Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector
X-linked agammaglobulinemia (XLA) is an immune disorder caused by mutations in Bruton’s tyrosine kinase (BTK). BTK is expressed in B and myeloid cells, and its deficiency results in a lack of mature B cells and protective antibodies. We previously reported a lentivirus (LV) BTK replacement therapy t...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2021-03-01
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| Series: | Molecular Therapy: Methods & Clinical Development |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2329050121000073 |
| _version_ | 1818624471310794752 |
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| author | Brenda J. Seymour Swati Singh Hannah M. Certo Karen Sommer Blythe D. Sather Socheath Khim Courtnee Clough Malika Hale Joseph Pangallo Byoung Y. Ryu Iram F. Khan Jennifer E. Adair David J. Rawlings |
| author_facet | Brenda J. Seymour Swati Singh Hannah M. Certo Karen Sommer Blythe D. Sather Socheath Khim Courtnee Clough Malika Hale Joseph Pangallo Byoung Y. Ryu Iram F. Khan Jennifer E. Adair David J. Rawlings |
| author_sort | Brenda J. Seymour |
| collection | DOAJ |
| description | X-linked agammaglobulinemia (XLA) is an immune disorder caused by mutations in Bruton’s tyrosine kinase (BTK). BTK is expressed in B and myeloid cells, and its deficiency results in a lack of mature B cells and protective antibodies. We previously reported a lentivirus (LV) BTK replacement therapy that restored B cell development and function in Btk and Tec double knockout mice (a phenocopy of human XLA). In this study, with the goal of optimizing both the level and lineage specificity of BTK expression, we generated LV incorporating the proximal human BTK promoter. Hematopoietic stem cells from Btk−/−Tec−/− mice transduced with this vector rescued lineage-specific expression and restored B cell function in Btk−/−Tec−/− recipients. Next, we tested addition of candidate enhancers and/or ubiquitous chromatin opening elements (UCOEs), as well as codon optimization to improve BTK expression. An Eμ enhancer improved B cell rescue, but increased immunoglobulin G (IgG) autoantibodies. Addition of the UCOE avoided autoantibody generation while improving B cell development and function and reducing vector silencing. An optimized vector containing a truncated UCOE upstream of the BTK promoter and codon-optimized BTK cDNA resulted in stable, lineage-regulated BTK expression that mirrored endogenous BTK, making it a strong candidate for XLA therapy. |
| first_indexed | 2024-12-16T18:57:29Z |
| format | Article |
| id | doaj.art-4469c8d686fc425a84ca48e687ffdce5 |
| institution | Directory Open Access Journal |
| issn | 2329-0501 |
| language | English |
| last_indexed | 2024-12-16T18:57:29Z |
| publishDate | 2021-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Therapy: Methods & Clinical Development |
| spelling | doaj.art-4469c8d686fc425a84ca48e687ffdce52022-12-21T22:20:28ZengElsevierMolecular Therapy: Methods & Clinical Development2329-05012021-03-0120635651Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vectorBrenda J. Seymour0Swati Singh1Hannah M. Certo2Karen Sommer3Blythe D. Sather4Socheath Khim5Courtnee Clough6Malika Hale7Joseph Pangallo8Byoung Y. Ryu9Iram F. Khan10Jennifer E. Adair11David J. Rawlings12Center for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USAFred Hutchinson Cancer Research Center, Seattle, WA 98109, USA; Department of Medical Oncology, University of Washington, Seattle, WA 98195, USACenter for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USA; Departments of Pediatrics and Immunology, University of Washington, Seattle, WA 98195, USA; Corresponding author: David J. Rawlings, Center for Immunity and Immunotherapies, Seattle Children’s Research Institute, 1900 Ninth Avenue, Seattle, WA 98101, USA.X-linked agammaglobulinemia (XLA) is an immune disorder caused by mutations in Bruton’s tyrosine kinase (BTK). BTK is expressed in B and myeloid cells, and its deficiency results in a lack of mature B cells and protective antibodies. We previously reported a lentivirus (LV) BTK replacement therapy that restored B cell development and function in Btk and Tec double knockout mice (a phenocopy of human XLA). In this study, with the goal of optimizing both the level and lineage specificity of BTK expression, we generated LV incorporating the proximal human BTK promoter. Hematopoietic stem cells from Btk−/−Tec−/− mice transduced with this vector rescued lineage-specific expression and restored B cell function in Btk−/−Tec−/− recipients. Next, we tested addition of candidate enhancers and/or ubiquitous chromatin opening elements (UCOEs), as well as codon optimization to improve BTK expression. An Eμ enhancer improved B cell rescue, but increased immunoglobulin G (IgG) autoantibodies. Addition of the UCOE avoided autoantibody generation while improving B cell development and function and reducing vector silencing. An optimized vector containing a truncated UCOE upstream of the BTK promoter and codon-optimized BTK cDNA resulted in stable, lineage-regulated BTK expression that mirrored endogenous BTK, making it a strong candidate for XLA therapy.http://www.sciencedirect.com/science/article/pii/S2329050121000073gene therapyXLAUCOElentivirusDNase hypersensitive sitescodon optimization |
| spellingShingle | Brenda J. Seymour Swati Singh Hannah M. Certo Karen Sommer Blythe D. Sather Socheath Khim Courtnee Clough Malika Hale Joseph Pangallo Byoung Y. Ryu Iram F. Khan Jennifer E. Adair David J. Rawlings Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector Molecular Therapy: Methods & Clinical Development gene therapy XLA UCOE lentivirus DNase hypersensitive sites codon optimization |
| title | Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector |
| title_full | Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector |
| title_fullStr | Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector |
| title_full_unstemmed | Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector |
| title_short | Effective, safe, and sustained correction of murine XLA using a UCOE-BTK promoter-based lentiviral vector |
| title_sort | effective safe and sustained correction of murine xla using a ucoe btk promoter based lentiviral vector |
| topic | gene therapy XLA UCOE lentivirus DNase hypersensitive sites codon optimization |
| url | http://www.sciencedirect.com/science/article/pii/S2329050121000073 |
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