From healthy donors, Ruggiero et al. isolated T cells expressing 19 TCRαβs targeting WT1, a transcription factor overexpressed in multiple malignancies. Five high-avidity TCRs restricted by HLA-A02:01, specific to naturally processed immunodominant or noncanonical WT1 epitopes, and which recognized primary AML blasts, were identified. CRISPR-Cas9 genome-edited T cells, generated by targeted integration into the TCRα constant locus combined with deletion of endogenous TCRαβ genes, were enriched in memory stem T cells, which killed primary leukemias
and glioblastoma cells without off-target toxicity in mouse models.
Contributed by Paula Hochman
ABSTRACT: T cell receptor (TCR)-based therapy has the potential to induce durable clinical responses in patients with cancer by targeting intracellular tumor antigens with high sensitivity and by promoting T cell survival. However, the need for TCRs specific for shared oncogenic antigens and the need for manufacturing protocols able to redirect T cell specificity while preserving T cell fitness remain limiting factors. By longitudinal monitoring of T cell functionality and dynamics in 15 healthy donors, we isolated 19 TCRs specific for Wilms' tumor antigen 1 (WT1), which is overexpressed by several tumor types. TCRs recognized several peptides restricted by common human leukocyte antigen (HLA) alleles and displayed a wide range of functional avidities. We selected five high-avidity HLA-A*02:01-restricted TCRs, three that were specific to the less explored immunodominant WT137-45 and two that were specific to the noncanonical WT1-78-64 epitopes, both naturally processed by primary acute myeloid leukemia (AML) blasts. With CRISPR-Cas9 genome editing tools, we combined TCR-targeted integration into the TCR α constant (TRAC) locus with TCR β constant (TRBC) knockout, thus avoiding TCRαβ mispairing and maximizing TCR expression and function. The engineered lymphocytes were enriched in memory stem T cells. A unique WT137-45-specific TCR showed antigen-specific responses and efficiently killed AML blasts, acute lymphoblastic leukemia blasts, and glioblastoma cells in vitro and in vivo in the absence of off-tumor toxicity. T cells engineered to express this receptor are being advanced into clinical development for AML immunotherapy and represent a candidate therapy for other WT1-expressing tumors.
Author Info: (1) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (2) Experimental
Author Info: (1) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (2) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (3) Intellia Therapeutics, Cambridge, MA 02139, USA. (4) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (5) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (6) San Raffaele Telethon Institute for Gene Therapy (SR-TIGET), Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. National Research Council, Institute for Biomedical Technologies, Segrate, Italy. (7) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (8) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (9) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. School of Medicine and Surgery, Milano-Bicocca University, 20126 Milan, Italy. (10) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (11) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. Vita-Salute San Raffaele University, 20132 Milan, Italy. (12) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (13) Immunogenetics, Leukemia Genomics and Immunobiology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. Hematology and Bone Marrow Transplantation Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (14) Immunohematology and Transfusion Medicine Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (15) Hematology and Bone Marrow Transplantation Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (16) Vita-Salute San Raffaele University, 20132 Milan, Italy. Pathology Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (17) Pathology Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (18) Intellia Therapeutics, Cambridge, MA 02139, USA. (19) Intellia Therapeutics, Cambridge, MA 02139, USA. (20) Neural Stem Cell Biology Unit, Division of Neurosciences, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (21) Genomics of the Innate Immune System Unit, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (22) Vita-Salute San Raffaele University, 20132 Milan, Italy. Genomics of the Innate Immune System Unit, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (23) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (24) Intellia Therapeutics, Cambridge, MA 02139, USA. (25) Intellia Therapeutics, Cambridge, MA 02139, USA. (26) Intellia Therapeutics, Cambridge, MA 02139, USA. (27) Intellia Therapeutics, Cambridge, MA 02139, USA. (28) Intellia Therapeutics, Cambridge, MA 02139, USA. (29) Intellia Therapeutics, Cambridge, MA 02139, USA. (30) Vita-Salute San Raffaele University, 20132 Milan, Italy. Hematology and Bone Marrow Transplantation Unit, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. (31) Experimental Hematology Unit, Division of Immunology, Transplantation and Infectious Diseases, Ospedale San Raffaele Scientific Institute, 20132 Milan, Italy. Vita-Salute San Raffaele University, 20132 Milan, Italy.
