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Published on: November 12, 2019
Designed Ankyrin Repeat Proteins as Her2 Targeting Domains in Chimeric Antigen Receptor-Engineered T Cells
Elizabeth Siegler1, Si Li2, Yu Jeong Kim2
11 Department of Biomedical Engineering, University of Southern California , Los Angeles, California.
Abstract:
Chimeric antigen receptor (CAR) engineering is a branch of cancer immunotherapy that equips immune cells to target tumor antigens expressed on the cell surface using antibody-derived single-chain variable fragments (scFvs). However, other antibody mimetics, such as designed ankyrin repeat proteins (DARPins), can also serve as antigen-binding domains in CARs. This study shows that CAR-engineered T (CAR-T) cells utilizing Her2-targeting DARPins G3 and 929 can target human epidermal growth factor receptor 2 (Her2)-overexpressing cancer cells as effectively as CAR-T cells with the scFv 4D5 in vitro, and G3 CAR-T cells can slow or eliminate tumor growth in vivo as effectively as 4D5 CAR-T cells. Some DARPins may offer an attractive alternative to scFv usage in CARs, as they are smaller, thermodynamically stable, poorly immunogenic, and can be generated with different binding properties from DARPin libraries.
Insights
Designed ankyrin repeat proteins (DARPins) show promise as alternatives to antibody fragments in CAR-T cell therapy. DARPins effectively target HER2-cancer cells, matching traditional CAR-T cell efficacy in vitro and in vivo.
Area of Science:
- Oncology
- Immunology
- Biotechnology
Background:
- Chimeric antigen receptor (CAR) engineering is a key cancer immunotherapy strategy.
- CARs typically use single-chain variable fragments (scFvs) derived from antibodies to target tumor antigens.
- Alternative antigen-binding domains are being explored to enhance CAR functionality.
Purpose of the Study:
- To evaluate the efficacy of designed ankyrin repeat proteins (DARPins) as antigen-binding domains in CAR-engineered T (CAR-T) cells.
- To compare the performance of DARPin-based CAR-T cells with traditional scFv-based CAR-T cells against HER2-overexpressing cancers.
Main Methods:
- Engineering CAR-T cells utilizing Her2-targeting DARPins (G3 and 929).
- In vitro assays to assess the targeting of human epidermal growth factor receptor 2 (Her2)-overexpressing cancer cells.
- In vivo studies to evaluate tumor growth inhibition by G3 CAR-T cells compared to 4D5 scFv CAR-T cells.
Main Results:
- CAR-T cells with Her2-targeting DARPins (G3 and 929) demonstrated comparable efficacy to scFv 4D5 CAR-T cells in vitro.
- G3 CAR-T cells effectively slowed or eliminated tumor growth in vivo, similar to 4D5 CAR-T cells.
- DARPin-based CARs showed potential for targeting Her2-positive cancers.
Conclusions:
- Designed ankyrin repeat proteins (DARPins) are viable alternatives to scFvs for CAR engineering.
- DARPins offer advantages such as smaller size, enhanced stability, and reduced immunogenicity.
- DARPin-based CAR-T cells present a promising therapeutic strategy for Her2-overexpressing cancers.
