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Cell Therapy and Immunotherapy
Genome editing of donor-derived T cells
to generate allogeneic chimeric antigen receptor-modified T cells: optimizing aβ T-cell-depleted haploidentical hematopoietic stem cell transplantation
Ferrata Storti Foundation
Haematologica 2021 Volume 106(3):847-858
Volker Wiebking,1 Ciaran M. Lee,2 Nathalie Mostrel,1 Premanjali Lahiri,3 Rasmus Bak,4,5 Gang Bao,2 Maria Grazia Roncarolo,1,6 Alice Bertaina1# and Matthew H. Porteus1,6#
1Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA; 2Department of Bioengineering, Rice University, Houston, TX, USA; 3Laboratory for Cell and Gene Medicine, Stanford University School of Medicine, Stanford, CA, USA; 4Department of Biomedicine, Aarhus University, Aarhus, Denmark; 5Aarhus Institute of Advanced Studies (AIAS), Aarhus University, Aarhus, Denmark and 6Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA.
#AB and MHP contributed equally as co-senior authors.
ABSTRACT
Allogeneic hematopoietic stem cell transplantation is an effective therapy for high-risk leukemias. In children, graft manipulation based on the selective removal of aβ T cells and B cells has been shown to reduce the risk of acute and chronic graft-versus-host disease, thus allowing the use of haploidentical donors which expands the popu- lation of recipients in whom allogeneic hematopoietic stem cell trans- plantation can be used. Leukemic relapse, however, remains a challenge. T cells expressing chimeric antigen receptors can potently eliminate leukemia, including those in the central nervous system. We hypothe- sized that by engineering the donor aβ T cells that are removed from the graft by genome editing to express a CD19-specific chimeric antigen receptor, while simultaneously inactivating the T-cell receptor, we could create a therapy that enhances the anti-leukemic efficacy of the stem cell transplant without increasing the risk of graft-versus-host disease. Using genome editing with Cas9 ribonucleoprotein and adeno-associated virus serotype 6, we integrated a CD19-specific chimeric antigen receptor in- frame into the TRAC locus. More than 90% of cells lost T-cell receptor expression, while >75% expressed the chimeric antigen receptor. The initial product was further purified with less than 0.05% T-cell receptor- positive cells remaining. In vitro, the chimeric antigen receptor T cells effi- ciently eliminated target cells and produced high cytokine levels when challenged with CD19+ leukemia cells. In vivo, the gene-modified T cells eliminated leukemia without causing graft-versus-host disease in a xenograft model. Gene editing was highly specific with no evidence of off-target effects. These data support the concept that the addition of aβ T-cell-derived, genome-edited T cells expressing CD19-specific chimeric antigen receptors could enhance the anti-leukemic efficacy of aβ T-cell- depleted haploidentical hematopoietic stem cell transplantation without increasing the risk of graft-versus-host disease.
Introduction
Despite improvements over the last decades, treatment outcomes remain unfa- vorable in pediatric patients with relapsed or refractory B-cell precursor acute lym- phoblastic leukemia.1,2 Allogeneic hematopoietic stem cell transplantation (HSCT) has been successfully employed to treat high-risk leukemias,3 providing a graft-ver- sus-leukemia4 effect but also carrying the risk of graft-versus-host disease (GvHD).5
Correspondence:
MATTHEW H. PORTEUS
mporteus@stanford.edu
ALICE BERTAINA
aliceb1@stanford.edu
Received: August 15, 2019. Accepted: March 19, 2020. Pre-published: April 2, 2020.
https://doi.org/10.3324/haematol.2019.233882
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