Dataset Information

Efficient Ex Vivo Engineering and Expansion of Highly Purified Human Hematopoietic Stem and Progenitor Cell Populations for Gene Therapy.

ABSTRACT: Ex vivo gene therapy based on CD34+ hematopoietic stem cells (HSCs) has shown promising results in clinical trials, but genetic engineering to high levels and in large scale remains challenging. We devised a sorting strategy that captures more than 90% of HSC activity in less than 10% of mobilized peripheral blood (mPB) CD34+ cells, and modeled a transplantation protocol based on highly purified, genetically engineered HSCs co-infused with uncultured progenitor cells. Prostaglandin E2 stimulation allowed near-complete transduction of HSCs with lentiviral vectors during a culture time of less than 38 hr, mitigating the negative impact of standard culture on progenitor cell function. Exploiting the pyrimidoindole derivative UM171, we show that transduced mPB CD34+CD38- cells with repopulating potential could be expanded ex vivo. Implementing these findings in clinical gene therapy protocols will improve the efficacy, safety, and sustainability of gene therapy and generate new opportunities in the field of gene editing.

SUBMITTER: Zonari E

PROVIDER: S-EPMC5390102 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Efficient Ex Vivo Engineering and Expansion of Highly Purified Human Hematopoietic Stem and Progenitor Cell Populations for Gene Therapy.

Zonari Erika E Desantis Giacomo G Petrillo Carolina C Boccalatte Francesco E FE Lidonnici Maria Rosa MR Kajaste-Rudnitski Anna A Aiuti Alessandro A Ferrari Giuliana G Naldini Luigi L Gentner Bernhard B

Stem cell reports 20170316 4

Ex vivo gene therapy based on CD34<sup>+</sup> hematopoietic stem cells (HSCs) has shown promising results in clinical trials, but genetic engineering to high levels and in large scale remains challenging. We devised a sorting strategy that captures more than 90% of HSC activity in less than 10% of mobilized peripheral blood (mPB) CD34<sup>+</sup> cells, and modeled a transplantation protocol based on highly purified, genetically engineered HSCs co-infused with uncultured progenitor cells. Prost ...[more]

PMID: 28330619

Similar Datasets

Project description:Umbilical cord blood (UCB) has been established as an alternative source for hematopoietic stem/progenitor cells (HSPC) for cell and gene therapies. Limited cell yields of UCB units have been tackled with the development of cytokine-based ex vivo expansion platforms. To improve the effectiveness of these platforms, namely targeting clinical approval, in this study, we optimized the cytokine cocktails in two clinically relevant expansion platforms for HSPC, a liquid suspension culture system (CS_HSPC) and a co-culture system with bone marrow derived mesenchymal stromal cells (BM MSC) (CS_HSPC/MSC). Using a methodology based on experimental design, three different cytokines [stem cell factor (SCF), fms-like tyrosine kinase 3 ligand (Flt-3L), and thrombopoietin (TPO)] were studied in both systems during a 7-day culture under serum-free conditions. Proliferation and colony-forming unit assays, as well as immunophenotypic analysis were performed. Five experimental outputs [fold increase (FI) of total nucleated cells (FI TNC), FI of CD34+ cells, FI of erythroid burst-forming unit (BFU-E), FI of colony-forming unit granulocyte-monocyte (CFU-GM), and FI of multilineage colony-forming unit (CFU-Mix)] were followed as target outputs of the optimization model. The novel optimized cocktails determined herein comprised concentrations of 64, 61, and 80 ng/mL (CS_HSPC) and 90, 82, and 77 ng/mL (CS_HSPC/MSC) for SCF, Flt-3L, and TPO, respectively. After cytokine optimization, CS_HSPC and CS_HSPC/MSC were directly compared as platforms. CS_HSPC/MSC outperformed the feeder-free system in 6 of 8 tested experimental measures, displaying superior capability toward increasing the number of hematopoietic cells while maintaining the expression of HSPC markers (i.e., CD34+ and CD34+CD90+) and multilineage differentiation potential. A tailored approach toward optimization has made it possible to individually maximize cytokine contribution in both studied platforms. Consequently, cocktail optimization has successfully led to an increase in the expansion platform performance, while allowing a rational side-by-side comparison among different platforms and enhancing our knowledge on the impact of cytokine supplementation on the HSPC expansion process.

Dataset Information

Efficient Ex Vivo Engineering and Expansion of Highly Purified Human Hematopoietic Stem and Progenitor Cell Populations for Gene Therapy.

Publications

Efficient Ex Vivo Engineering and Expansion of Highly Purified Human Hematopoietic Stem and Progenitor Cell Populations for Gene Therapy.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets