Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation
Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously, we demonstrated that dual co-stimulation can be effectively harnessed by a parallel (p)CAR architecture in which a CD28-containing second generation CAR is co-expressed with a 4-1BB containing chimeric...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2022-02-01
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| Series: | Frontiers in Immunology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fimmu.2022.836549/full |
| _version_ | 1819274114266824704 |
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| author | Leena Halim Kushal K. Das Daniel Larcombe-Young Adam Ajina Andrea Candelli Reuben Benjamin Reuben Benjamin Richard Dillon Richard Dillon David M. Davies John Maher John Maher John Maher John Maher |
| author_facet | Leena Halim Kushal K. Das Daniel Larcombe-Young Adam Ajina Andrea Candelli Reuben Benjamin Reuben Benjamin Richard Dillon Richard Dillon David M. Davies John Maher John Maher John Maher John Maher |
| author_sort | Leena Halim |
| collection | DOAJ |
| description | Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously, we demonstrated that dual co-stimulation can be effectively harnessed by a parallel (p)CAR architecture in which a CD28-containing second generation CAR is co-expressed with a 4-1BB containing chimeric co-stimulatory receptor (CCR). When compared to linear CARs, pCAR-engineered T-cells elicit superior anti-tumor activity in a range of pre-clinical models. Since CD19 is the best validated clinical target for cellular immunotherapy, we evaluated a panel of CD19-specific CAR and pCAR T-cells in this study. First, we generated a panel of single chain antibody fragments (scFvs) by alanine scanning mutagenesis of the CD19-specific FMC63 scFv (VH domain) and these were incorporated into second generation CD28+CD3ζ CARs. The resulting panel of CAR T-cells demonstrated a broad range of CD19 binding ability and avidity for CD19-expressing tumor cells. Each scFv-modified CAR was then converted into a pCAR by co-expression of an FMC63 scFv-targeted CCR with a 4-1BB endodomain. When compared to second generation CARs that contained an unmodified or mutated FMC63 scFv, each pCAR demonstrated a significant enhancement of tumor re-stimulation potential and IL-2 release, reduced exhaustion marker expression and enhanced therapeutic efficacy in mice with established Nalm-6 leukemic xenografts. These data reinforce the evidence that the pCAR platform delivers enhanced anti-tumor activity through effective provision of dual co-stimulation. Greatest anti-tumor activity was noted for intermediate avidity CAR T-cells and derived pCARs, raising the possibility that effector to target cell avidity is an important determinant of efficacy. |
| first_indexed | 2024-12-23T23:03:17Z |
| format | Article |
| id | doaj.art-039ee14d3847466d8a8145d5ff65a345 |
| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-12-23T23:03:17Z |
| publishDate | 2022-02-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Immunology |
| spelling | doaj.art-039ee14d3847466d8a8145d5ff65a3452022-12-21T17:26:52ZengFrontiers Media S.A.Frontiers in Immunology1664-32242022-02-011310.3389/fimmu.2022.836549836549Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-StimulationLeena Halim0Kushal K. Das1Daniel Larcombe-Young2Adam Ajina3Andrea Candelli4Reuben Benjamin5Reuben Benjamin6Richard Dillon7Richard Dillon8David M. Davies9John Maher10John Maher11John Maher12John Maher13Chimeric Antigen Receptor (CAR) Mechanics Laboratory, Guy’s Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomLumicks, Amsterdam, NetherlandsChimeric Antigen Receptor (CAR) Mechanics Laboratory, Guy’s Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomChimeric Antigen Receptor (CAR) Mechanics Laboratory, Guy’s Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomLumicks, Amsterdam, NetherlandsFaculty of Life Sciences and Medicine, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomDepartment of Clinical Haematology, King’s College Hospital National Health Service (NHS) Foundation Trust, London, United KingdomDepartment of Clinical Haematology, Guy’s and St Thomas’ National Health Service (NHS) Foundation Trust, London, United KingdomDepartment of Medicine and Molecular Genetics, King’s College London, London, United KingdomChimeric Antigen Receptor (CAR) Mechanics Laboratory, Guy’s Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomChimeric Antigen Receptor (CAR) Mechanics Laboratory, Guy’s Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, United KingdomLeucid Bio, Guy’s Hospital, London, United KingdomDepartment of Clinical Immunology and Allergy, King’s College Hospital National Health Service (NHS) Foundation Trust, London, United KingdomDepartment of Immunology, Eastbourne Hospital, Eastbourne, United KingdomCo-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously, we demonstrated that dual co-stimulation can be effectively harnessed by a parallel (p)CAR architecture in which a CD28-containing second generation CAR is co-expressed with a 4-1BB containing chimeric co-stimulatory receptor (CCR). When compared to linear CARs, pCAR-engineered T-cells elicit superior anti-tumor activity in a range of pre-clinical models. Since CD19 is the best validated clinical target for cellular immunotherapy, we evaluated a panel of CD19-specific CAR and pCAR T-cells in this study. First, we generated a panel of single chain antibody fragments (scFvs) by alanine scanning mutagenesis of the CD19-specific FMC63 scFv (VH domain) and these were incorporated into second generation CD28+CD3ζ CARs. The resulting panel of CAR T-cells demonstrated a broad range of CD19 binding ability and avidity for CD19-expressing tumor cells. Each scFv-modified CAR was then converted into a pCAR by co-expression of an FMC63 scFv-targeted CCR with a 4-1BB endodomain. When compared to second generation CARs that contained an unmodified or mutated FMC63 scFv, each pCAR demonstrated a significant enhancement of tumor re-stimulation potential and IL-2 release, reduced exhaustion marker expression and enhanced therapeutic efficacy in mice with established Nalm-6 leukemic xenografts. These data reinforce the evidence that the pCAR platform delivers enhanced anti-tumor activity through effective provision of dual co-stimulation. Greatest anti-tumor activity was noted for intermediate avidity CAR T-cells and derived pCARs, raising the possibility that effector to target cell avidity is an important determinant of efficacy.https://www.frontiersin.org/articles/10.3389/fimmu.2022.836549/fullchimeric antigen receptoravidityCD19parallel CARco-stimulationCD28 |
| spellingShingle | Leena Halim Kushal K. Das Daniel Larcombe-Young Adam Ajina Andrea Candelli Reuben Benjamin Reuben Benjamin Richard Dillon Richard Dillon David M. Davies John Maher John Maher John Maher John Maher Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation Frontiers in Immunology chimeric antigen receptor avidity CD19 parallel CAR co-stimulation CD28 |
| title | Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation |
| title_full | Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation |
| title_fullStr | Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation |
| title_full_unstemmed | Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation |
| title_short | Engineering of an Avidity-Optimized CD19-Specific Parallel Chimeric Antigen Receptor That Delivers Dual CD28 and 4-1BB Co-Stimulation |
| title_sort | engineering of an avidity optimized cd19 specific parallel chimeric antigen receptor that delivers dual cd28 and 4 1bb co stimulation |
| topic | chimeric antigen receptor avidity CD19 parallel CAR co-stimulation CD28 |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2022.836549/full |
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