Marie Pouzolles1, Alice Machado1, Mickaël Guilbaud2, Magali Irla3, Sarah Gailhac1, Pierre Barennes4, Daniela Cesana5, Andrea Calabria5, Fabrizio Benedicenti5, Arnauld Sergé6, Indu Raman7, Quan-Zhen Li8, Eugenio Montini5, David Klatzmann9, Oumeya Adjali10, Naomi Taylor11, Valérie S Zimmermann12. 1. Institut de Génétique Moléculaire de Montpellier, University of Montpellier, CNRS, Montpellier, France. 2. INSERM UMR1089, Université de Nantes, Centre Hospitalier Universitaire de Nantes, Nantes, France. 3. Center of Immunology Marseille-Luminy (CIML), INSERM U1104, CNRS UMR7280, Aix-Marseille Université UM2, Marseille, France. 4. Sorbonne Université, INSERM, Immunology-Immunopathology-Immunotherapy (i3), Paris, France. 5. San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS, San Raffaele Scientific Institute, Milan, Italy. 6. Aix Marseille University, CNRS, INSERM, Institut Paoli-Calmettes, CRCM, Marseille, France. 7. Microarray Core Facility, University of Texas Southwestern Medical Center, Dallas, Tex. 8. Microarray Core Facility, University of Texas Southwestern Medical Center, Dallas, Tex; Department of Immunology, University of Texas Southwestern Medical Center, Dallas, Tex. 9. Sorbonne Université, INSERM, Immunology-Immunopathology-Immunotherapy (i3), Paris, France; AP-HP, Hôpital Pitié-Salpêtrière, Biotherapy (CIC-BTi) and Inflammation-Immunopathology-Biotherapy Department (i2B), Paris, France. 10. INSERM UMR1089, Université de Nantes, Centre Hospitalier Universitaire de Nantes, Nantes, France. Electronic address: oumeya.adjali@univ-nantes.fr. 11. Institut de Génétique Moléculaire de Montpellier, University of Montpellier, CNRS, Montpellier, France; Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Md. Electronic address: taylorn4@mail.nih.gov. 12. Institut de Génétique Moléculaire de Montpellier, University of Montpellier, CNRS, Montpellier, France. Electronic address: zimmermann@igmm.cnrs.fr.
Abstract
BACKGROUND: Patients with T-cell immunodeficiencies are generally treated with allogeneic hematopoietic stem cell transplantation, but alternatives are needed for patients without matched donors. An innovative intrathymic gene therapy approach that directly targets the thymus might improve outcomes. OBJECTIVE: We sought to determine the efficacy of intrathymic adeno-associated virus (AAV) serotypes to transduce thymocyte subsets and correct the T-cell immunodeficiency in a zeta-associated protein of 70 kDa (ZAP-70)-deficient murine model. METHODS: AAV serotypes were injected intrathymically into wild-type mice, and gene transfer efficiency was monitored. ZAP-70-/- mice were intrathymically injected with an AAV8 vector harboring the ZAP70 gene. Thymus structure, immunophenotyping, T-cell receptor clonotypes, T-cell function, immune responses to transgenes and autoantibodies, vector copy number, and integration were evaluated. RESULTS: AAV8, AAV9, and AAV10 serotypes all transduced thymocyte subsets after in situ gene transfer, with transduction of up to 5% of cells. Intrathymic injection of an AAV8-ZAP-70 vector into ZAP-70-/- mice resulted in a rapid thymocyte differentiation associated with the development of a thymic medulla. Strikingly, medullary thymic epithelial cells expressing the autoimmune regulator were detected within 10 days of gene transfer, correlating with the presence of functional effector and regulatory T-cell subsets with diverse T-cell receptor clonotypes in the periphery. Although thymocyte reconstitution was transient, gene-corrected peripheral T cells harboring approximately 1 AAV genome per cell persisted for more than 40 weeks, and AAV vector integration was detected. CONCLUSIONS: Intrathymic AAV-transduced progenitors promote a rapid restoration of the thymic architecture, with a single wave of thymopoiesis generating long-term peripheral T-cell function.
BACKGROUND:Patients with T-cell immunodeficiencies are generally treated with allogeneic hematopoietic stem cell transplantation, but alternatives are needed for patients without matched donors. An innovative intrathymic gene therapy approach that directly targets the thymus might improve outcomes. OBJECTIVE: We sought to determine the efficacy of intrathymic adeno-associated virus (AAV) serotypes to transduce thymocyte subsets and correct the T-cell immunodeficiency in a zeta-associated protein of 70 kDa (ZAP-70)-deficient murine model. METHODS:AAV serotypes were injected intrathymically into wild-type mice, and gene transfer efficiency was monitored. ZAP-70-/- mice were intrathymically injected with an AAV8 vector harboring the ZAP70 gene. Thymus structure, immunophenotyping, T-cell receptor clonotypes, T-cell function, immune responses to transgenes and autoantibodies, vector copy number, and integration were evaluated. RESULTS:AAV8, AAV9, and AAV10 serotypes all transduced thymocyte subsets after in situ gene transfer, with transduction of up to 5% of cells. Intrathymic injection of an AAV8-ZAP-70 vector into ZAP-70-/- mice resulted in a rapid thymocyte differentiation associated with the development of a thymic medulla. Strikingly, medullary thymic epithelial cells expressing the autoimmune regulator were detected within 10 days of gene transfer, correlating with the presence of functional effector and regulatory T-cell subsets with diverse T-cell receptor clonotypes in the periphery. Although thymocyte reconstitution was transient, gene-corrected peripheral T cells harboring approximately 1 AAV genome per cell persisted for more than 40 weeks, and AAV vector integration was detected. CONCLUSIONS: Intrathymic AAV-transduced progenitors promote a rapid restoration of the thymic architecture, with a single wave of thymopoiesis generating long-term peripheral T-cell function.
Authors: Matthew Elverman; Melissa A Goddard; David Mack; Jessica M Snyder; Michael W Lawlor; Hui Meng; Alan H Beggs; Ana Buj-Bello; Karine Poulard; Anthony P Marsh; Robert W Grange; Valerie E Kelly; Martin K Childers Journal: Muscle Nerve Date: 2017-05-22 Impact factor: 3.217
Authors: Suk See De Ravin; Xiaolin Wu; Susan Moir; Sandra Anaya-O'Brien; Nana Kwatemaa; Patricia Littel; Narda Theobald; Uimook Choi; Ling Su; Martha Marquesen; Dianne Hilligoss; Janet Lee; Clarissa M Buckner; Kol A Zarember; Geraldine O'Connor; Daniel McVicar; Douglas Kuhns; Robert E Throm; Sheng Zhou; Luigi D Notarangelo; I Celine Hanson; Mort J Cowan; Elizabeth Kang; Coleen Hadigan; Michael Meagher; John T Gray; Brian P Sorrentino; Harry L Malech Journal: Sci Transl Med Date: 2016-04-20 Impact factor: 17.956