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Published on: February 17, 2022
CAR-T Therapy Targets Extra Domain B of Fibronectin Positive Solid Tumor Cells
Jie Tang1, Nan Liu1, Yongjie Zhu1
1Department of Targeting Therapy & Immunology and Laboratory of Animal Tumor Models, Cancer Center and Department of Respiratory and Critical care Medicine and Frontiers Science Center for Disease-related Molecular Network, West China Hospital, Sichuan University, Chengdu, Sichuan, China.
Background:
CAR-T cell immunotherapy has achieved remarkable success in malignant B-cell malignancies, but progress in solid tumors is slow, and one of the key reasons is the lack of ideal targets. Cancer-specific extra domain B of fibronectin (EDB-FN) is widely upregulated in solid tumors and expressed at low levels in normal tissues. Many imaging and targeted cancer therapies based on EDB-FN targets have been developed and tested in clinical trials, making EDB-FN an ideal target for immunotherapy.
Methods:
We constructed two EDB-FN-targeted CAR-Ts based on the peptide APT0 and the single-chain antibody CGS2 in a lentiviral infection manner for the first time. Luciferase cytotoxicity assay to assess CAR-T killing of tumor cells. An enzyme-linked immunosorbent assay was used to detect the release of the cytokine IFN-γ. Fluorescence imaging to evaluate the dynamics of CAR-T cell and tumor cell coculture. Knockdown assays were used to validate the target specificity of CAR-T cells.
Results:
In this research, two CAR-Ts targeting EDB-FN, APT0 CAR-T, and CGS2 CAR-T, were constructed. In vitro, both CAR-T cells produced broad-spectrum killing of multiple EDB-FN-positive solid tumor cell lines and were accompanied by cytokine IFN-γ release. Regarding safety, the two CAR-T cells did not affect T cells' normal growth and proliferation and were not toxic to HEK-293T human embryonic kidney epithelial cells.
Conclusion:
APT0 CAR-T and CGS2 CAR-T cells are two new CAR-Ts targeting EDB-FN. Both CAR-T cells can successfully identify and specifically kill various EDB-FN-positive solid tumor cells with potential clinical applications.
Insights
Researchers developed novel Chimeric Antigen Receptor T-cell (CAR-T) therapies targeting extra domain B of fibronectin (EDB-FN) for solid tumors. These EDB-FN-targeted CAR-T cells demonstrated effective tumor cell killing and hold potential for clinical application.
Area of Science:
- Immunotherapy
- Oncology
- Biotechnology
Background:
- CAR-T cell therapy shows success in B-cell malignancies but lags in solid tumors due to target limitations.
- Extra domain B of fibronectin (EDB-FN) is a promising cancer-specific target, upregulated in solid tumors and minimally expressed in normal tissues.
- EDB-FN's characteristics make it an ideal target for developing novel immunotherapies.
Purpose of the Study:
- To engineer and evaluate novel Chimeric Antigen Receptor T-cells (CAR-Ts) targeting the cancer-specific antigen EDB-FN.
- To assess the efficacy and specificity of EDB-FN-targeted CAR-Ts against solid tumor cell lines.
- To investigate the safety profile of these novel CAR-T constructs.
Main Methods:
- Construction of two EDB-FN-targeted CAR-Ts using a peptide (APT0) and a single-chain antibody (CGS2) via lentiviral transduction.
- In vitro assessment of CAR-T cytotoxicity using luciferase assays and cytokine release (IFN-γ) via ELISA.
- Evaluation of CAR-T cell dynamics and target specificity through fluorescence imaging and knockdown assays.
Main Results:
- Successfully constructed APT0 CAR-T and CGS2 CAR-T targeting EDB-FN.
- Both CAR-T variants exhibited broad-spectrum killing of EDB-FN-positive solid tumor cell lines, accompanied by IFN-γ release.
- CAR-T cells demonstrated no toxicity towards normal T-cells or HEK-293T cells, indicating a favorable safety profile.
Conclusions:
- APT0 CAR-T and CGS2 CAR-T represent novel immunotherapeutic agents targeting EDB-FN for solid tumors.
- These CAR-T cells effectively identify and eliminate various EDB-FN-positive solid tumor cells in vitro.
- The developed CAR-T therapies show significant potential for future clinical applications in solid tumor treatment.
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