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Inhibition of vascular permeability factor (vascular endothelial growth factor) with antipeptide antibodies
T M Sioussat1, H F Dvorak, T A Brock
1Department of Pathology, Beth Israel Hospital, Boston, Massachusetts.
Abstract:
Vascular permeability factor (VPF), also known as vascular endothelial cell growth factor (VEGF), is a 34- to 43-kDa dimeric protein synthesized and secreted by a variety of tumor and normal cells. At nanomolar concentrations, VPF causes an increase in microvascular permeability and is thought to be responsible for enhanced permeability of tumor blood vessels and for the fluid accumulation associated with solid and ascites tumors. In addition, VPF/VEGF is a mitogen for endothelial cells and may play an important role in maintaining vascular endothelium and in promoting tumor angiogenesis. Antibodies were raised against a series of synthetic peptides derived from the predicted human VPF amino acid sequence. The antibodies were assayed for their ability to bind native and denatured/reduced VPF. Antibodies to peptides from the N- and C-termini bound both denatured/reduced and native VPF; antibodies directed to internal segments (e.g., amino acids 27-48 and 85-101) strongly bound denatured/reduced VPF but were substantially less effective at binding native VPF. These results suggest that the N- and C-termini are exposed regions of the protein in solution. Individually, antibodies to the N- and C-termini each partially blocked VPF permeability activity, and, in combination, blocked nearly 100% of this activity. Also, the N- and C-terminal antibodies blocked the VPF-mediated stimulation of both endothelial cell growth and increase in free cytosolic calcium.
Insights
Antibodies targeting the N- and C-termini of vascular permeability factor (VPF), also known as vascular endothelial cell growth factor (VEGF), effectively block its ability to increase vascular permeability and stimulate cell growth.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Vascular permeability factor (VPF), or vascular endothelial cell growth factor (VEGF), is a protein secreted by various cells.
- VPF/VEGF increases microvascular permeability, contributing to tumor fluid accumulation and angiogenesis.
- It also acts as a mitogen for endothelial cells, crucial for vascular health.
Purpose of the Study:
- To develop antibodies against VPF/VEGF to investigate its structure and function.
- To determine which regions of VPF/VEGF are accessible in its native form.
- To evaluate the functional impact of blocking specific VPF/VEGF regions.
Main Methods:
- Antibodies were generated against synthetic peptides from the human VPF/VEGF sequence.
- Antibody binding was tested against native and denatured/reduced VPF/VEGF.
- The effect of antibodies on VPF/VEGF permeability, endothelial cell growth, and cytosolic calcium levels was assessed.
Main Results:
- Antibodies to N- and C-terminal peptides bound both native and denatured VPF/VEGF.
- Antibodies to internal peptides primarily bound denatured VPF/VEGF, indicating N- and C-termini exposure.
- N- and C-terminal antibodies individually partially blocked VPF/VEGF activity; combined, they blocked nearly all activity.
- These antibodies also inhibited VPF/VEGF-stimulated endothelial cell growth and calcium increase.
Conclusions:
- The N- and C-termini of VPF/VEGF are exposed in solution.
- Targeting these terminal regions with antibodies can effectively neutralize VPF/VEGF's biological activities.
- This suggests a potential therapeutic strategy for conditions involving VPF/VEGF dysregulation.