Vitamin B12 mediated oral delivery systems for granulocyte-colony stimulating factor and erythropoietin

As a prelude to the development of orally active erythropoietin (EPO) and granulocyte-colony stimulating factor (G-CSF), conjugates have been formed between these molecules and vitamin B12. During the formation of these conjugates intramolecular cross-linking of the proteins was avoided by the use of hydrazidyl derivatives of vitamin B12. A potentially biodegradable linkage was formed between vitamin B12 and G-CSF by reaction of the buried thiol in G-CSF with a long chain dithiopyridyl derivative of vitamin B12. In vitro and in vivo testing of the conjugates showed that their bioactivity was substantially maintained and that they were actively transported in an intrinsic factor dependent fashion across CaCo-2 cells and from the intestine to the circulation in a biologically active form.