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Combinatorial T cell engineering eliminates on-target off-tumor toxicity of CD229 CAR T cells while maintaining anti-tumor activity

Erica R. Vander Mause, Jillian M. Baker, Kenneth A. Dietze, Sabarinath V. Radhakrishnan, Thierry Iraguha, Patricia Davis, Jens Panse, James E. Marvin, Michael L. Olson, Mary Steinbach, David P. Ng, Carol S. Lim, Djordje Atanackovic, Tim Luetkens
doi: https://doi.org/10.1101/2021.12.06.471279
Erica R. Vander Mause
1Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD, United States
2Hematology and Hematologic Malignancies, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, United States
3Department of Pharmaceutics & Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT, United States
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Jillian M. Baker
1Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD, United States
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Kenneth A. Dietze
1Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD, United States
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Sabarinath V. Radhakrishnan
2Hematology and Hematologic Malignancies, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, United States
4Division of Hematology and Oncology, Medical College of Wisconsin, Milwaukee, USA
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Thierry Iraguha
9Department of Medicine and Transplant/Cell Therapy Program, University of Maryland School of Medicine and Marlene and Stewart Greenebaum Comprehensive Cancer Center, Baltimore, MD, United States
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Patricia Davis
5Division of Experimental and Clinical Pathology, ARUP Laboratories, Salt Lake City, USA
6Department of Pathology, University of Utah, Salt Lake City, UT, United States
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Jens Panse
7Department of Oncology, Hematology, Hemostaseology, and Stem Cell Transplantation, University Hospital RWTH Aachen, Aachen, Germany
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James E. Marvin
8Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, United States
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Michael L. Olson
6Department of Pathology, University of Utah, Salt Lake City, UT, United States
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Mary Steinbach
2Hematology and Hematologic Malignancies, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, United States
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David P. Ng
6Department of Pathology, University of Utah, Salt Lake City, UT, United States
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Carol S. Lim
3Department of Pharmaceutics & Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT, United States
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Djordje Atanackovic
1Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD, United States
9Department of Medicine and Transplant/Cell Therapy Program, University of Maryland School of Medicine and Marlene and Stewart Greenebaum Comprehensive Cancer Center, Baltimore, MD, United States
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Tim Luetkens
1Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD, United States
2Hematology and Hematologic Malignancies, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, United States
9Department of Medicine and Transplant/Cell Therapy Program, University of Maryland School of Medicine and Marlene and Stewart Greenebaum Comprehensive Cancer Center, Baltimore, MD, United States
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  • For correspondence: tluetkens@som.umaryland.edu
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ABSTRACT

T cells expressing chimeric antigen receptors have shown remarkable therapeutic activity against different types of cancer. However, their wider use has been hampered by the potential for life-threatening toxicities due to the unintended targeting of healthy cells expressing low levels of the targeted antigen. We have now developed an affinity-tuning approach for the generation of minimally modified, low-affinity antibody variants derived from existing high-affinity antibodies. Using this approach, we engineered low affinity variants of the fully human CD229-specific antibody 2D3. Parental 2D3 originally efficiently targeted multiple myeloma cells but also healthy T cells expressing low levels of CD229. We demonstrate that CAR T cells based on a low affinity variant of 2D3, engineered to also express CJUN to increase CAR T cell expansion, maintain the parental antibody’s anti-tumor activity but lack its targeting of healthy T cells in vitro and in vivo. In addition, we found that low affinity CD229 CAR T cells show reduced trogocytosis potentially augmenting CAR T cell persistence. The fast off-rate CAR produced using our affinity tuning approach eliminates a key liability of CD229 CAR T cells and paves the way for the effective and safe treatment of patients with multiple myeloma and other lymphoid malignancies.

One sentence summary Rational T cell engineering yields low affinity CD229 CAR T cells overexpressing CJUN, which maintain the parental cells’ anti-tumor activity but eliminate killing of healthy T cells, increasing CAR T cell expansion, and decreasing trogocytosis.

Competing Interest Statement

ERV, TL, and DA are inventors on provisional patent application 63/285843 describing low-affinity CD229 antibodies and CAR T cells. SVR, DA, and TL are inventors on PCT application US2017/42840 'Antibodies and CAR T Cells for the Treatment of Multiple Myeloma' describing the therapeutic use of CD229 CAR T cells for the treatment of multiple myeloma.

Footnotes

  • Added in vivo data. Added optimization of low affinity CAR construct using CJUN.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted June 19, 2022.
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Combinatorial T cell engineering eliminates on-target off-tumor toxicity of CD229 CAR T cells while maintaining anti-tumor activity
Erica R. Vander Mause, Jillian M. Baker, Kenneth A. Dietze, Sabarinath V. Radhakrishnan, Thierry Iraguha, Patricia Davis, Jens Panse, James E. Marvin, Michael L. Olson, Mary Steinbach, David P. Ng, Carol S. Lim, Djordje Atanackovic, Tim Luetkens
bioRxiv 2021.12.06.471279; doi: https://doi.org/10.1101/2021.12.06.471279
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Combinatorial T cell engineering eliminates on-target off-tumor toxicity of CD229 CAR T cells while maintaining anti-tumor activity
Erica R. Vander Mause, Jillian M. Baker, Kenneth A. Dietze, Sabarinath V. Radhakrishnan, Thierry Iraguha, Patricia Davis, Jens Panse, James E. Marvin, Michael L. Olson, Mary Steinbach, David P. Ng, Carol S. Lim, Djordje Atanackovic, Tim Luetkens
bioRxiv 2021.12.06.471279; doi: https://doi.org/10.1101/2021.12.06.471279

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