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Sleeping Beauty (SB) transposon, a type of nonviral integrative vectors, provides an alternative to modify primary T cells. Creative biolabs has developed SB transposon CAR vector pSBCAR1 MUC1 (5E5) h(28ζ), which is constructed for the engineering of T cells to target human MUC1. The T cells are genetically modified through transduction with a nonviral vector expressing scFv of anti-MUC1 antibody linked to CD28 and CD3ζ signaling domains. And the vector product was designed for the treatment of Colorectal cancer.
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.1 IL-2 cytokine production in supernatant from 5E5 CAR T cells cultured on peptide-bound plates for 24 hr. Peptides were plated starting at concentrations of 2,000 ng/mL and diluted 256-fold until 1.82 ag/mL. 10 mg/mL of OKT3 mAb was used as a control T cell Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.2 IFN-r cytokine production in supernatant from 5E5 CAR T cells cultured on peptide-bound plates for 24 hr. Peptides were plated starting at concentrations of 2,000 ng/mL and diluted 256-fold until 1.82 ag/mL. 10 mg/mL of OKT3 mAb was used as a control T cell Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.3 Evaluation of 5E5 CAR T Cells Reactivity to a Panel of Human Primary Cells. 5E5 and CD19 CAR T cells were tested in a chromium release lysis assays at effector:target ratios of 1:1 to 30:1. 5E5 CAR see Blue line. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.4 Evaluation of 5E5 CAR T Cells Reactivity to a Panel of Human Primary Cells. 5E5 and CD19 CAR T cells were tested in a chromium release lysis assays at effector:target ratios of 1:1 to 30:1. 5E5 CAR see Blue line. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.5 Analysis of the cytokine-producing T cells from an intracellular cytokine assay of 5E5 CAR, cultured with K562, K562-meso, Jurkat, and NNP4 cells. NNP4 is a sample of primary epithelial ovarian cancer cells obtained from a pleural tap. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.6 Antitumor Efficacy of 5E5 CAR T Cells In Vitro. In vitro cytotoxicity assay of T cells, 5E5 CAR, CD19 CAR, or NTD, cultured with Jurkat cell line at the indicated effector-to-target ratios. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.7 Antitumor Efficacy of 5E5 CAR T Cells In Vivo. Serial bioluminescence imaging of NSG mice injected intravenously with 5E6 CBG+ Jurkat cells and infused with CAR-T, 1E7 or 5E6 cells as indicated, 8 days after tumor engraftment. The bottom right panel displays the mean bioluminescence by treatment group. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.8 Kaplan-Meyer survival curve of mice bearing Jurkat tumor by treatment group. Median survival of PBS group = 36.5, NTD group = 42, CD19 CAR group = 37.5, 5e6 5E5-CAR group = 63 and 1e7 5E5-CAR group = 94 days post T cell infusion. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.9 5E5 CAR T Cells Are Reactive against Multiple Tumor Histotypes. Indirect ELISA quantifying T cell (NTD or transduced with the indicated CAR) secretion of IFN-g during 24 hr culture with the indicated tumor cell lines. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.10 5E5 CAR T Cell-Treated Mice Exhibited Superior Tumor Rejection and Prolonged Survival against Disseminated Pancreatic Cancer. Mice were injected with 1 3 105 CBG+ Hs766T cells and imaged 6 days prior to T cell infusion and serially after treatment until 113 days post T cell infusion. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.11 5E5 CAR T Cell-Treated Mice Exhibited Superior Tumor Rejection and Prolonged Survival against Disseminated Pancreatic Cancer. Serial bioluminescence of NSG mice injected intraperitoneally with 1e5 CBG+ Hs766T metastatic pancreatic cancer cell line and infused with PBS or 1e6 of the indicated T cells when the mean tumor bioluminescence was in the 1e7-1e8 p/s range. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.12 5E5 CAR T Cell-Treated Mice Exhibited Superior Tumor Rejection and Prolonged Survival against Disseminated Pancreatic Cancer. Kaplan-Meyer survival curve of mice bearing the Hs766T tumor by treatment group (N = 6 mice per CD19 CAR and 5E5 CAR groups, N = 5 mice per PBS and NTD groups). Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
CAR Construction : 5E5 scfv-41BB-CD3ζ Fig.13 5E5 CAR T Cell-Treated Mice Exhibited Superior Tumor Rejection and Prolonged Survival against Disseminated Pancreatic Cancer. Immunohistochemistry of bone marrow, spleen, pancreas, liver, kidney, and tumor from mice bearing Hs766T tumor subcutaneously and treated with 5E5 CAR. Posey Jr, A. D., Schwab, R. D., Boesteanu, A. C., Steentoft, C., Mandel, U., Engels, B., ... & June, C. H. (2016). Engineered CAR T cells targeting the cancer-associated Tn-glycoform of the membrane mucin MUC1 control adenocarcinoma. Immunity, 44(6), 1444-1454. |
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