Targeted antagonism of galactosamine toxicity in normal rat hepatocytes in vitro.
We present evidence that normal hepatocytes can be specifically protected from galactosamine toxicity in vitro by targeting an antagonist to these cells via receptor-mediated endocytosis. The strategy is based upon the following principles: 1) galactosamine is a highly selective hepatotoxin that causes a dose-dependent depletion of uridine intermediates; 2) galactosamine toxicity can be antagonized by supplemental administration of uridine; 3) normal hepatocytes possess unique cell-surface receptors that can internalize galactose terminal (asialo-)glycoproteins with subsequent degradation of the glycoprotein ligand. Based on these facts, we hypothesized that chemical coupling of a galactosamine antagonist to an asialoglycoprotein could result in cell-specific delivery and protection of normal hepatocytes by targeting the antagonist via asialoglycoprotein receptors. Using a model system consisting of freshly isolated rat hepatocytes (receptor (+)) and Morris 7777 rat hepatoma (receptor (-)) cells, sensitivity to galactosamine in vitro was determined and found to be similar for both types of cells. A targetable antagonist was synthesized by coupling uridine monophosphate to asialoorosomucoid in a molar ratio of 5 to 1. Exposure of Morris 7777 cells to the targetable antagonist in the presence of a toxic concentration of galactosamine did not protect these cells as evidenced by a steady decline in the number of viable cells in a fashion identical to cells treated with galactosamine alone. However, normal hepatocytes that received the conjugate in the presence of galactosamine were protected as their viable cell number remained the same as control (untreated) cells. Competition by an excess of asialoglycoprotein inhibited the protective effect of the conjugate, supporting the concept that the asialoglycoprotein component of the conjugate was responsible for the specific delivery of the antagonist to the target cells.
Wu
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Department of Medicine, University of Connecticut School of Medicine, Farmington 06032.
Keegan-Rogers
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Franklin
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Midford
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Wu
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D000818:Animals; D001212:Asialoglycoproteins; D002478:Cells, Cultured; D005688:Galactosamine / Q000037:antagonists & inhibitors / Q000633:toxicity*; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008114:Liver Neoplasms, Experimental / Q000378:metabolism; D008297:Male; D009961:Orosomucoid / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D011107:Polylysine / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D014542:Uridine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology
D001212:Asialoglycoproteins; D009961:Orosomucoid; C017030:asialoorosomucoid; C055350:asialoorosomucoid-polylysine-UMP conjugate; D011107:Polylysine; D005688:Galactosamine; D014542:Uridine Monophosphate