Positional isomers of monopegylated interferon alpha-2a: isolation, characterization, and biological activity.
The success of recombinant interferon alpha in the clinic in part is limited by two properties of the protein: short serum half-life and immunogenicity. To improve these properties, interferon alpha-2a was conjugated with polyethylene glycol (PEG-5000). A homogeneous preparation of monopegylated interferon alpha-2a was subjected to vigorous analytical and activity characterization. A newly developed ampholyte-free chromatofocussing-like cation-exchange HPLC method utilizing a sulfopropyl resin was used to separate the monopegylated protein into 11 species. Peptide mapping, sequencing, and mass spectrometric analyses indicated that these species are positional isomers where each isomer represents a single polymer molecule conjugated to one specific lysine residue. No species with a modification at the amino terminus was observed. All 11 isomers show antiviral and antiproliferative activities in the same range as the parent monopegylated interferon alpha-2a.
Monkarsh
S P
SP
Department of Biopharmaceuticals, Hoffmann-La Roche Inc., Nutley, New Jersey 07110, USA.
Ma
Y
Y
Aglione
A
A
Bailon
P
P
Ciolek
D
D
DeBarbieri
B
B
Graves
M C
MC
Hollfelder
K
K
Michel
H
H
Palleroni
A
A
Porter
J E
JE
Russoman
E
E
Roy
S
S
Pan
Y C
YC
D000818:Animals; D000998:Antiviral Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D003588:Cytopathogenic Effect, Viral / Q000187:drug effects; D004285:Dogs; D006801:Humans; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D007536:Isomerism; D015394:Molecular Structure; D011092:Polyethylene Glycols / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D011994:Recombinant Proteins
D000998:Antiviral Agents; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; C100416:peginterferon alfa-2a