AQUEOUS STABILITY OF RECOMBINANT HUMAN THROMBOPOIETIN AS A FUNCTION OF PROCESSING SCHEMES

Preformulation studies conducted with recombinant human thrombopoietin (rhTPO), a 332 amino acid glycoprotein which stimulates platelet prod uction, show distinctions in degradation profiles as a function of pro cessing schemes. The stability-limiting degradation pathways change as a function of purification stage and method and are dependent upon th e presence of contaminating protease. The stability-limiting degradati on pathway of affinity-purified and in-process rhTPO preparations is p rimarily attributed to proteolysis initiated by a protease present as a fermentation contaminant. The proteolysis increases with increasing pH as a function of temperature. The degradation profiles for these pr eparations show that bioactivity initially increases and then decrease s with increasing pH as a function of temperature. This is consistent with proteolysis to active forms which ultimately undergo degradation to less active forms. Similar studies conducted with rhTPO preparation s purified by a combination of more conventional chromatographic steps show different stability-limiting degradation pathways and a differen t pH-stability profile when compared to affinity purified or in-proces s preparations. In this case, degradation is accompanied by decreases in activity under all conditions, consistent with the conversion to le ss active forms. These results illustrate the importance of preformula tion and stability characterization of protein pharmaceuticals in supp ort of both process and formulation development. Issues related to sto rage and handling of in-process preparations differ from those with fo rmulated product since the stability-limiting degradation pathways cha nge as a function of purification stage.