Ex Vivo Manipulation of Mouse and Human Hematopoietic Progenitors

7.10.1. Ex Vivo Expansion

SCF has been an important component of conditions for ex vivo expansion of hematopoietic stem/progenitor cells in liquid culture for transplantation, in mouse and human systems (5,7,134). From selected human ex vivo expansion studies, SCF (with other cytokines such as IL-3, IL-6, G-CSF, EPO, and MGDF) is seen to be a critical component for expansion in suspension culture of CD34+ cells from bone marrow, cord blood, or mobilized peripheral blood (5,7,77,78,134). Depending on the starting population, and on the cytokine combinations used, maintenance or expansion of primitive cells (CD34+, LTC-IC/E-LTC-IC, LTR+) (70,74) and extensive expansion of erythroid, myeloid, and megakaryocytic progenitors and more differentiated cells occurs over periods of 1-3 wk. Some evidence indicates that hematopoietic stem/progenitor cells may have better LTR if they are not cycling (135,136), and if they express CXCR4, the receptor for the chemokine stromal cell-derived factor (SDF)-1, and other integrin-type adhesins that may facilitate homing. SCF may have a potential clinical benefit in both of these respects: by returning ex vivo expanded cells to relative quiescence before transplantation (137) and by upregulating expression of CXCR4 (137). Several groups have shown that clinical transplantation of mobilized peripheral blood progenitor cells (PBPCs) expanded ex vivo with a cocktail of cytokines that included MGDF, G-CSF, and SCF is promising in terms of short-term recovery of neutrophils (139-141) and platelets (140) after myeloablative chemotherapy; in these studies, unmanipulated PBPCs were transplanted along with the ex vivo-expanded PBPCs.

7.10.2. Gene Therapy

Retroviral DNA is unable to pass the nuclear envelope, such that at least one round of mitosis is a requirement for infection/DNA integration. SCF, typically along with other cytokines such as IL-3, IL-6, Flt3 ligand, and TPO, has been a critical component for enhancing retrovirally mediated gene transfection into primitive hematopoietic cells in developing the ex vivo portions of gene therapy protocols (5,7,142-144).

Transfection is highly efficient in murine systems, but less so for nonhuman primate and human. SCF probably contributes to initiation of target cell cycling (145-147) and may also upregulate binding sites for amphotrophic retroviral vectors on the target cells (148). In the human system, various reports have described transfection of genes including neomycin resistance, murine adenosine deaminase, glucocerebrosidase (deficient in the lysosomal storage disorder Gaucher's disease), and multidrug resistance (MDR) into LTC-IC and committed hematopoietic progenitors from human bone marrow, cord blood, and peripheral blood (5,7,142-144).

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