Characterization of therapeutic proteins in disposition studies

Protein therapeutics must be quantitated in biologic matrices that often include the therapeutic's endogenous counterpart. The immunoassays and bioassays that are frequently the means of this quantitation are subject to numerous interferences and limitations, and may not provide adequate identification of the analyte. Radiolabels (125I external label, 3H, and 35S internal labels) may also be used to aid in the characterization of the in vivo fate of therapeutic proteins. Meaningful interpretation of results from protein disposition studies using radiolabels requires careful characterization of the structure and function of the labeled product. Both external and internal radiolabels are subject to in vivo loss of the label due to dehalogenation, or proteolytic liberation of labeled amino acids which may be rapidly reincorporated into unrelated, newly synthesized proteins. The advantages and disadvantages of various analytical approaches used in disposition studies will be described for tumor necrosis factor, insulin-like growth factor, relaxin, growth hormone and interferon-γ. It is concluded that a battery of analytical approaches is required to evaluate the molecular form of the detected radio-, immuno, or bioactivity. Ideally, PAGE, HPLC and other more discriminating analytical approaches should be an integral part of the evaluation of theapeutic protein disposition