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1.
J Biol Chem ; 284(46): 32108-15, 2009 Nov 13.
Article in English | MEDLINE | ID: mdl-19717564

ABSTRACT

A major limitation in biopharmaceutical development is selectively targeting drugs to diseased tissues. Growth factors and viruses have solved this problem by targeting tissue-specific cell-surface heparan sulfates. Neuregulin (NRG), a growth factor important in both nervous system development and cancer, has a unique heparin-binding domain (HBD) that targets to cell surfaces expressing its HER2/3/4 receptors (Esper, R. M., Pankonin, M. S., and Loeb, J. A. (2006) Brain Res. Rev. 51, 161-175). We have harnessed this natural targeting ability of NRG by fusing the HBD of NRG to soluble HER4. This fusion protein retains high affinity heparin binding to heparin and to cells that express heparan sulfates resulting in a more potent NRG antagonist. In vivo, it is targeted to peripheral nerve segments where it blocks the activity of NRG as a Schwann cell survival factor. The fusion protein also efficiently blocks autocrine and paracrine signaling and reduces the proliferation of MCF10CA1 breast cancer cells. These findings demonstrate the utility of the HBD of NRG in biopharmaceutical targeting and provide a new way to block HER signaling in cancer cells.


Subject(s)
Autocrine Communication , ErbB Receptors/metabolism , Heparin/metabolism , Heparitin Sulfate/metabolism , Neuregulin-1/metabolism , Paracrine Communication , Animals , Apoptosis , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , CHO Cells , Cell Proliferation , Chick Embryo , Cricetinae , Cricetulus , ErbB Receptors/antagonists & inhibitors , ErbB Receptors/genetics , Female , Humans , Molecular Sequence Data , Neuregulin-1/genetics , Phosphorylation , Receptor, ErbB-4 , Schwann Cells
2.
Brain Res ; 1258: 1-11, 2009 Mar 03.
Article in English | MEDLINE | ID: mdl-19150438

ABSTRACT

The neuregulins are a family of polypeptide factors implicated in a wide range of neurological and psychiatric disorders including multiple sclerosis, schizophrenia, and Alzheimer's disease. Many alternatively-spliced forms of the NRG1 gene are released as soluble factors that can diffuse to near and distant sites within the nervous system where they can accumulate through binding to highly specific heparan-sulfate proteoglycans in the extracellular matrix. Here we have determined the sites of synthesis and accumulation of heparin-binding neuregulin forms in human neocortex, white matter, cerebral spinal fluid, and serum by immunostaining and measurement of neuregulin activity. While neuregulin precursors are expressed predominately within cortical neurons, soluble neuregulin accumulates preferentially on the surface of white matter astrocytes. Consistently, neuregulin activity can be released from the extracellular matrix of human brain by protease treatment. Neuregulin activity is also detectable in human cerebral spinal fluid where its expression appears to be altered in neuronal disorders. While cerebral spinal fluid neuregulin levels were unaltered in patients with multiple sclerosis, they were slightly reduced in amyotrophic lateral sclerosis and Parkinson's disease (p<0.15), but significantly increased in Alzheimer's disease (p<0.01). While not detected in human serum, a novel neuregulin antagonist activity was identified in human serum that could have prevented its detection. These results suggest that human neuregulin is selectively targeted from cortical neurons to white matter extracellular matrix where it exists in steady-state equilibrium with cerebral spinal fluid where it has the potential to serve as a biological marker in human neuronal disorders.


Subject(s)
Brain/metabolism , Neuregulin-1/cerebrospinal fluid , Neuregulin-1/metabolism , Alzheimer Disease/cerebrospinal fluid , Amyotrophic Lateral Sclerosis/cerebrospinal fluid , Astrocytes/metabolism , Autopsy , Blotting, Western , Extracellular Matrix/metabolism , Heparin/metabolism , Humans , Immunohistochemistry , Multiple Sclerosis/cerebrospinal fluid , Nerve Fibers, Myelinated/metabolism , Neuregulin-1/blood , Neurons/metabolism , Parkinson Disease/cerebrospinal fluid
3.
Brain Res Rev ; 51(2): 161-75, 2006 Aug.
Article in English | MEDLINE | ID: mdl-16412517

ABSTRACT

The neuregulins are a family of growth and differentiation factors with a wide range of functions in the nervous system. The power and diversity of the neuregulin signaling system comes in part from a large number of alternatively-spliced forms of the NRG1 gene that can produce both soluble and membrane-bound forms. The soluble forms of neuregulin are unique from other factors in that they have a structurally distinct heparin-binding domain that targets and potentiates its actions. In addition, a finely tuned, bidirectional mechanism regulates when and where neuregulin is released from neurons in response to neurotrophic factors produced by both neuronal targets and supporting glial cells. Together, this produces a balanced intercellular signaling system that can be localized to distinct regions for both normal development and maintenance of the mature nervous system. Recent evidence suggests that neuregulin signaling plays important roles in many neurological disorders including multiple sclerosis, traumatic brain and spinal cord injury, peripheral neuropathy, and schizophrenia. Here, we review the basic biology of neuregulins and relate this to research suggesting their involvement with and potential therapeutic uses for neurological disorders.


Subject(s)
Brain Diseases/metabolism , Central Nervous System/metabolism , Growth Substances/metabolism , Neuregulins/metabolism , Animals , Brain Diseases/genetics , Brain Diseases/physiopathology , Cell Communication/physiology , Cell Differentiation/physiology , Central Nervous System/embryology , Central Nervous System/growth & development , Growth Substances/chemistry , Growth Substances/genetics , Humans , Neuregulins/chemistry , Neuregulins/genetics , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , Signal Transduction/physiology , Solubility
4.
J Biol Chem ; 280(1): 383-8, 2005 Jan 07.
Article in English | MEDLINE | ID: mdl-15528194

ABSTRACT

Neuregulins are a family of growth and differentiation factors that act through activation of cell-surface erbB receptor tyrosine kinases and have essential functions both during development and on the growth of cancer cells. One alternatively spliced neuregulin-1 form has a distinct heparin-binding immunoglobulin-like domain that enables it to adhere to heparan sulfate proteoglycans at key locations during development and substantially potentiates its activity. We examined the structural specificity needed for neuregulin-1-heparin interactions using a gel mobility shift assay together with an assay that measures the ability of specific oligosaccharides to block erbB receptor phosphorylation in L6 muscle cells. Whereas the N-sulfate group of heparin was most important, the 2-O-sulfate and 6-O-sulfate groups also contributed to neuregulin-1 binding in these two assays. Optimal binding to neuregulin-1 required eight or more heparin disaccharides; however, as few as two disaccharides were still able to bind neuregulin-1 to a lesser extent. The physiological importance of this specificity was shown both by chemical and siRNA treatment of cultured muscle cells. Pretreatment of muscle cells with chlorate that blocks all sulfation or with an siRNA that selectively blocks N-sulfation significantly reduced erbB receptor activation by neuregulin-1 but had no effect on the activity of neuregulin-1 that lacks the heparin-binding domain. These results suggest that the regulation of glycosaminoglycan sulfation is an important biological mechanism that can modulate both the localization and potentiation of neuregulin-1 signaling.


Subject(s)
Heparan Sulfate Proteoglycans/metabolism , Heparin/metabolism , Neuregulin-1/metabolism , Signal Transduction , Animals , Binding Sites , Cell Line , ErbB Receptors/agonists , ErbB Receptors/metabolism , Heparan Sulfate Proteoglycans/chemistry , Heparin/chemistry , Phosphorylation , Protein Binding , RNA, Small Interfering/metabolism , RNA, Small Interfering/pharmacology , Rats , Structure-Activity Relationship , Sulfates
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