Biomedical Engineering Reference
In-Depth Information
parapox orf virus (OV) shares about 25% amino acid identity
to mammalian VEGF-A, but does not have heparin-binding
capability. Recombinant proteins were generated VEGF
receptor agonist that stimulates proliferation and sprout
formation of macro- and microvascular endothelial cells.
At the same time, the fusion protein was a more potent
competitor of VEGF-A than the wild-type VEGF-E in
binding to primary human endothelial cells.
Insulin-like growth factor-1 (IGF-1) is a small and highly
diffusible growth factor well known as an important media-
tor of cell growth and differentiation [53]. However, sys-
temic administration of IGF-1 has significant unwanted side
effects in tissues other than those of interest [54]. To try to
solve this problem, a fusion protein consisting of the IGF-1
polypeptide and the HBD of heparin-binding epidermal
growth factor-like growth factor (HB-EGF) was developed
[55]. This fusion protein bound selectively to heparin as well
as cell surfaces, without showing any reduction in biological
activity. Moreover, the fusion protein was selectively
retained by cartilage explants enriched with HSPGs, sug-
gesting an improved and localized delivery of the growth
factor to where it was needed.
Recent studies from our group have shown that the
interaction of NRG1's HBD with HSPGs leads to targeted
deposition of the factor in specific regions of the developing
nervous system [67]. In vitro studies suggest that the HBD
facilitates the accumulation of NRG1 at HSPG-rich synaptic
basal lamina and provides a sustained source of ligand
required for sustained erbB receptor activation, important
for the induction of acetylcholine receptor gene expression
[1,65,67,69]. Structural heparin-binding studies using vari-
ous desulfated heparins showed the importance as well as
specificity of these sulfate groups by comparing the effects
of fully sulfated heparin to either completely desulfated or
partially desulfated heparin using a gel mobility shift assay
[68]. Both the completely desulfated heparin and the De-N-
sulfated sugars were unable to bind NRG1, suggesting a
critical importance of N-sulfation for NRG1-heparin bind-
ing (Figure 27.2a). Removing the 2-O- and 6-O-sulfate
groups, however, also reduced NRG1-heparin binding, but
to a lesser extent (Figure 27.2b). Moreover, N-sulfate groups
were needed for NRG1 to accumulate on the cell surface and
activate its erbB receptors. Thus, NRG1 has a unique,
structurally distinct, and highly tissue-specific HBD that
is critical for its tissue localization during development as
well as for optimal signal transduction.
27.8 THE NEUREGULIN 1 GROWTH FACTOR
HAS A UNIQUE AND HIGHLY SPECIFIC HBD
27.9 USING NEUREGULIN'S HBD TO GENERATE
A TARGETED NEUREGULIN ANTAGONIST
The neuregulin 1 (NRG1) gene produces a large family of
alternatively spliced growth and differentiation factors with
a wide range of important functions in nervous system and
heart development as well as in human diseases ranging
from breast cancer to schizophrenia [56-62]. Although
NRG1 has many isoforms generated through alternative
splicing, all forms share a highly conserved and tightly
folded epidermal growth factor (EGF) -like domain, which
binds EGF receptor family members HER3 (erbB3) and
HER4 (erbB4), which can heterodimerize with HER2
(erbB2) to transduce signals through an intracellular tyro-
sine kinase domain [57,60]. The receptor activation initiates
several downstream signaling pathways including the mito-
gen-activated protein (MAP) kinase and phosphatidylinosi-
tol 3-kinase (PI3K) and mediates diverse effects on cellular
processes including survival, proliferation, and differentia-
tion [63]. A key feature of the secreted forms of NRG1 is a
spatially separated heparin-binding domain (HBD) that is
unique from other HBDs because of its disulfide-linked C2
immunoglobulin domain structure that allows it to be capa-
ble of maintaining its structural specificity for high-affinity
heparin binding [64]. This C2 loop consists of a long stretch
of alternating positively charged amino acids and is sepa-
rated from the EGF-like domain by a glycosylated spacer.
The net result is a highly specific HBD that recognizes
specific sulfate groups on HS and results in precise tissue
localization during development [65-68].
Because of the established role of NRG1 and its family of
EGF receptors to promote the growth of breast cancer or
other types of solid tumors as well as its potential role in
neurological and psychiatric diseases, augmenting or block-
ing NRG1-erbB signaling has become an attractive thera-
peutic strategy [1,70-73]. A number of efforts have been
made to block NRG signaling, including the FDA approved
humanized monoclonal antibody drug against the erbB2
receptor called Herceptin [74]. It has become a clinically
effective adjuvant therapy for a subgroup of breast cancer
patients [75,76] but has to be used at extremely high
concentrations that have been linked to cardiac side effects
and only works on a subgroup of breast cancer patients that
overexpress HER2 [77,78]. A major obstacle of blocking
NRG1 signaling in both cancer and developmental studies
comes from its ability to become highly concentrated in the
ECM where it can produce sustained HER activation
required for downstream gene activations [1,65]. All these
studies provide a rationale to develop novel effective drugs
with better targeting specificity that block NRG1 in breast
cancers or other tumors that proliferate in response to NRG1
signaling.
To solve this important problem, we generated a novel
fusion protein called HBD-S-H4 that fuses the human
NRG1's HBD to the soluble ectodomain of human
Search WWH ::
Custom Search