Related Experiment Video
Updated: Jun 7, 2026
Intravital Microscopy of Tumor-associated Vasculature Using Advanced Dorsal Skinfold Window Chambers on Transgenic Fluorescent Mice
Published on: January 19, 2018
Vascular infarction by subcutaneous application of tissue factor targeted to tumor vessels with NGR-peptides:
Johannes Dreischalück1, Christian Schwöppe, Tilmann Spieker
1Department of Medicine A, University of Muenster, D-48129 Muenster, Germany.
Abstract:
tTF-NGR consists of the extracellular domain of the (truncated) tissue factor (tTF), a central molecule for coagulation in vivo, and the peptide GNGRAHA (NGR), a ligand of the surface protein aminopeptidase N (CD13). After deamidation of the NGR-peptide moiety, the fusion protein is also a ligand for integrin αvβ3 (CD51/CD61). Both surface proteins are upregulated on endothelial cells of tumor vessels. tTF-NGR showed binding to specific binding sites on endothelial cells in vitro as shown by flow cytometry. Subcutaneous injection of tTF-NGR into athymic mice bearing human HT1080 fibrosarcoma tumors induced tumor growth retardation and delay. Contrast enhanced ultrasound detected a decrease in tumor blood flow in vivo after application of tTF-NGR. Histological analysis of the tumors revealed vascular disruption due to blood pooling and thrombotic occlusion of tumor vessels. Furthermore, a lack of resistance was shown by re-exposure of tumor-bearing mice to tTF-NGR after regrowth following a first cycle of treatment. However, after subcutaneous (s.c.) push injection with therapeutic doses (1-5 mg/kg bw) side effects have been observed, such as skin bleeding and reduced performance. Since lethality started within the therapeutic dose range (LD10 approximately 2 mg/kg bw) no safe therapeutic window could be found. Limiting toxicity was represented by thrombo-embolic events in major organ systems as demonstrated by histology. Thus, subcutaneous injection of tTF-NGR represents an active, but toxic application procedure and compares unfavourably to intravenous infusion.
Insights
The fusion protein tTF-NGR targets tumor vasculature, inhibiting tumor growth and blood flow in mice. However, subcutaneous administration causes significant toxicity, including bleeding and organ damage, limiting its therapeutic potential.
Area of Science:
- Biotechnology
- Oncology
- Vascular Biology
Background:
- Truncated tissue factor (tTF) fused with the NGR peptide targets aminopeptidase N (CD13) and integrin αvβ3, both upregulated on tumor endothelial cells.
- These targets are crucial for tumor angiogenesis and vascular integrity.
Purpose of the Study:
- To evaluate the anti-tumor efficacy and toxicity of the fusion protein tTF-NGR.
- To investigate the mechanism of action and potential therapeutic window of tTF-NGR.
Main Methods:
- In vitro flow cytometry to assess binding to endothelial cells.
- In vivo studies in athymic mice bearing HT1080 fibrosarcoma xenografts.
- Contrast-enhanced ultrasound for tumor blood flow assessment.
- Histological analysis for vascular disruption and toxicity evaluation.
Main Results:
- tTF-NGR demonstrated specific binding to endothelial cells.
- Subcutaneous injection of tTF-NGR significantly inhibited tumor growth and reduced tumor blood flow.
- Histological analysis revealed vascular disruption, blood pooling, and thrombotic occlusion in tumors.
- Significant toxicities, including skin bleeding, reduced performance, and thrombo-embolic events, were observed at therapeutic doses.
- Lethality occurred within the therapeutic dose range, indicating a narrow or absent safe therapeutic window.
Conclusions:
- tTF-NGR exhibits potent anti-tumor activity by targeting tumor vasculature.
- Subcutaneous administration of tTF-NGR is associated with severe dose-limiting toxicities, precluding its safe clinical use via this route.
- Intravenous infusion may represent a more favorable administration route, warranting further investigation.
