The Drug development process - Pharmaceutical Biotechnology: Concepts and Aapplications - page 78

The macromolecular structure of drugs and the fact that relatively minor structural alterations
can potentially have a major infl uence upon bioactivity are often complicating factors. For ex-
ample, an immunoassay may be blind to the oxidation of an amino acid residue, or very limited
proteolytic processing, although such events can activate or decrease bioactivity.
As outlined previously, i.v. or s.c. administration is by far the most common delivery approach
in the context of biopharmaceuticals. Whole-body distribution studies are undertaken mainly in
order to assess tissue targeting and to identify the major elimination routes. The large molecular
weight of therapeutic proteins, along with additional properties (e.g. charge), generally impairs
their passage through biomembranes; hence, their initial distribution is usually limited to the
volume of the extracellular space (mainly the plasma volume). Distribution volume usually subse-
quently increases, as the protein is taken up into tissue during its metabolism/elimination.
The metabolism/elimination of therapeutic proteins occurs via processes identical to those pertain-
ing to native endogenous proteins. Ultimately this entails proteolytic degradation, with amino acid
residues released either being incorporated into newly synthesized protein or being further degraded
by standard metabolic pathways. Although the therapeutic protein may be subject to limited proteoly-
sis in the blood, extensive and full metabolism occurs intracellularly, subsequent to product cellular
uptake. Clearance of protein drugs from systemic circulation commences with passage across the
capillary endothelia. The rate of passage depends upon the protein's physicochemical properties (e.g.
mass and charge). Final product excretion is, in the main, either renal and/or hepatic mediated.
Many proteins of molecular mass 30 kDa are eliminated by the kidneys via glomerular fi l-
tration. In addition to size, fi ltration is also dependent upon the protein's charge characteristics.
Owing to the presence of glycosaminoglycans, the glomerular fi lter is itself negatively charged, so
negatively charged proteins are poorly fi ltered due to charge repulsion.
After initial fi ltration many proteins are actively reabsorbed (endocytosed) by the proximal
tubules and subjected to lysosomal degradation, with subsequent amino acid reabsorption. Thus,
very little intact protein actually enters the urine.
Uptake of protein by hepatocytes can occur via one of two mechanisms: (a) receptor-mediated en-
docytosis or (b) non-selective pinocytosis, again with subsequent protein proteolysis. Similarly, a pro-
portion of some proteins are likely degraded within the target tissue, as binding to their functional cell
surface receptors triggers endocytotic internalization of the receptor ligand complex (Figure 4.7).
Cellular uptake of some glycosylated therapeutic proteins occurs via specifi c sugar-binding cell
surface receptors. Cell surface mannose receptors, for example, are capable of binding glycopro-
teins whose sugar side chains terminate in mannose, fucose, N -acetyl glucosamine or N -acetyl
galactosamine. Evidence suggests that a liver-specifi c form of the mannose receptor mediates
clearance of luteinizing hormone (LH, Chapter 11). The sugar side chains of many glycoproteins
exhibit terminal sialic acid residues (sialic acid caps). The hepatic asialoglycoprotein receptor
binds glycoproteins whose sialic acid caps have been removed, likely mediating their removal
from general circulation.
Pharmacokinetic and indeed pharmacodynamic characteristics of therapeutic proteins can be
rendered (even more) complicated by a number of factors, including:

• The presence of serum-binding proteins. Some biopharmaceuticals (including insulin-like

growth factor (IGF), GH and certain cytokines) are notable in that the blood contains proteins
that specifi cally bind them. Such binding proteins can function naturally as transporters or acti-
vators, and binding can affect characteristics such as serum elimination rates.