Generation and characterization of an anti-delta like ligand-4 Nanobody to induce non-productive angiogenesis
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Monitoring Functionality and Morphology of Vasculature Recruited by Factors Secreted by Fast-growing Tumor-generating Cells
Published on: November 23, 2014
Rasoul Baharlou1, Nader Tajik2, Mahdi Habibi-Anbouhi3
1Department of Immunology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran.
Abstract:
Antibody-based targeting of angiogenesis is a key approach for cancer treatment. Delta-like ligand 4 (DLL4) plays a pivotal role in tumor neovascular development and angiogenesis during tumor progression. It forecasts the prognosis of human malignancies and blocking its signaling can help to inhibit neovascularization and tumor metastasis. Nanobodies are the smallest antigen-binding domains of heavy chain antibodies in camelidae. The aim of this study was to develop a Nanobody against DLL4 and apply binding and functional approaches to target it. In this work, a Nanobody library against human recombinant DLL4 was developed. After panning, the periplasmic-extract (PE) of individual colonies were screened through ELISA. The interactions between Nanobody and DLL4 were assessed using immunohistochemistry and FACS. The functional assessment was carried out via tube formation assay. We selected a Nanobody (3Nb3) with a high binding signal to DLL4, associated with a binding affinity of 3.6 nM. It was demonstrated that 3Nb3 binds to native DLL4 on the surface of MKN cells and gastric carcinoma tissue, and also inhibits the maturation of capillary-like structures in HUVECs. The results were indicative of the potential of Nanobody for DLL4 identification and can broaden the scope for development of cancer diagnosis and treatment techniques.
Insights
Researchers developed a novel Nanobody targeting Delta-like ligand 4 (DLL4) for cancer therapy. This Nanobody effectively binds to DLL4 and inhibits tumor angiogenesis, showing potential for cancer diagnosis and treatment.
Area of Science:
- Oncology
- Immunology
- Biotechnology
Background:
- Antibody-based therapies targeting angiogenesis are crucial for cancer treatment.
- Delta-like ligand 4 (DLL4) is a key regulator of tumor neovascularization and a prognostic marker in malignancies.
- Blocking DLL4 signaling can inhibit tumor growth, metastasis, and angiogenesis.
Purpose of the Study:
- To develop a Nanobody targeting human Delta-like ligand 4 (DLL4).
- To assess the binding and functional capabilities of the developed Nanobody against DLL4.
- To explore the potential of Nanobodies for cancer diagnosis and treatment by targeting DLL4.
Main Methods:
- Development of a Nanobody library against human recombinant DLL4.
- Screening of Nanobodies using ELISA, immunohistochemistry, and Flow Cytometry (FACS).
- Functional assessment via tube formation assay in human umbilical vein endothelial cells (HUVECs).
Main Results:
- A high-affinity Nanobody (3Nb3) with a binding affinity of 3.6 nM against DLL4 was selected.
- 3Nb3 demonstrated binding to native DLL4 on MKN cells and gastric carcinoma tissue.
- The Nanobody inhibited capillary-like structure formation in HUVECs, indicating anti-angiogenic activity.
Conclusions:
- The developed Nanobody (3Nb3) is a promising tool for identifying DLL4.
- This Nanobody shows potential for developing novel cancer diagnostic and therapeutic strategies.
- Nanobody technology offers a new avenue for targeting DLL4 in cancer treatment.
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