Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia
Background C-type lectin-like molecule 1 (CLL-1) is highly expressed in acute myeloid leukemia (AML) but is absent in primitive hematopoietic progenitors, making it an attractive target for a chimeric antigen receptor (CAR) T-cell therapy. Here, we optimized our CLL-1 CAR for anti-leukemic activity...
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| Format: | Article |
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BMJ Publishing Group
2020-10-01
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| Series: | Journal for ImmunoTherapy of Cancer |
| Online Access: | https://jitc.bmj.com/content/8/2/e001229.full |
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| author | Maksim Mamonkin Malcolm K Brenner Norihiro Watanabe Pinar Ataca Atilla Mary K McKenna Haruko Tashiro Madhuwanti Srinivasan Feiyan Mo Brian Wesley Simons Alexandra McLean Stevens Michele S Redell Helen E Heslop Erden Atilla |
| author_facet | Maksim Mamonkin Malcolm K Brenner Norihiro Watanabe Pinar Ataca Atilla Mary K McKenna Haruko Tashiro Madhuwanti Srinivasan Feiyan Mo Brian Wesley Simons Alexandra McLean Stevens Michele S Redell Helen E Heslop Erden Atilla |
| author_sort | Maksim Mamonkin |
| collection | DOAJ |
| description | Background C-type lectin-like molecule 1 (CLL-1) is highly expressed in acute myeloid leukemia (AML) but is absent in primitive hematopoietic progenitors, making it an attractive target for a chimeric antigen receptor (CAR) T-cell therapy. Here, we optimized our CLL-1 CAR for anti-leukemic activity in mouse xenograft models of aggressive AML.Methods First, we optimized the CLL-1 CAR using different spacer, transmembrane and costimulatory sequences. We used a second retroviral vector to coexpress transgenic IL15. We measured the effects of each construct on T cell phenotype and sequential (recursive) co culture assays with tumor cell targets to determine the durability of the anti tumor activity by flow cytometry. We administered CAR T cells to mice engrafted with patient derived xenografts (PDX) and AML cell line and determined anti tumor activity by bioluminescence imaging and weekly bleeding, measured serum cytokines by multiplex analysis. After euthanasia, we examined formalin-fixed/paraffin embedded sections. Unpaired two-tailed Student’s t-tests were used and values of p<0.05 were considered significant. Survival was calculated using Mantel-Cox log-rank test.Results In vitro, CLL-1 CAR T cells with interleukin-15 (IL15) were less terminally differentiated (p<0.0001) and had superior expansion compared with CD28z-CD8 CAR T cells without IL15 (p<0.001). In both AML PDX and AML cell line animal models, CLL-1 CAR T coexpressing transgenic IL15 initially expanded better than CD28z-CD8 CAR T without IL15 (p<0.0001), but produced severe acute toxicity associated with high level production of human tumor necrosis factor α (TNFα), IL15 and IL2. Histopathology showed marked inflammatory changes with tissue damage in lung and liver. This acute toxicity could be managed by two strategies, individually or in combination. The excessive TNF alpha secretion could be blocked with anti-TNF alpha antibody, while excessive T cell expansion could be arrested by activation of an inducible caspase nine safety switch by administration of dimerizing drug. Both strategies successfully prolonged tumor-free survival.Conclusion Combinatorial treatment with a TNFα blocking antibody and subsequent activation of the caspase-9 control switch increased the expansion, survival and antileukemic potency of CLL-1 CAR T-cells expressing transgenic IL15 while avoiding the toxicities associated with excessive cytokine production and long-term accumulation of activated T-cells. |
| format | Article |
| id | doaj-art-7a32e4bf724e46cfb4155f71a646cc64 |
| institution | Kabale University |
| issn | 2051-1426 |
| language | English |
| publishDate | 2020-10-01 |
| publisher | BMJ Publishing Group |
| record_format | Article |
| series | Journal for ImmunoTherapy of Cancer |
| spelling | doaj-art-7a32e4bf724e46cfb4155f71a646cc642024-11-10T14:20:08ZengBMJ Publishing GroupJournal for ImmunoTherapy of Cancer2051-14262020-10-018210.1136/jitc-2020-001229Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemiaMaksim Mamonkin0Malcolm K Brenner1Norihiro Watanabe2Pinar Ataca Atilla3Mary K McKenna4Haruko Tashiro5Madhuwanti Srinivasan6Feiyan Mo7Brian Wesley Simons8Alexandra McLean Stevens9Michele S Redell10Helen E Heslop11Erden Atilla12Baylor College of Medicine, Houston, TX, USA6 Department of Pediatrics, Baylor College of Medicine, Houston, Texas, USACenter for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA2 Center for Comparative Medicine, Baylor College of Medicine, Houston, Texas, USA3 Division of Pediatric Hematology/Oncology, Texas Children`s Hospital, Houston, Texas, USA3 Division of Pediatric Hematology/Oncology, Texas Children`s Hospital, Houston, Texas, USAGraduate Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA1 Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USABackground C-type lectin-like molecule 1 (CLL-1) is highly expressed in acute myeloid leukemia (AML) but is absent in primitive hematopoietic progenitors, making it an attractive target for a chimeric antigen receptor (CAR) T-cell therapy. Here, we optimized our CLL-1 CAR for anti-leukemic activity in mouse xenograft models of aggressive AML.Methods First, we optimized the CLL-1 CAR using different spacer, transmembrane and costimulatory sequences. We used a second retroviral vector to coexpress transgenic IL15. We measured the effects of each construct on T cell phenotype and sequential (recursive) co culture assays with tumor cell targets to determine the durability of the anti tumor activity by flow cytometry. We administered CAR T cells to mice engrafted with patient derived xenografts (PDX) and AML cell line and determined anti tumor activity by bioluminescence imaging and weekly bleeding, measured serum cytokines by multiplex analysis. After euthanasia, we examined formalin-fixed/paraffin embedded sections. Unpaired two-tailed Student’s t-tests were used and values of p<0.05 were considered significant. Survival was calculated using Mantel-Cox log-rank test.Results In vitro, CLL-1 CAR T cells with interleukin-15 (IL15) were less terminally differentiated (p<0.0001) and had superior expansion compared with CD28z-CD8 CAR T cells without IL15 (p<0.001). In both AML PDX and AML cell line animal models, CLL-1 CAR T coexpressing transgenic IL15 initially expanded better than CD28z-CD8 CAR T without IL15 (p<0.0001), but produced severe acute toxicity associated with high level production of human tumor necrosis factor α (TNFα), IL15 and IL2. Histopathology showed marked inflammatory changes with tissue damage in lung and liver. This acute toxicity could be managed by two strategies, individually or in combination. The excessive TNF alpha secretion could be blocked with anti-TNF alpha antibody, while excessive T cell expansion could be arrested by activation of an inducible caspase nine safety switch by administration of dimerizing drug. Both strategies successfully prolonged tumor-free survival.Conclusion Combinatorial treatment with a TNFα blocking antibody and subsequent activation of the caspase-9 control switch increased the expansion, survival and antileukemic potency of CLL-1 CAR T-cells expressing transgenic IL15 while avoiding the toxicities associated with excessive cytokine production and long-term accumulation of activated T-cells.https://jitc.bmj.com/content/8/2/e001229.full |
| spellingShingle | Maksim Mamonkin Malcolm K Brenner Norihiro Watanabe Pinar Ataca Atilla Mary K McKenna Haruko Tashiro Madhuwanti Srinivasan Feiyan Mo Brian Wesley Simons Alexandra McLean Stevens Michele S Redell Helen E Heslop Erden Atilla Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia Journal for ImmunoTherapy of Cancer |
| title | Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia |
| title_full | Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia |
| title_fullStr | Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia |
| title_full_unstemmed | Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia |
| title_short | Modulating TNFα activity allows transgenic IL15-Expressing CLL-1 CAR T cells to safely eliminate acute myeloid leukemia |
| title_sort | modulating tnfα activity allows transgenic il15 expressing cll 1 car t cells to safely eliminate acute myeloid leukemia |
| url | https://jitc.bmj.com/content/8/2/e001229.full |
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