Manufacture of T cells using the Sleeping Beauty system to enforce expression of a CD19-specific chimeric antigen receptor

Singh, Harjeet; Moyes, Judy S.; Huls, M. Helen; Cooper, Laurence J.N.

doi:10.1038/cgt.2014.69

Cited by 76 publications

(63 citation statements)

References 55 publications

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“…Release criteria to produce banks were as described (36)(37)(38). Details of the manufacturing and expansion process are provided in the Supplemental Methods.…”

Section: Multiparameter Flow Cytometrymentioning

confidence: 99%

“…To produce CAR T cells for infusion, peripheral blood (PB) mononuclear cells (PBMCs) simply obtained by venipuncture (Supplemental Table 1), were electroporated with SB transposon and transposase plasmids encoding CD19RCD28 and SB11. Electroporated cells were expanded on γ-irradiated clone 4 AaPCs for a median of 28 days (autologous: average 29 days, SD 1.4 days; allogeneic: average 28.5 days, SD 3.9 days) with IL-2 and IL-21, enabling selective growth of T cells stably encoding the CD19-specific CAR (Figure 2A), following methods reported previously (34)(35)(36)(37)(38). This process resulted in the selective outgrowth of T cells with integrated CAR (average cell expansion: ~2,200-fold autologous and ~2,500-fold allogeneic).…”

Section: Introductionmentioning

confidence: 99%

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Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells

Kebriaei¹,

Singh

Huls

et al. 2016

Journal of Clinical Investigation

437

390

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BACKGROUND.T cells expressing antigen-specific chimeric antigen receptors (CARs) improve outcomes for CD19-expressing B cell malignancies. We evaluated a human application of T cells that were genetically modified using the Sleeping Beauty (SB) transposon/transposase system to express a CD19-specific CAR. METHODS.T cells were genetically modified using DNA plasmids from the SB platform to stably express a second-generation CD19-specific CAR and selectively propagated ex vivo with activating and propagating cells (AaPCs) and cytokines. Twenty-six patients with advanced non-Hodgkin lymphoma and acute lymphoblastic leukemia safely underwent hematopoietic stem cell transplantation (HSCT) and infusion of CAR T cells as adjuvant therapy in the autologous (n = 7) or allogeneic settings (n = 19).RESULTS. SB-mediated genetic transposition and stimulation resulted in 2,200-to 2,500-fold ex vivo expansion of genetically modified T cells, with 84% CAR expression, and without integration hotspots. Following autologous HSCT, the 30-month progression-free and overall survivals were 83% and 100%, respectively. After allogeneic HSCT, the respective 12-month rates were 53% and 63%. No acute or late toxicities and no exacerbation of graft-versus-host disease were observed. Despite a low antigen burden and unsupportive recipient cytokine environment, CAR T cells persisted for an average of 201 days for autologous recipients and 51 days for allogeneic recipients.CONCLUSIONS. CD19-specific CAR T cells generated with SB and AaPC platforms were safe, and may provide additional cancer control as planned infusions after HSCT. These results support further clinical development of this nonviral gene therapy approach.

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“…Release criteria to produce banks were as described (36)(37)(38). Details of the manufacturing and expansion process are provided in the Supplemental Methods.…”

Section: Multiparameter Flow Cytometrymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells

Kebriaei¹,

Singh

Huls

et al. 2016

Journal of Clinical Investigation

437

390

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“…1). Use of gene-engineered T cells obviates the requirement for surgery because these T cells can be isolated from blood samples, and viral or non-viral transduction of receptors confers tumor Ag specificity 23) .…”

Section: Engineered T Cells Including T Cell Receptor Transduced T (mentioning

confidence: 99%

Prospects for personalized combination immunotherapy for solid tumors based on adoptive cell therapies and immune checkpoint blockade therapies

Kato

Yaguchi

Iwata

et al. 2017

Jpn. J. Clin. Immunol.

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summaryImmune checkpoint blockade (ICB) and adoptive cell therapies (ACT) with antigen-receptor gene-engineered T cells have been shown to be successful for a limited number of patients with solid tumors. Responders to ICB therapy typically have T cell-inflamed tumors. Thus, it is important to develop strategies that convert non-T cell-inflamed tumors to T cell-inflamed tumors. Although chimeric antigen receptor transduced T (CAR-T) cell therapy targeting hematological malignancies demonstrated durable clinical responses, the success of gene-engineered T cell therapies in solid tumors is hampered by a lack of unique antigens, antigen loss in cancer cells, and the immune-suppressive tumor microenvironment (TME) of solid tumors. However, gene-engineered T cells possess strong killing activity and cytokine production capacity, which can induce antigen spreading and modulate the TME of non-T cell-inflamed tumors seen in non-responders to ICB therapy. Immune responses against cancer are highly heterogeneous, not only between tumor types, but also within a patient or between different patients with the same type of cancer, indicating that personalized immunotherapy should be employed, based on the immune status of the individual patient. Here, we offer our perspective for personalized combination immunotherapy for solid tumors based on ACT and ICB therapies.

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“…DNA transposons have been used to efficiently insert gene cassettes into the host genomic DNA [67][68][69]. DNA transposon-based systems, such as the Sleeping Beauty (SB) and the PiggyBac transposon, have been used to engineer CAR T cells for clinical applications [70][71][72]. Although there is a theoretical risk of insertional oncogenesis, no transforming event in mature T cells after viral or non-viral gene transfer was so far observed.…”

Section: Car T Cells Are Successful In First Clinical Explorationsmentioning

confidence: 99%

CARs on the Highway: Chimeric Antigen Receptor Modified T Cells for the Adoptive Cell Therapy of Malignant Diseases

Holzinger¹,

Abken²

2017

Immunotherapy - Myths, Reality, Ideas, Future

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Adoptive therapy of malignant diseases by chimeric antigen receptor (CAR) redirected T cells takes advantage of the patient's own immune system to recognize and destroy cancer cells. This is impressively demonstrated by the induction of complete and lasting remissions of leukemia with CAR-engineered T cells in early phase trials. Recent developments in optimizing the CAR design, in the recognition of target cells and the production of modified cells for clinical use, have paved the path for a broader application than currently explored. The chapter reviews the differences in CAR design, the success in the treatment of hematologic malignancies, the challenges in treating solid cancer, the treatment-related toxicities, and strategies to improve safety of CAR T cell therapy. Challenges for future applications are discussed.

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Manufacture of T cells using the Sleeping Beauty system to enforce expression of a CD19-specific chimeric antigen receptor

Cited by 76 publications

References 55 publications

Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells

Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells

Prospects for personalized combination immunotherapy for solid tumors based on adoptive cell therapies and immune checkpoint blockade therapies

CARs on the Highway: Chimeric Antigen Receptor Modified T Cells for the Adoptive Cell Therapy of Malignant Diseases

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