PT  - JOURNAL ARTICLE
AU  - Stefan Radtke
AU  - Dnyanada Pande
AU  - Margaret Cui
AU  - Anai M. Perez
AU  - Yan-Yi Chan
AU  - Mark Enstrom
AU  - Stefanie Schmuck
AU  - Andrew Berger
AU  - Tom Eunson
AU  - Jennifer E. Adair
AU  - Hans-Peter Kiem
TI  - Sort-purification of human CD34<sup>+</sup>CD90<sup>+</sup> cells reduces target cell population and improves lentiviral transduction for gene therapy
AID  - 10.1101/850479
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 850479
4099  - http://biorxiv.org/content/early/2020/01/21/850479.short
4100  - http://biorxiv.org/content/early/2020/01/21/850479.full
AB  - Hematopoietic stem cell (HSC) gene therapy has the potential to cure many genetic, malignant and infectious diseases. We have shown in a nonhuman primate (NHP) HSC gene therapy and transplantation model that the CD34+CD90+ cell fraction was exclusively responsible for multilineage engraftment and hematopoietic reconstitution. Here we show the translational potential of this HSC-enriched CD34 subset for lentivirus-mediated gene therapy. Alternative HSC-enrichment strategies include the purification of CD133+ cells or CD38low/- subsets of CD34+ cells from human blood products. We directly compared these strategies to the isolation of CD90+ cells using a GMP-grade flow-sorting protocol with clinical applicability. We show that CD90+ cell selection results in 40-fold fewer target cells in comparison to CD133+ or CD38low/- CD34 subsets without compromising the engraftment potential in vivo. Single cell RNA sequencing confirmed nearly complete depletion of lineage committed progenitor cells in CD90+ fractions compared to alternative selections. Importantly, lentiviral transduction efficiency in purified CD90+ cells resulted in up to 3-fold higher levels of engrafted gene-modified blood cells. These studies should have important implications for the manufacturing of patient-specific HSC gene therapy and genome editing products.