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ROR1-targeting switchable CAR-T cells for cancer therapy

Oncogene volume 41, pages 4104–4114 (2022)Cite this article

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A Correction to this article was published on 21 February 2024

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Abstract

The success of chimeric antigen receptor T cell (CAR-T) therapy in the treatment of hematologic malignancies has prompted the development of numerous CAR-T technologies, including switchable CAR-T (sCAR-T) systems that combine a universal CAR-T with bispecific adapter proteins. Owing to their controllability and versatility, sCAR-Ts have received considerable attention. To explore the therapeutic utility of sCAR-Ts targeting the receptor tyrosine kinase ROR1, which is expressed in hematologic and solid malignancies, and to identify bispecific adaptor proteins that efficiently mediate universal CAR-T engagement, a panel of switches based on ROR1-targeting Fabs with different epitopes and affinities was compared in in vitro and in vivo models of ROR1-expressing cancers. For switches targeting overlapping or identical epitopes, potency correlated with affinity. Surprisingly, however, we identified a switch targeting a unique epitope with low affinity but mediating potent and selective antitumor activity in vitro and in vivo. Converted to a conventional CAR-T, the same anti-ROR1 mAb (324) outperformed a clinically investigated conventional CAR-T that is based on an anti-ROR1 mAb (R12) with ~200-fold higher affinity. Thus, demonstrating therapeutic utility on their own, sCAR-Ts also facilitate higher throughput screening for the identification of conventional CAR-T candidates for preclinical and clinical studies.

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Fig. 1: A ROR1-targeting sCAR-T system.
Fig. 2: Orthogonal binding of anti-ROR1 Fab switches to effector cells and target cells.
Fig. 3: Switch-mediated in vitro cytotoxicity of sCAR-Ts.
Fig. 4: Switch-mediated in vitro activation of sCAR-Ts.
Fig. 5: Switch-mediated in vitro cytotoxicity and activation of sCAR-Ts against TNBC cells.
Fig. 6: Switch-mediated in vivo cytotoxicity of sCAR-Ts in an NSG/JeKo-1_ffluc xenograft mouse model.
Fig. 7: In vitro and in vivo activity of sCAR-Ts targeting colon cancer cells.

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Acknowledgements

CR acknowledges support by National Institutes of Health (NIH) grants R01 CA174844, R01 CA181258, R01 CA204484, R21 CA229961, and R21 CA263240, and by the Klorfine Foundation.

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Authors and Affiliations

  1. Department of Immunology and Microbiology, UF Scripps Biomedical Research, University of Florida, Jupiter, FL, 33458, USA

    Haiyong Peng, Jing Chang & Christoph Rader

  2. Medizinische Klinik und Poliklinik II, Universitätsklinikum Würzburg, Oberdürrbacher Strasse 6, 97080, Würzburg, Germany

    Thomas Nerreter, Katrin Mestermann, Jakob Wachter & Michael Hudecek

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Contributions

HP and CR conceived and designed the research. HP conducted the in vitro and in vivo experiments with switches and sCAR-Ts. Supervised by MH, KM and JW selected affinity variants. JC assisted with protein production. TN and MH provided lentiviral vectors and advice for generating CAR-Ts. HP, TN, and CR wrote the manuscript. All authors read, edited, and approved the manuscript.

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Correspondence to Haiyong Peng or Christoph Rader.

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Competing interests

CR and HP are named inventors on a licensed patent family (assignee, University of Florida and Boehringer Ingelheim) that claims a set of anti-ROR1 mAbs used in this study including 324 (United States Patent 10,618,959). CR is named inventor on a licensed patent family (assignee, United States of America) that claims another set of anti-ROR1 mAbs used in this study including R12 (United States Patent 9,758,586).

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Peng, H., Nerreter, T., Mestermann, K. et al. ROR1-targeting switchable CAR-T cells for cancer therapy. Oncogene 41, 4104–4114 (2022). https://doi.org/10.1038/s41388-022-02416-5

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