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De novo identification of tumor-specific internalizing human antibody-receptor pairs by phage-display methods
Changshou Gao1, Shenlan Mao, Francesca Ronca
1Department of Chemistry, The Scripps Research Institute and the Skaggs Institute for Chemical Biology, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Abstract:
Three tumor-specific, internalizing human single-chain Fvs (scFvs) were obtained by direct selection against tumor cells from a large, nonimmune scFv-phage library pre-subtracted with various normal human cells. After scFv selection and characterization for cell binding and internalization, the scFvs were also employed in immunoprecipitations to identify putative receptors. In the case of a prostate tumor-cell specific scFv PR5, the receptor that mediated endocytosis was shown to be the transferrin receptor. For two pancreatic adenocarcinoma specific scFvs SW1 and PAN10, the alpha(3)beta(1) integrin was identified. The scFv SW1 was studied in further detail and found to induce functional effects as a ligand-mimetic by mediating cell adhesion and migration. The results demonstrated the feasibility of utilizing enhanced phage-display methods as a rapid and general approach for not only direct isolation of human internalizing scFvs, but also for identifying tumor cell-surface receptors from various classes. The use of scFv constructs that target tumor cells and undergo internalization could have significant impact on the future of cancer and gene therapy.
Insights
Researchers developed a rapid phage-display method to isolate tumor-targeting single-chain Fvs (scFvs) and identify their receptors. This approach successfully identified the transferrin receptor and alpha(3)beta(1) integrin, paving the way for cancer and gene therapy advancements.
Area of Science:
- Biotechnology
- Molecular Biology
- Cancer Research
Background:
- Targeting tumor-specific antigens is crucial for effective cancer therapy.
- Phage-display technology offers a powerful platform for antibody fragment discovery.
- Identifying internalized single-chain Fvs (scFvs) and their receptors can enhance therapeutic strategies.
Purpose of the Study:
- To develop and validate an enhanced phage-display method for isolating tumor-specific, internalizing human scFvs.
- To identify the specific cell-surface receptors that mediate the internalization of these scFvs.
- To explore the potential therapeutic applications of these scFvs in cancer and gene therapy.
Main Methods:
- Utilized a large, nonimmune scFv-phage library pre-subtracted with normal human cells for direct selection against tumor cells.
- Characterized selected scFvs for tumor cell binding and internalization capabilities.
- Employed immunoprecipitation assays to identify putative receptor targets for the scFvs.
Main Results:
- Successfully isolated three tumor-specific, internalizing human scFvs.
- Identified the transferrin receptor as the mediator for prostate tumor-specific scFv PR5 internalization.
- Identified the alpha(3)beta(1) integrin as the receptor for pancreatic adenocarcinoma-specific scFvs SW1 and PAN10.
- Demonstrated that scFv SW1 acts as a ligand-mimetic, influencing cell adhesion and migration.
Conclusions:
- Enhanced phage-display is a rapid and versatile method for isolating internalizing scFvs and identifying tumor cell-surface receptors.
- The identified scFvs and their receptors provide novel targets for cancer therapy.
- Internalizing scFv constructs hold significant promise for future cancer and gene therapy applications.