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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 (SM3/HMFG2) h(28BBζ), 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 a CD28 transmembrane domain/ endodomain and CD137 (4-1BB), CD3-zeta signaling domains. And the vector product was designed for the treatment of Colorectal cancer.
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.1 In vitro antitumor efficacy of CAR-MUC1 T cells co-culture with OME for 6hrs. Flow cytometry tested the lysis of different tumor cells by CAR-MUC1 T cells and GFP+ T cells, respectively. CAR-MUC1 T and GFP+ T cells were coculture with tumors cell from OME for 6 h. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.2 In vitro antitumor efficacy of CAR-MUC1 T cells co-culture with HN4 for 6hrs. Flow cytometry tested the lysis of different tumor cells by CAR-MUC1 T cells and GFP+ T cells, respectively. CAR-MUC1 T and GFP+ T cells were coculture with tumors cell from HN4 for 6 h. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.3 In vitro antitumor efficacy of CAR-MUC1 T cells co-culture with CAL33 for 6hrs. Flow cytometry tested the lysis of different tumor cells by CAR-MUC1 T cells and GFP+ T cells, respectively. CAR-MUC1 T and GFP+ T cells were coculture with tumors cell from CAL33 for 6 h. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.4 IL-2 secretion of CAR-MUC1 T cells and GFP+ T cells in coculture supernatants after different E/T ratio were measured by ELISA assay. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.5 IFN-γ secretion of CAR-MUC1 T cells and GFP+ T cells in coculture supernatants after different E/T ratio were measured by ELISA assay. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.6 TNF-α secretion of CAR-MUC1 T cells and GFP+ T cells in coculture supernatants after different E/T ratio were measured by ELISA assay. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.7 In vitro antitumor efficacy of CAR-MUC1 T cells co-culture with tumor cell after exogenous addition of IL22 recombinant protein. After 72 h of exogenous addition of IL22 recombinant protein, detected the killing of CAR-MUC1 T cells against OME cell line and different HNSCC cell lines in different E/T ratios. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.8 Cytokine secretion of CAR-MUC1 T cells and GFP+ T cells in coculture supernatants with different E/T ratio after exogenous addition of IL22 addition. After 72 hours of exogenous addition of IL22 recombinant protein, cytokine secretion in coculture supernatants after different E/T ratio were measured by ELISA assay. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.9 Microscopic observation of the ability of two different kinds of CAR-T cells and GFP+T cells to capture cells in 1:2 E/T ratio. In CAR-MUC1 T cells group, a little number of CAR-MUC1 T cells agglomerate around tumor cells; In CAR-MUC1-IL22 T cells group, large number of CAR-MUC1-IL22 T cells agglomerate around tumor cells (Red arrow is point to the tumor cell; the blue arrow is point to the T cell or two kinds of CAR-T cells). Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.10 In vitro antitumor efficacy of CAR-MUC1 T cells co-culture with tumor cell after for 72 h. Three different T cells were incubated with tumor cell (Cal33) and non-neoplastic epithelial cell (OME) at a low E/T ratio (1:1; 1:2; 1:5; 1:10) for 72 h, Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.11 Representative T-cell expansion profile of three different T cells. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.12 ELISA detected the secretion of IL22 in the culture supernatant. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.13 ELISA detected the secretion of IL22 in the coculture supernatant of different E/T ratio. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.14 ELISA detected the secretion of IL-2/IFN-γ/TNF-α in the coculture supernatant of low E/T ratio. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.15 CAR-T cells induce tumor degradation of HNSCC in vivo. 1E6 fLuc+ HN4 tumor cells were injected into the subcutaneous tissue in NSG mouse. After 10 d, tail intravenous injection of CAR T cells (n = 5). The mice were sacrificed on day 40 for tumor analysis. B, Bioluminescence images of four treatment groups mice. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.16 CAR-T cells induce tumor degradation of HNSCC in vivo. Total body bioluminescence units quantitate were measured by bioluminescence photometry and flux values (photons per second, P < .001). Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.17 CAR-T cells induce tumor degradation of HNSCC in vivo. Tumor volume growth curves of four treatment groups (P < .001). Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.18 CAR-T cells induce tumor degradation of HNSCC in vivo. T-cell infiltration statistics in four treatment groups (P < .01). Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
CAR Construction : SM3/HMFG2 scfv-41BB-CD3ζ-IL22 Fig.19 CAR-T cells induce tumor degradation of HNSCC in vivo. CD3 in tumor tissue was detected using the anti-CD3 antibody (Abcam 5690, dilution 1:200); cell nuclei were stained with DAPI. Mei, Z., Zhang, K., Lam, A. K. Y., Huang, J., Qiu, F., Qiao, B., & Zhang, Y. (2020). MUC1 as a target for CAR‐T therapy in head and neck squamous cell carinoma. Cancer medicine, 9(2), 640-652. |
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