T cells redirected to interleukin-13Rα2 with interleukin-13 mutein--chimeric antigen receptors have anti-glioma activity but also recognize interleukin-13Rα1.
OBJECTIVE
Outcomes for patients with glioblastoma remain poor despite aggressive multimodal therapy. Immunotherapy with genetically modified T cells expressing chimeric antigen receptors (CARs) targeting interleukin (IL) 13Rα2, human epidermal growth factor receptor 2, epidermal growth factor variant III or erythropoietin-producing hepatocellular carcinoma A2 has shown promise for the treatment of glioma in preclinical models. On the basis of IL13Rα2 immunotoxins that contain IL13 molecules with one or two amino acid substitutions (IL13 muteins) to confer specificity to IL13Rα2, investigators have constructed CARS with IL13 muteins as antigen-binding domains. Whereas the specificity of IL13 muteins in the context of immunotoxins is well characterized, limited information is available for CAR T cells.


METHODS
We constructed four second-generation CARs with IL13 muteins with one or two amino acid substitutions, and evaluated the effector function of IL13-mutein CAR T cells in vitro and in vivo.


RESULTS
T cells expressing all four CARs recognized IL13Rα1 or IL13Rα2 recombinant protein in contrast to control protein (IL4R) as judged by interferon-γ production. IL13 protein produced significantly more IL2, indicating that IL13 mutein-CAR T cells have a higher affinity to IL13Rα2 than to IL13Rα1. In cytotoxicity assays, CAR T cells killed IL13Rα1- and/or IL13Rα2-positive cells in contrast to IL13Rα1- and IL13Rα2-negative controls. Although we observed no significant differences between IL13 mutein-CAR T cells in vitro, only T cells expressing IL13 mutein-CARs with an E13K amino acid substitution had anti-tumor activity in vivo that resulted in a survival advantage of treated animals.


CONCLUSIONS
Our study highlights that the specificity/avidity of ligands is context-dependent and that evaluating CAR T cells in preclinical animal model is critical to assess their potential benefit.
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA.
Simone
S
Krebs
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA.
Kevin K H
KK
Chow
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA.
Zhongzhen
Z
Yi
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA.
Tania
T
Rodriguez-Cruz
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA; Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA.
Meenakshi
M
Hegde
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA.
Claudia
C
Gerken
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA; Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA; Department of Pediatrics, Baylor College of Medicine, Houston, Texas, USA.
Nabil
N
Ahmed
Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, Texas, USA; Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA; Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA; Department of Pediatrics, Baylor College of Medicine, Houston, Texas, USA; Department of Pathology and Immunology, Baylor College of Medicine, Houston, Texas, USA. Electronic address: smgottsc@txch.org.
Stephen
S
Gottschalk
D019943:Amino Acid Substitution; D000818:Animals; D015972:Gene Expression Regulation, Neoplastic / Q000276:immunology; D005909:Glioblastoma / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D006801:Humans; D007167:Immunotherapy; D000922:Immunotoxins / Q000235:genetics; D053661:Interleukin-13 Receptor alpha1 Subunit / Q000235:genetics / Q000276:immunology; D053663:Interleukin-13 Receptor alpha2 Subunit / Q000235:genetics* / Q000276:immunology; D051379:Mice; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D023041:Xenograft Model Antitumor Assays
D000922:Immunotoxins; D053661:Interleukin-13 Receptor alpha1 Subunit; D053663:Interleukin-13 Receptor alpha2 Subunit; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins
CAR; GBM; IL13Rα1; IL13Rα2; T cells; cancer; immunotherapy