Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy
Background: Chimeric antigen receptor (CAR) T cell treatment involves in vitro production of T cells from patient blood with synthetic receptors specific to a cancer antigen. They circumvent the major histocompatibility complex to recognize the tumor antigen, reducing hematologic malignancy remissio...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-05-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1159507/full |
| _version_ | 1797824133687410688 |
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| author | Yanyan Chen Pufeng Huang Mengda Niu Chuanhuizi Tian Tingting Zhang Zhiping Peng |
| author_facet | Yanyan Chen Pufeng Huang Mengda Niu Chuanhuizi Tian Tingting Zhang Zhiping Peng |
| author_sort | Yanyan Chen |
| collection | DOAJ |
| description | Background: Chimeric antigen receptor (CAR) T cell treatment involves in vitro production of T cells from patient blood with synthetic receptors specific to a cancer antigen. They circumvent the major histocompatibility complex to recognize the tumor antigen, reducing hematologic malignancy remission rates by 80%. Considering the efficacy of CAR-T treatment, the present work aimed at generating functional clusters of differentiation (CD)8 + T cells from human induced pluripotent stem cells (hiPSC) and to generate hiPS-CAR-T cells with high antigen-specific cytotoxicity.Methods: The Alkaline phosphatase assay and MycoEasy rapid mycoplasma detection kit was implemented for detection of hiPSCs and mycoplasma, respectively. The CD34+ HSPCs were harvested in AggreWellTM 400 using a 37-micron reversible strainer. Likewise, the lymphoid progenitor and CD4+CD8+ DP T cells were also harvested. The Cell Counting Kit-8 (CCK-8) assay was used to mark cytotoxicity and ELISA was used to detect IFN-γ secretion. Further, flow cytometry and transwell chambers were used to assess cell cycle, and migration and invasion. Finally, the in vivo antitumor effects of the CAR-T cells were evaluated using experimental animals (mice).Results: Results revealed that a serum-free, feeder layer-free differentiation system significantly yielded hiPSC-based T cell immunotherapy with interleukin-2, interleukin-15, and activators at the differentiation stage to promote the maturation of these cells into human induced pluripotent stem (hiPS)-T cells. The infection of hiPSCs with the CD19 CAR lentivirus resulted in the production of the hiPSC-CAR-T cells. We validated the function of hiPS-CAR-T cells in vivo and in vitro experimentation which revealed no significant differences in cell morphology and function between hiPSC-derived hiPS-CAR-T cells and peripheral blood-derived CAR-T cells.Conclusion: This study developed a culture method that is efficient and clinically useful to make functional CD8+ T cells from hiPSC and to get hiPS-CAR-T cells with high antigen-specific cytotoxicity that are not very different from CAR T cells found in peripheral blood. As a result, our findings may open the way for the clinical use of hiPSC to create functional CD8+ T and hiPS-CAR-T cells cells for use in cell-based cancer therapy. |
| first_indexed | 2024-03-13T10:34:29Z |
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| id | doaj.art-e02517fafee74415b94a41207b45adc7 |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-03-13T10:34:29Z |
| publishDate | 2023-05-01 |
| publisher | Frontiers Media S.A. |
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| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-e02517fafee74415b94a41207b45adc72023-05-18T07:36:21ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852023-05-011110.3389/fbioe.2023.11595071159507Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapyYanyan ChenPufeng HuangMengda NiuChuanhuizi TianTingting ZhangZhiping PengBackground: Chimeric antigen receptor (CAR) T cell treatment involves in vitro production of T cells from patient blood with synthetic receptors specific to a cancer antigen. They circumvent the major histocompatibility complex to recognize the tumor antigen, reducing hematologic malignancy remission rates by 80%. Considering the efficacy of CAR-T treatment, the present work aimed at generating functional clusters of differentiation (CD)8 + T cells from human induced pluripotent stem cells (hiPSC) and to generate hiPS-CAR-T cells with high antigen-specific cytotoxicity.Methods: The Alkaline phosphatase assay and MycoEasy rapid mycoplasma detection kit was implemented for detection of hiPSCs and mycoplasma, respectively. The CD34+ HSPCs were harvested in AggreWellTM 400 using a 37-micron reversible strainer. Likewise, the lymphoid progenitor and CD4+CD8+ DP T cells were also harvested. The Cell Counting Kit-8 (CCK-8) assay was used to mark cytotoxicity and ELISA was used to detect IFN-γ secretion. Further, flow cytometry and transwell chambers were used to assess cell cycle, and migration and invasion. Finally, the in vivo antitumor effects of the CAR-T cells were evaluated using experimental animals (mice).Results: Results revealed that a serum-free, feeder layer-free differentiation system significantly yielded hiPSC-based T cell immunotherapy with interleukin-2, interleukin-15, and activators at the differentiation stage to promote the maturation of these cells into human induced pluripotent stem (hiPS)-T cells. The infection of hiPSCs with the CD19 CAR lentivirus resulted in the production of the hiPSC-CAR-T cells. We validated the function of hiPS-CAR-T cells in vivo and in vitro experimentation which revealed no significant differences in cell morphology and function between hiPSC-derived hiPS-CAR-T cells and peripheral blood-derived CAR-T cells.Conclusion: This study developed a culture method that is efficient and clinically useful to make functional CD8+ T cells from hiPSC and to get hiPS-CAR-T cells with high antigen-specific cytotoxicity that are not very different from CAR T cells found in peripheral blood. As a result, our findings may open the way for the clinical use of hiPSC to create functional CD8+ T and hiPS-CAR-T cells cells for use in cell-based cancer therapy.https://www.frontiersin.org/articles/10.3389/fbioe.2023.1159507/fullimmunotherapycell differentiationhuman induced pluripotent stem cellslymphomagenesishiPS-CAR-T cells |
| spellingShingle | Yanyan Chen Pufeng Huang Mengda Niu Chuanhuizi Tian Tingting Zhang Zhiping Peng Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy Frontiers in Bioengineering and Biotechnology immunotherapy cell differentiation human induced pluripotent stem cells lymphomagenesis hiPS-CAR-T cells |
| title | Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy |
| title_full | Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy |
| title_fullStr | Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy |
| title_full_unstemmed | Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy |
| title_short | Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy |
| title_sort | regeneration of t cells from human induced pluripotent stem cells for car t cell medicated immunotherapy |
| topic | immunotherapy cell differentiation human induced pluripotent stem cells lymphomagenesis hiPS-CAR-T cells |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1159507/full |
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