With the completion of the Human Genome Project, molecular biologists are continuously discovering new classes of proteins
and new therapeutic uses for known proteins. The ~30,000 genes defined by the Human Genome Project translate into 300,000
to one million proteins.1 Converting these proteins into effective biopharmaceuticals, however, may require challenging formulation development, because
mostrecombinant proteins have limited chemical or physical stability in liquid state. For this reason, a renewed interest in conjugating
proteins with polyethylene glycol (PEG) can be expected, with the final goal of bringing PEGylated protein drugs to market.
PROTEIN PROPERTIES AND PEGYLATION USE
Depending on their ionic charge, size, and structure, proteins vary in their thermal stability, solubility, and susceptibility
to proteolysis. The intermolecular packing and surface chemistry of proteins determine many of these biopharmaceutical properties,
and many stress factors can cause protein unfolding and degradation, ultimately leading to loss of biological activity.2 Slight changes in pH, ionic strength, or temperature, for example, all can reduce biological activity in vivo.3 Other potential stress factors include proteases and oxidation. In vivo, theseprotein propertiescan translate into a high clearance rate of a therapeutic protein from the body. In addition, short plasma half-life and reactions
with the immune system complicate effective delivery of therapeutic proteins in humans.4 Bypassing these problems can be accomplished by either stabilizing the proteins or increasing their solubility, thus allowing
for low dosage volumes and longer circulation times.
To achieve the desired stability and solubility, proteins can be modified using methods such as crosslinking,5 fusion to other proteins, changing the oligomerization state, glycosylation, mutations of cysteine residues, or polymer
attachment.6 Currently, one of the current most successful methods for stabilizing proteins and increasing their solubility is to use
polymer therapeutics, i.e., to link an active molecule to a polymer molecule such as polyethylene glycol (PEG). Polymer therapeutics
includes polymer drugs, polymer conjugates, and polymeric micelles.7 In general, conjugating a protein to a polymer can accomplish several desirable objectives: a longer in vivo half-life; reduced immunogenicity, toxicity, and clearance rate through the kidneys; successful transportation across a cell
membrane; protection against proteolysis; modification of electro-osmotic flow; increased pH and thermal stability; a low
volume of distribution and sustained adsorption from the injection site; and improved formulation properties of the protein.
These superior properties can increase effective potency8 , improve response to the drug, increase patient tolerance and reduce side effects, reduce overall dosage, decrease office
visits, and lower the cost to the patient. A better drug profile and an improved quality of life are thus achieved.9 In the biotechnology industry, better biophysical characterization and understanding of PEGylated protein properties would
allow for better control of the conjugation reaction and an improved drug product.
POLYETHYLENE GLYCOL'S PROPERTIES
PEG is FDA approved for human administration by mouth, injection, or dermal application. It has been used for many years in
various capacities. Early work used PEG in crystallography for crystal growth, purification in two-phase systems, incorporation
into liposomes for an increased serum lifetime, or attachment to surfaces to reduce protein adsorption.10 In addition, PEG has been used to promote correct protein folding.
A. Sorina Morar, PhD, is a scientist, Process Development, Diosynth Biotechnology, 3000 Weston Parkway, Carey, NC 25713, 919.388.5649, fax: 919.678.0366, firstname.lastname@example.org.
Articles by A. Sorina Morar, PhD