Biomedical Engineering Reference
In-Depth Information
circulating half-life of peptides or some proteins by fusing
them to HSA on either the N- or C-terminus has been
developed and proven to be successful in drug development.
One major difference between Fc fusions and HSA-fusion
protein is that the Fc fusions are homodimers but the HSA
fusions are monomers. The most clinically advanced exam-
ple of peptide-HSA fusion, Syncria
1
(albiglutide), a gluca-
gon-like peptide-1 (GLP-1) peptide genetically fused to
HSA, created originally by Human Genome Sciences and
licensed to GlaxoSmithKline, is in Phase III clinical trials
for the treatment of type 2 diabetes mellitus [28]. Several
other HSA-fusion therapeutics are in different stages of
clinical trials, including Zalbin, an interferon (IFN)-
a
2b
fused to HSA which was codeveloped by Human Genome
Science and Novartis (discontinued in development), for
treatment of chronic hepatitis C; Albugranin, a granulocyte
colony-stimulating factor fused to HSA, for neutropenia;
Albulin, an insulin fused to HSA for diabetes; and AlbuBNP
(also called Cardeva
TM
), the B-type natriuretic peptide fused
to HSA for chronic heart failure [29-32].
processes by allowing application of the same established
procedures for antibody drug manufacturing [36].
8.4.1 The Structure of Fc-Fusion Proteins
In 1989, Daniel Capon at Genetech described the first
successful example of using a recombinant soluble receptor
to block ligand activity [37]. They fused the ligand-binding
domains D1-D2 of the cluster of differentiation 4 (CD4) to
the N-terminal of the immunoglobulin G1 (IgG1) Fc domain
and demonstrated that the fusion protein can specifically
inhibit CD4 activities [38,39]. Since the fusion protein was
composed of IgG constant region and ligand-binding
domains of CD4, a cell adhesion molecule, it was called
an immunoadhesin. These types of proteins are now also
commonly referred to as Ig-fusion or Fc-fusion proteins
[40]. Figure 8.1A illustrates the overall generic structure of a
typical Fc-fusion protein, for example, a receptor-Fc-fusion
protein. The C-terminus of the ligand-binding domain or the
whole soluble extracellular domain (ECD) of a single recep-
tor chain is genetically fused to the N-terminus of the hinge
region, followed by the CH2 and CH3 domains of a human
IgG heavy chain. Similar to antibodies or other Fc-contain-
ing molecules, the fusion protein forms a homodimer owing
to the strong interactions of the CH3 domains. The homo-
dimer is stabilized by interchain disulfide bonds in the hinge
regions. This covalently linked homodimer structure pro-
vides some of the antibody-like structural properties, such as
high affinity binding due to the avidity effect. The hinge
regions provide flexible spacers between the two receptor
ECDs and the IgG constant regions to allow each part of the
molecule the necessary flexibility to function independently.
Several structural variations from the traditional Fc-
fusion structure were developed, including the heterodi-
meric Fc-fusion protein, which contains two distinct fusion
partners as shown in Figure 8.1E and monomeric Fc-fusion
protein in which only one fusion partner is attached to the
dimeric Fc region (Figure 8.1C) [41,42]. The most advanced
monomeric Fc fusion is human coagulation Factor IX-Fc for
hemophilia B currently in the Phase IIa clinical trials (Table
8.1). The drug is being developed by Biogen Idec (formerly
Syntonix) to improve the pulmonary delivery efficiency of
Fc-fused proteins by reducing the molecular size. Compared
to traditional Fc-fusion proteins, monomeric Fc-fusion pro-
teins have shown higher bioavailability for pulmonary deliv-
ery in nonhuman primates, longer circulating half-life, and
better biological activities [52,53].
8.3.3 Other Approaches
Several other approaches, including the use of albumin-bind-
ing peptides or proteins to attach to serumalbumin, developed
by Genentech and Dyax, and the use of chemical conjugation
of peptides to covalently link toHSA to extend serumhalf-life,
were reviewed by McGregor [26]. Another interesting
approach was developed by CovX (acquired by Pfizer),
in which an antibody having aldolase activity catalyzes the
site-specific covalent attachment of peptides to itself [33].
8.4 Fc-FUSION PROTEINS IN DRUG
DEVELOPMENT
The Fc-fusion technique is the most used and successful
technique for extension of serum half-life in both research
and drug development. Many of the favorable attributes that
have contributed to the successful utilization of antibodies as
drugs are embodied in the immunoglobulin constant region
or Fc domain. The Fc domain prolongs the serum half-life of
proteins, mainly because of the pH-dependent binding of the
neonatal Fc receptor (FcRn), which salvages the protein
from being degraded in endosomes [34,35] and to limited
renal clearance resulting from the increase of mass due to the
homodimeric nature of Fc-fusion proteins. Fc-dependent
effector functions are important attributes for many indica-
tions, particularly for oncology applications. The Fc region
in many cases improves the biophysical properties of its
fusion partner, such as the solubility and stability of the
protein. In addition, high expression, secretion to tissue
culture medium, and protein-A affinity purification of
Fc-fusion proteins simplify the downstream manufacturing
8.4.2 Fc-Fusion Therapeutics
Fc-fusion proteins have been widely used as research tools to
study the structure and function of many receptors, identify
and isolate novel ligands, and understand better the recep-
tor-ligand interactions [54,55]. More significantly,
they
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