title: Enhanced genetic modification of adult growth factor mobilized peripheral blood hematopoietic stem and progenitor cells with rapamycin.
abstract: Genetic modification of adult human hematopoietic stem and progenitor cells (HSPCs) with lentiviral vectors leads to long-term gene expression in the progeny of the HSPCs and has been used to successfully treat several monogenic diseases. In some cases, the gene-modified cells have a selective growth advantage over nonmodified cells and eventually are the dominant engrafted population. However, in disease indications for which the gene-modified cells do not have a selective advantage, optimizing transduction of HSPC is paramount to successful stem cell-based gene therapy. We demonstrate here that transduction of adult CD34+ HSPCs with lentiviral vectors in the presence of rapamycin, a widely used mTORC1 inhibitor, results in an approximately threefold increase in stable gene marking with minimal effects on HSPC growth and differentiation. Using this approach, we have demonstrated that we can enhance the frequency of gene-modified HSPCs that give rise to clonogenic progeny in vitro without excessive increases in the number of vector copies per cell or changes in integration pattern. The genetic marking of HSPCs and expression of transgenes is durable, and transplantation of gene-modified HSPCs into immunodeficient mice results in high levels of gene marking of the lymphoid and myeloid progeny in vivo. The prior safe clinical history of rapamycin in other applications supports the use of this compound to generate gene-modified autologous HSPCs for our HIV gene therapy clinical trials.
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Lijing
authors.initials: L
authors.lastname: Li
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Mónica
authors.initials: M
authors.lastname: Torres-Coronado
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Angel
authors.initials: A
authors.lastname: Gu
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Anitha
authors.initials: A
authors.lastname: Rao
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Agnes M
authors.initials: AM
authors.lastname: Gardner
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Elizabeth W
authors.initials: EW
authors.lastname: Epps
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Nancy
authors.initials: N
authors.lastname: Gonzalez
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Chy-Anh
authors.initials: CA
authors.lastname: Tran
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Xiwei
authors.initials: X
authors.lastname: Wu
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA.
authors.forename: Jin-Hui
authors.initials: JH
authors.lastname: Wang
authors.affiliation: Shared Resources-Cellular Process, City of Hope, Duarte, California, USA; Center for Blood Cell Therapies at Peter McCallum Center, Melbourne, Australia; Departments of Molecular and Cellular Biology and Virology, City of Hope, Duarte, California, USA ddigiusto@coh.org.
authors.forename: David L
authors.initials: DL
authors.lastname: DiGiusto
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D005434:Flow Cytometry; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods; D006412:Hematopoietic Stem Cells / Q000187:drug effects*; D006801:Humans; D066298:In Vitro Techniques; D016086:Lentivirus; D051379:Mice; D016688:Mice, Inbred NOD; D016133:Polymerase Chain Reaction; D020123:Sirolimus / Q000494:pharmacology*; D014161:Transduction, Genetic / Q000379:methods*
chemical_list: D016179:Granulocyte Colony-Stimulating Factor; D020123:Sirolimus
keywords: Genetic; HIV; Hematopoietic stem cells; Mice; SCID; Sirolimus; Transduction; Transplantation
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