Cytokine and chemokine networks influencing stem cell proliferation, differentiation, and marrow homing

The hematopoietic stem cell (HSC) is an attractive target for gene therapy of genetic diseases of the immune and hematopoietic system, and for drug-resistance strategies in which genes conferring resistance to a variety of chemotherapeutic agents can be transduced. Stem cells are relatively easy to obtain; e.g., by marrow aspiration or G-CSF mobilization into the peripheral blood, and can be enriched e.g., by the use of anti-CD34 + monoclonal antibody. For conventional retroviral transduction, normally quiescent HSC must be activated into the cell cycle by priming with appropriate cytokines, and it has been critical to identify cytokine combinations that preserve the self-renewal capacity of long-term repopulating HSC. It has become apparent that strategies designed to optimize HSC cycling and proviral integration can compromise the capacity of transduced HSC to compete in vivo against endogenous HSC or HSC that have not been activated into cell cycle. Lentiviral vectors can integrate genes into non-cycling cells but there is an increased efficiency of transduction if Go HSC are activated into G1-phase of the cell cycle. This reduced efficiency of long-term engraftment of ex vivo cultured HSC may be due to impaired self-renewal capacity or reduced marrow homing efficiency. The latter may be attributed to down modulation of chemokine receptors necessary for chemotactic homing to the marrow. Alternatively, or in addition, there may be down modulation of (1) HSC adhesion molecules necessary for endothelial adhesion and egress from the circulation: (2) metalloproteinases secreted by HSC that facilitate their migration through extracellular matrix and promote release of critical soluble regulatory factors in the marrow microenvironment. A more controversial view is that cell death pathways, for example those involving FasR (CD95) may be activated in cycling HSC, resulting in their selective destruction upon transplantation and localization to sites rich in Fas ligand such as the liver.