PiggyBac-Engineered T Cells Expressing CD19-Specific CARs that Lack IgG1 Fc Spacers Have Potent Activity against B-ALL Xenografts

• Published in: Molecular therapy Vol. 26; no. 8; pp. 1883 - 1895
• Main Authors: Bishop, David C., Xu, Ning, Tse, Benjamin, O’Brien, Tracey A., Gottlieb, David J., Dolnikov, Alla, Micklethwaite, Kenneth P.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.08.2018; Elsevier Limited; American Society of Gene & Cell Therapy
• Subjects: acute lymphoblastic leukemia; Animals; B cell malignancy; Cancer; CAR T cell; CD19; CD19 antigen; CD28 antigen; CD4 antigen; Cell Line, Tumor; Cell lines; cellular immunotherapy; chimeric antigen receptor; Chimeric antigen receptors; Clinical trials; Cytokines; Cytotoxicity; DNA Transposable Elements; Flow cytometry; Funding; Humans; Immunoglobulin G; Immunotherapy, Adoptive - methods; Jurkat Cells; K562 Cells; Leukemia; Lymphocytes; Lymphocytes B; Lymphocytes T; Medical research; Mice; murine xenograft model; non-viral vector; Original; piggyBac transposase; Plasmids; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma - genetics; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma - immunology; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma - therapy; Receptors, Antigen, T-Cell - metabolism; Receptors, IgG - genetics; Spacer; Statistical analysis; T-Lymphocytes - transplantation; transposon; Treatment Outcome; Vectors (Biology); Xenograft Model Antitumor Assays; Xenografts; Australia; New South Wales Australia; transposon; piggyBac transposase; B cell malignancy; acute lymphoblastic leukemia; CD19; chimeric antigen receptor; cellular immunotherapy; murine xenograft model; non-viral vector; CAR T cell
• ISSN: 1525-0016; 1525-0024
• DOI: 10.1016/j.ymthe.2018.05.007

Clinical trials of CD19-specific chimeric antigen receptor (CAR19) T cells have demonstrated remarkable efficacy against relapsed and refractory B cell malignancies. The piggyBac transposon system offers a less complex and more economical means for generating CAR19 T cells compared to viral vectors. We have previously optimized a protocol for the generation of CAR19 T cells using the piggyBac system, but we found that CAR19 T cells had poor in vivo efficacy and persistence, probably due to deleterious FcγR interactions with the CAR’s IgG1 Fc-containing spacer domain. We therefore designed three CD19-specifc CARs that lacked the IgG1 Fc region, and we incorporated combinations of CD28 or 4-1BB transmembrane and co-stimulatory domains. PiggyBac-generated CAR19 T cells expressing these re-designed constructs all demonstrated reactivity in vitro specifically against CD19+ cell lines. However, those combining CD28 transmembrane and co-stimulatory domains showed CD4 predominance and inferior cytotoxicity. At high doses, CAR19 T cells were effective against B-ALL in a xenograft mouse model, regardless of co-stimulatory domain. At diminishing doses, 4-1BB co-stimulation led to greater potency and persistence of CAR19 T cells, and it provided protection against B-ALL re-challenge. Production of potent CAR T cells using piggyBac is simple and cost-effective, and it may enable wider access to CAR T cell therapy. CAR T cell production using viral vectors can be complex and expensive. Bishop and colleagues demonstrate that the piggyBac transposase is a valid alternative for generating CAR T cells that retain high in vivo efficacy. This simple and economical approach may facilitate greater access to CAR T cell therapy in the future.