Conjugation of High-Molecular Weight Poly(ethylene glycol) to Cytokines: Granulocyte-Macrophage Colony-Stimulating Factors as Model Substrates

The ability of the small receptor-binding protein, recombinant murine granulocyte-macrophage colony-stimulating factor (GM-CSF), to increase the abundance of certain blood cell types in mice was enhanced markedly by covalent attachment of a single long strand of PEG (30-40 kDa). Potency was not increased further by coupling a second strand. Such conjugates can be synthesized efficiently by reaction of protein amino groups with PEG propionaldehydes in the presence of NaBH₃CN or with PEG p-nitrophenyl carbonates. Both methods have been used to prepare recombinant human GM-CSF conjugates of pre-determined composition, e.g. PEG₁GM-CSF and PEG2GM-CSF, in high yield. These compounds, or analogous derivatives of other cytokines, purified by ion-exchange and size-exclusion chromatography, may be suitable candidates for pharmaceutical development.