Abstract
Recombinant human interferon-β (rhIFN-β) is the leading therapeutic intervention shown to change the cause of relapsing-remitting multiple sclerosis, and both a nonglycosylated and a significantly more active glycosylated variant of rhIFN-β are used in treatment. This study investigates the function of the rhIFN-β1a glycan moiety and its individual carbohydrate residues, using the myxovirus resistance (Mx) mRNA as a biomarker in Mx-congenic mice. We showed that the Mx mRNA level in blood leukocytes peaked 3 h after s.c. administration of rhIFN-β1a. In addition, a clear dose-response relationship was confirmed, and the Mx response was shown to be receptor-mediated. Using specific glycosidases, different glycosylation analogs of rhIFN-β1a were obtained, and their activities were determined. The glycosylated rhIFN-β1a showed significantly higher activity than its deglycosylated counterpart, due to a protein stabilization/solubilization effect of the glycan. It is interesting to note that the terminating sialic acids were essential for these effects. Conclusively, the structure/bioactivity relationship of rhIFN-β1a was determined in vivo, and it provided a novel insight into the role of the rhIFN-β1a glycan and its carbohydrate residues. The possibilities of improving the pharmacological properties of rhIFN-β1a using glycoengineering are discussed.
Footnotes
-
This work was supported by grants from Warwara Larsen Foundation, Novo Nordisk/Novozymes, Lundbeck Foundation, and the Danish Multiple Sclerosis Society.
-
L.D.-O. and M.T.-A. contributed equally to this work.
-
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
-
doi:10.1124/jpet.108.138263.
-
ABBREVIATIONS: IFN, interferon(s); IFNAR, IFN-α receptor; Mx, myxovirus resistance; rh, recombinant human; RRMS, relapsing-remitting multiple sclerosis; ko, knockout; HSA, human serum albumin; hpt, hours posttreatment; HPRT1, hypoxanthine phosphoribosyltransferase 1; SQ, starting quantity; bp, base pair; PAGE, polyacrylamide gel electrophoresis; MeCN, acetonitrile; FA, formic acid; MALDI-TOF, matrix-assisted laser desorption ionization time-of-flight; MS, mass spectrometry; ANOVA, one-way analysis of variance; NT, no treatment.
- Received February 20, 2008.
- Accepted April 28, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|