Circulating hematopoietic stem cells serve as novel targets for in utero gene therapy

Abstract

In utero gene therapy has been proposed as a method for permanent correction of somatic disorders that affect the hematopoietic system before disease initiation (1, 2). However, clinical trials using transplantation of allogenic fetal liver, bone marrow, or adult stem cells have been unsuccessful, largely due to the failure of sustained hematopoietic reconstitution in fetal recipients (3). Here, we reveal that retroviral transduction of unique repopulating stem cells found in the human fetal circulation is superior to full-gestation cord blood or adult sources. In contrast to postnatal human stem cells, the fetal circulation is highly enriched for actively cycling blood stem cells, thereby forming the basis for enhanced transduction. Our findings indicate that active, transducible hematopoietic reconstituting cells are present in the circulation of the human fetus and that they represent novel target cells for future in utero gene therapy trials using autologous transplantation.

Differential gene expression of human stem progenitor cells derived from early stages of in utero human hematopoiesis

“…FB and MPB samples were diluted (1:4) with ␣-minimal essential medium (GibcoBRL, Grand Island, NY) or phosphate-buffered saline (PBS), and mononuclear cells (MNCs) were isolated as previously described. 8,9 Figure 1 provides a schematic of our experimental approach using purified FB-and MPB-HSPCs. Given the individual variation between different human samples, FB and MPB samples from each population were pooled for SSH.…”

“…5 To further illustrate the importance of this phenotype, HSPC function segregates to the subset of Lin Ϫ CD34 ϩ CD38 Ϫ cells at all stages of human hematopoietic development, ranging from fetal to adult sources of HSPCs. [6][7][8][9] Despite the ability to isolate human HSPCs, our understanding of mechanisms governing self-renewal or differentiation of these rare cells is limited. For example, the intrinsic and extrinsic factors that regulate the mitotic transitions of human HSPCs have yet to be determined.…”

“…Over a series of investigations, a novel population of human HSPCs has been recently identified in the fetal circulation that possesses distinct proliferation and differentiation characteristics compared with adult circulating HSPCs harvested from mobilized peripheral blood (MPB). 8,9 Lin Ϫ CD34 ϩ CD38 Ϫ FB-HSPCs are distinct from adult Lin Ϫ CD34 ϩ CD38 Ϫ MPB-HSPCs in their frequency among total mononuclear cells, number of human SCID repopulating cells (SRCs), 8 enhanced retroviral transduction ability into HSPCs, in vivo myelopoietic reconstitution potential of FB-HSPCs, and the number of cycling HSPCs in FB that retains SRC capacity. 9 Accordingly, FB-HSPCs provide a unique source of human HSPCs that are actively cycling 8,9 with the opportunity to study and define the molecular mechanisms involved in HSPCs proliferation when self-renewal divisions are more frequent than in the adult hematopoietic system.…”

Efficient engraftment of in utero transplanted mice with retrovirally transduced hematopoietic stem cells

“…29,30 In addition, experimental studies with the ovine model showed the possibility of collecting fetal liver cells in sufficient numbers for the autologous transplantation of the animals. 31 The combined use of ex vivo HSC gene therapy and IUT may improve the therapeutic potential of both strategies used as individual therapies, thus facilitating the treatment of diseases diagnosed during fetal development.…”

Megakaryocyte Growth and Development Factor Is a Potent Growth Factor for Primitive Hematopoietic Progenitors in the Human Fetus

“…Administration of MGDF has also been shown to increase the mobilization of progenitors and stem cells (40 -42). Gene therapy performed on hematopoietic stem cells isolated from fetal and/or neonatal blood is being studied as a means to treat a number of inherited disorders of the hematopoi-1055 GROWTH OF FETAL PROGENITORS WITH MGDF etic system (43)(44)(45)(46)(47). Mobilization of stem cells by MGDF administration may increase the yield of stem cells that can be harvested for therapy and returned to the fetal or neonatal circulation.…”