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Development of Human CBF1-Targeting Single-Stranded DNA Aptamers with Antiangiogenic Activity In Vitro
Mari Tezuka-Kagajo1,2, Masashi Maekawa1,3, Atsushi Ogawa4
1Department of Biochemistry and Molecular Genetics and Ehime University Graduate School of Medicine, Toon, Japan.
Abstract:
C promoter binding factor 1 (CBF1) (alias RBPJ) is a critical transcription factor involved in Notch signaling. The activation of Notch signaling through CBF1 maintains the angiostatic state of endothelial cells suppressing angiogenesis, that is, the formation of new blood vessels. Vascular endothelial growth factor (VEGF) induces angiogenesis by promoting the proteasomal degradation of CBF1, in addition to endothelial cell proliferation. To date, angiogenic inhibitors targeting VEGF have been successfully used in clinics for cancer and age-related macular degeneration. Most antiangiogenic drugs, however, only target VEGF or VEGF receptors. In this study, to expand the repertoire of antiangiogenic therapeutics, we developed 15 single-stranded deoxyribonucleic acid (ssDNA) aptamers capable of binding to CBF1 with high affinity (Kd; 10-300 nM). To this end, systematic evolution of ligands by the exponential enrichment (SELEX) method was applied. One of the CBF1-binding ssDNA aptamers, Apt-3, inhibited angiogenesis through the activation of Notch signaling in vitro. We found that Apt-3 directly interacted with the LAG1 domain of CBF1. We suggest that the Apt-3 ssDNA aptamer may contribute to the development of a novel angiogenic inhibitor, which does not target VEGF.
Insights
Researchers developed novel DNA aptamers that inhibit blood vessel formation by targeting the transcription factor CBF1, offering a new therapeutic strategy beyond VEGF inhibitors for conditions like cancer.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- CBF1 (RBPJ) is a transcription factor crucial for Notch signaling, maintaining endothelial cell quiescence and suppressing angiogenesis.
- VEGF promotes angiogenesis by degrading CBF1, and current anti-angiogenic therapies primarily target VEGF or its receptors.
- There is a need for novel anti-angiogenic therapeutics with different mechanisms of action.
Purpose of the Study:
- To develop novel single-stranded DNA (ssDNA) aptamers targeting CBF1 for anti-angiogenic therapy.
- To characterize the binding affinity and inhibitory potential of these aptamers.
- To explore a new therapeutic avenue for angiogenesis-related diseases by targeting CBF1.
Main Methods:
- Systematic evolution of ligands by exponential enrichment (SELEX) was employed to generate ssDNA aptamers against CBF1.
- Binding affinity of aptamers to CBF1 was determined (Kd; 10-300 nM).
- In vitro assays were used to assess the anti-angiogenic activity of the aptamers and their interaction with CBF1 domains.
Main Results:
- Fifteen ssDNA aptamers with high affinity for CBF1 were successfully developed.
- One aptamer, Apt-3, demonstrated significant inhibition of angiogenesis by activating Notch signaling.
- Apt-3 was found to directly bind to the LAG1 domain of CBF1.
Conclusions:
- The ssDNA aptamer Apt-3 shows potential as a novel anti-angiogenic therapeutic agent.
- Targeting CBF1 offers a new strategy for developing anti-angiogenic drugs independent of VEGF.
- This approach could lead to new treatments for angiogenesis-driven diseases such as cancer and AMD.
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