The inability to transduce a sufficient fraction of the stem cell pool is a major limitation to gene therapy for sickle cell disease. In this project we attempt to overcome this obstacle by: a) identifying the optimal conditions for the transduction of human hemopoietic stem cells in vitro. Novel cytokines, such as flk-2 ligand, will be tested in our transduction protocols, as will the effects of substrata (CS-1, VCAM-1, or fibronectin), which may provide concentrated foci of retroviral particles. Due to the similarity between human and baboon hemopoiesis, transduction conditions that are optimal in humans will be applied to our primate model; b) Optimal mobilization conditions will be identified in primates so that the greatest possible number of stem cells can be harvested for ex vivo manipulation; c) Repeated cycles of mobilization and reinfusion will be performed using a vector containing a dominant selectable marker. In vivo selection will be applied using a drug to which cells containing the marker are resistant. A schedule of drug administration will be used that is intended to simultaneously provide mild conditioning and in vivo selection; d) As a control for conditioning and selection in c), repeated cycles of mobilization, transduction, and reinfusion, without myeloablation or selection will be performed; e) The relative efficiencies of MLV- versus AAV- based vectors to transduce long-term repopulating cells will be compared in individual animals; and f) the efficacy and safety of a new class of vectors based on human foamy virus and lentivirus will be tested.
| Constantinou, Varnavas C; Bouinta, Asimina; Karponi, Garyfalia et al. (2017) Poor stem cell harvest may not always be related to poor mobilization: lessons gained from a mobilization study in patients with ?-thalassemia major. Transfusion 57:1031-1039 |
| Gori, Jennifer L; Butler, Jason M; Kunar, Balvir et al. (2017) Endothelial Cells Promote Expansion of Long-Term Engrafting Marrow Hematopoietic Stem and Progenitor Cells in Primates. Stem Cells Transl Med 6:864-876 |
| Psatha, Nikoletta; Karponi, Garyfalia; Yannaki, Evangelia (2016) Optimizing autologous cell grafts to improve stem cell gene therapy. Exp Hematol 44:528-39 |
| Li, Li B; Ma, Chao; Awong, Geneve et al. (2016) Silent IL2RG Gene Editing in Human Pluripotent Stem Cells. Mol Ther 24:582-91 |
| Karponi, Garyfalia; Psatha, Nikoletta; Lederer, Carsten Werner et al. (2015) Plerixafor+G-CSF-mobilized CD34+ cells represent an optimal graft source for thalassemia gene therapy. Blood 126:616-9 |
| Vierstra, Jeff; Reik, Andreas; Chang, Kai-Hsin et al. (2015) Functional footprinting of regulatory DNA. Nat Methods 12:927-30 |
| Qi, Heyuan; Liu, Mingdong; Emery, David W et al. (2015) Functional validation of a constitutive autonomous silencer element. PLoS One 10:e0124588 |
| Liu, Mingdong; Maurano, Matthew T; Wang, Hao et al. (2015) Genomic discovery of potent chromatin insulators for human gene therapy. Nat Biotechnol 33:198-203 |
| Polak, Paz; Karli?, Rosa; Koren, Amnon et al. (2015) Cell-of-origin chromatin organization shapes the mutational landscape of cancer. Nature 518:360-364 |
| Chandrasekaran, Devikha; Nakamoto, Betty; Watts, Korashon L et al. (2014) Modeling promising nonmyeloablative conditioning regimens in nonhuman primates. Hum Gene Ther 25:1013-22 |
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