Large-scale manufacturing and characterization of CMV-CD19CAR T cells
Background Adoptive transfer of CD19-specific chimeric antigen receptor (CD19CAR) T cells can induce dramatic disease regression in patients with B cell malignancies. CD19CAR T cell therapy may be limited by insufficient engraftment and persistence, resulting in tumor relapse. We previously demonstr...
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BMJ Publishing Group
2022-01-01
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| Series: | Journal for ImmunoTherapy of Cancer |
| Online Access: | https://jitc.bmj.com/content/10/1/e003461.full |
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| author | Stephen J Forman Xiuli Wang Ryan Urak Miriam Walter Min Guan Tianxu Han Vibhuti Vyas Sheng-Hsuan Chien Brenna Gittins Mary C Clark Sally Mokhtari Angelo Cardoso Don J Diamond John Zaia Ryotaro Nakamura |
| author_facet | Stephen J Forman Xiuli Wang Ryan Urak Miriam Walter Min Guan Tianxu Han Vibhuti Vyas Sheng-Hsuan Chien Brenna Gittins Mary C Clark Sally Mokhtari Angelo Cardoso Don J Diamond John Zaia Ryotaro Nakamura |
| author_sort | Stephen J Forman |
| collection | DOAJ |
| description | Background Adoptive transfer of CD19-specific chimeric antigen receptor (CD19CAR) T cells can induce dramatic disease regression in patients with B cell malignancies. CD19CAR T cell therapy may be limited by insufficient engraftment and persistence, resulting in tumor relapse. We previously demonstrated a proof of principle that cytomegalovirus (CMV)-specific T cells can be isolated and enriched prior to CD19CAR transduction to produce CMV-CD19CAR T cells, and that these CMV-CD19CAR T cells can be expanded in vivo through CMV vaccination, resulting in better tumor control in a murine model. Here we developed a clinical platform for generating CMV-CD19CAR T cells.Methods Peripheral blood mononuclear cells (PBMCs) collected from CMV-seropositive healthy donors were stimulated with a good manufacturing practices-grade PepTivator overlapping CMVpp65 peptide pool and enriched for CMV-responsive interferon γ (IFNγ)+T cells using IFNγ Catchmatrix, within the CliniMACS Prodigy Cytokine Capture System (Miltenyi Biotec). Resulting CMV-specific T cells were transduced with a lentiviral vector encoding a second generation CD19R:CD28:ζ/EGFRt CAR and expanded with interleukin 2 (IL-2) and IL-15 for 15 days before characterization.Results CMV-specific T cells were enriched from 0.8%±0.5 of input PBMC to 76.3%±11.6 in nine full-scale qualification runs (absolute yield of 4.2±3.3×106 IFNγ+T cells from an input of 1×109 PBMCs). Average CD19CAR transduction efficiency of CMV-specific T cells was 27.0%±14.2 in the final products, which underwent rapid expansion, resulting in a total cell dose of 6.2±0.9 × 106 CD19CAR-tranduced T cells with CMV specificity (ie, functionally bispecific). CMV-CD19CAR T cells were polyclonal, expressed memory markers but had low expression of exhaustion markers, responded to both CD19 and CMVpp65 stimulation with rapid proliferation and exhibited antigen-specific effector functions against both CD19-expressing tumors and CMVpp65 antigen. The final products passed release criteria for clinical use.Conclusions We demonstrated the feasibility of our large-scale platform for generating CMV-CD19CAR T cells for clinical application. We plan to initiate a clinical trial at City of Hope using CMV-CD19CAR T cells for patients with intermediate/high-grade B cell non-Hodgkin’s lymphoma immediately after autologous hematopoietic cell transplantation followed by vaccination with a novel CMV vaccine based on Modified Vaccinia Ankara (Triplex) 28 days and 56 days post-T cell infusion. |
| format | Article |
| id | doaj-art-3a1427b2df1e43c0b87e330a814dd039 |
| institution | Kabale University |
| issn | 2051-1426 |
| language | English |
| publishDate | 2022-01-01 |
| publisher | BMJ Publishing Group |
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| series | Journal for ImmunoTherapy of Cancer |
| spelling | doaj-art-3a1427b2df1e43c0b87e330a814dd0392025-02-03T11:50:10ZengBMJ Publishing GroupJournal for ImmunoTherapy of Cancer2051-14262022-01-0110110.1136/jitc-2021-003461Large-scale manufacturing and characterization of CMV-CD19CAR T cellsStephen J Forman0Xiuli Wang1Ryan Urak2Miriam Walter3Min Guan4Tianxu Han5Vibhuti Vyas6Sheng-Hsuan Chien7Brenna Gittins8Mary C Clark9Sally Mokhtari10Angelo Cardoso11Don J Diamond12John Zaia13Ryotaro Nakamura142City of Hope, Duarte, CA, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Neurology, Jinan University First Affiliated Hospital, Guangzhou, Guangdong, ChinaDepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USADepartment of Clinical Translational Project Development, City of Hope, Duarte, California, USADepartment of Clinical Translational Project Development, City of Hope, Duarte, California, USACenter for Gene Therapy, Hematological Malignancy and Stem Cell Transplantation Institute, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USACenter for Gene Therapy, Hematological Malignancy and Stem Cell Transplantation Institute, City of Hope, Duarte, California, USADepartment of Hematology and Hematopoietic Cell Transplantation, City of Hope, Duarte, California, USABackground Adoptive transfer of CD19-specific chimeric antigen receptor (CD19CAR) T cells can induce dramatic disease regression in patients with B cell malignancies. CD19CAR T cell therapy may be limited by insufficient engraftment and persistence, resulting in tumor relapse. We previously demonstrated a proof of principle that cytomegalovirus (CMV)-specific T cells can be isolated and enriched prior to CD19CAR transduction to produce CMV-CD19CAR T cells, and that these CMV-CD19CAR T cells can be expanded in vivo through CMV vaccination, resulting in better tumor control in a murine model. Here we developed a clinical platform for generating CMV-CD19CAR T cells.Methods Peripheral blood mononuclear cells (PBMCs) collected from CMV-seropositive healthy donors were stimulated with a good manufacturing practices-grade PepTivator overlapping CMVpp65 peptide pool and enriched for CMV-responsive interferon γ (IFNγ)+T cells using IFNγ Catchmatrix, within the CliniMACS Prodigy Cytokine Capture System (Miltenyi Biotec). Resulting CMV-specific T cells were transduced with a lentiviral vector encoding a second generation CD19R:CD28:ζ/EGFRt CAR and expanded with interleukin 2 (IL-2) and IL-15 for 15 days before characterization.Results CMV-specific T cells were enriched from 0.8%±0.5 of input PBMC to 76.3%±11.6 in nine full-scale qualification runs (absolute yield of 4.2±3.3×106 IFNγ+T cells from an input of 1×109 PBMCs). Average CD19CAR transduction efficiency of CMV-specific T cells was 27.0%±14.2 in the final products, which underwent rapid expansion, resulting in a total cell dose of 6.2±0.9 × 106 CD19CAR-tranduced T cells with CMV specificity (ie, functionally bispecific). CMV-CD19CAR T cells were polyclonal, expressed memory markers but had low expression of exhaustion markers, responded to both CD19 and CMVpp65 stimulation with rapid proliferation and exhibited antigen-specific effector functions against both CD19-expressing tumors and CMVpp65 antigen. The final products passed release criteria for clinical use.Conclusions We demonstrated the feasibility of our large-scale platform for generating CMV-CD19CAR T cells for clinical application. We plan to initiate a clinical trial at City of Hope using CMV-CD19CAR T cells for patients with intermediate/high-grade B cell non-Hodgkin’s lymphoma immediately after autologous hematopoietic cell transplantation followed by vaccination with a novel CMV vaccine based on Modified Vaccinia Ankara (Triplex) 28 days and 56 days post-T cell infusion.https://jitc.bmj.com/content/10/1/e003461.full |
| spellingShingle | Stephen J Forman Xiuli Wang Ryan Urak Miriam Walter Min Guan Tianxu Han Vibhuti Vyas Sheng-Hsuan Chien Brenna Gittins Mary C Clark Sally Mokhtari Angelo Cardoso Don J Diamond John Zaia Ryotaro Nakamura Large-scale manufacturing and characterization of CMV-CD19CAR T cells Journal for ImmunoTherapy of Cancer |
| title | Large-scale manufacturing and characterization of CMV-CD19CAR T cells |
| title_full | Large-scale manufacturing and characterization of CMV-CD19CAR T cells |
| title_fullStr | Large-scale manufacturing and characterization of CMV-CD19CAR T cells |
| title_full_unstemmed | Large-scale manufacturing and characterization of CMV-CD19CAR T cells |
| title_short | Large-scale manufacturing and characterization of CMV-CD19CAR T cells |
| title_sort | large scale manufacturing and characterization of cmv cd19car t cells |
| url | https://jitc.bmj.com/content/10/1/e003461.full |
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