Survival of recombinant erythropoietin in the circulation: the role of carbohydrates.
Recombinant human erythropoietin produced in transfected Chinese hamster ovary cells is glycosylated much the same way as the erythropoietin present in human urine. To determine the role of carbohydrates in the stability of recombinant human erythropoietin in vivo, [125I]-labeled recombinant erythropoietin was intravenously infused into rats. The erythropoietin was slowly cleared from the blood with a half-life of approximately two hours. Asialoerythropoietin, which was produced by treatment of recombinant human erythropoietin with sialidase, was found to be cleared rapidly from circulation within ten minutes. These data suggest that the galactose binding protein of hepatic cells is involved in the clearance of asialoerythropoietin. Erythropoietin also contains N-glycans with a few N-acetyllactosamine repeats, which can be enriched by tomato lectin affinity chromatography. The lectin-bound fraction was cleared to a larger extent than was the unfractionated erythropoietin, while the component that did not bind the lectin was found to be stable in the circulation. Authentic N-acetyllactosamine repeats (polylactosaminoglycans) prepared from erythrocytes were similarly rapidly cleared from the circulation to the liver, and this clearance was inhibitable with asialo-alpha 1-acid glycoprotein. These results suggest that (a) the sialic acid of the recombinant erythropoietin is necessary for this glycoprotein hormone to circulate stably and (b) glycoproteins with more than three lactosaminyl repeat units may be cleared by the galactose binding protein of hepatocytes.
Fukuda
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La Jolla Cancer Research Foundation, Cancer Research Center, CA 92037.
Sasaki
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H
Lopez
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L
Fukuda
M
M
D000606:Amino Sugars; D000818:Animals; D002236:Carbohydrate Conformation; D002241:Carbohydrates / Q000097:blood / Q000502:physiology*; D006224:Cricetinae; D004355:Drug Stability; D004591:Electrophoresis, Polyacrylamide Gel; D004921:Erythropoietin / Q000097:blood* / Q000493:pharmacokinetics; D006801:Humans; D008297:Male; D008657:Metabolic Clearance Rate; D019158:N-Acetylneuraminic Acid; D051381:Rats; D011919:Rats, Inbred Strains; D011994:Recombinant Proteins / Q000097:blood* / Q000493:pharmacokinetics; D012794:Sialic Acids / Q000502:physiology; D012967:Sodium Dodecyl Sulfate; D014018:Tissue Distribution
D000606:Amino Sugars; D002241:Carbohydrates; D011994:Recombinant Proteins; D012794:Sialic Acids; D004921:Erythropoietin; D012967:Sodium Dodecyl Sulfate; C000458:N-acetyllactosamine; D019158:N-Acetylneuraminic Acid