase and VEGF protein synthesis could be prevented by proteasome inhibitor, lactacystine. Collectively, our results support that glucosidase II inhibitor EGCG interferes with protein processing and quality control in the ER, which diverts tyrosinase, VEGF, and likely other glycoproteins towards proteasomal degradation. This mechanism provides a novel therapeutic approach in dermatology and might play an important role in the antitumor effect or hepatotoxicity of EGCG.

Transport and transporters in the endoplasmic reticulum.

Enzyme activities localized in the luminal compartment of the endoplasmic reticulum are integrated into the cellular metabolism by transmembrane fluxes of their substrates, products and/or cofactors. Most compounds involved are bulky, polar or even charged; hence, they cannot be expected to diffuse through lipid bilayers. Accordingly, transport processes investigated so far have been found protein-mediated. The selective and often rate-limiting transport processes greatly influence the activity, kinetic features and substrate specificity of the corresponding luminal enzymes. Therefore, the phenomenological characterization of endoplasmic reticulum transport contributes largely to the understanding of the metabolic functions of this organelle. Attempts to identify the transporter proteins have only been successful in a few cases, but recent development in molecular biology promises a better progress in this field.

Glucuronide transport across the endoplasmic reticulum membrane is inhibited by epigallocatechin gallate and other green tea polyphenols.

Toxic endogenous or exogenous compounds can be inactivated by various conjugation reactions. Glucuronidation (i.e. conjugation with glucuronate) is especially important due to the large number of drugs and chemical carcinogens that are detoxified through this pathway. Stable and harmless glucuronides can be reactivated by enzymatic hydrolysis thus inhibitors of glucuronidase activity reduce the risk of chemical carcinogenesis. The aim of this study was to reveal whether this mechanism contributes to the anti-cancer effect of green tea flavanols, which has been shown in various animal models. Therefore, we investigated the effect of these polyphenols on deglucuronidation in rat liver microsomes and in Hepa 1c1c7 mouse hepatoma cells, using 4-methylumbelliferyl glucuronide as model substrate. Tea flavanols inhibited beta-glucuronidase in intact vesicles, where glucuronide transport across the microsomal membrane is rate-limiting, but were almost ineffective in permeabilized vesicles. Epigallocatechin gallate, the major green tea flavanol was shown to have a concentration-dependent inhibitory effect on both beta-glucuronidase activity and glucuronide transport in native vesicles. Epigallocatechin gallate also inhibited beta-glucuronidase activity in native Hepa 1c1c7 mouse hepatoma cells, while failed to affect the enzyme in alamethicin-permeabilized cells, where the endoplasmic membrane barrier was eliminated. Our findings indicate that tea flavanols inhibit deglucuronidation in the endoplasmic reticulum at the glucuronide transport stage. This phenomenon might potentially contribute to the cancer-preventing dietary or pharmacological effect attributed to these catechins.

Green tea flavonols inhibit glucosidase II.

Green tea is getting into the focus of scientific interest due to its beneficial health effects, most of which are attributed to its catechin content. Polyphenolic tea catechins have antioxidant, antiproliferative, antiangiogenic and proapoptotic effects, which makes them promising anticancer compounds. Other poly-hydroxy molecules have similar antitumor potentials through the inhibition of glucosidase II, which affects the glycoprotein maturation and quality control in the endoplasmic reticulum. We investigated the effect of tea catechins on glucosidase II activity in rat liver microsomes using 4-methylumbelliferyl glucoside and 4-nitrophenyl glucoside as substrates. A concentration-dependent inhibition with non-competitive kinetics was found. The IC50 and Ki values for certain tea catechins were comparable with those of N-butyldeoxynojirimycin, the widely used glucosidase inhibitor. The possible interference of tea catechins with the glycoprotein processing in the endoplasmic reticulum should be considered as a potential mechanism of their dietary or pharmacological effects.