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We have developed a novel system for expansion of gene-modified hematopoietic stem/progenitor cells to overcome the low efficiency of current gene transfer methodology. This system involves ‘selective amplifier genes’, that encode fusion proteins between the granulocyte colony-stimulating factor receptor (GCR) and the hormone-binding domain of estrogen receptor (ER). Hematopoietic progenitors expressing the chimeras showed estrogen-responsive growth in a controllable manner. However, endogenous estrogen may activate the fusion proteins in vivo, depending on the hormonal status of the subjects. We replaced ER with a mutant receptor (TmR) which specifically binds to 4-hydroxytamoxifen (Tm), to overcome limitations with wild-type ER. Interleukin-3 (IL-3)-dependent Ba/F3 cells and hematopoietic progenitor cells transduced with the resultant fusion proteins (GCRTmR and ΔGCRTmR) were examined for ligand-inducible growth. GCRTmR- and ΔGCRTmR-expressing Ba/F3 showed IL-3-independent growth in response to Tm, while the cells were unresponsive to estrogen at concentrations up to 10⁻⁷–10⁻⁶ M. Furthermore, murine bone marrow cells transduced with GCRTmR and ΔGCRTmR formed colonies in methylcellulose medium in response to Tm, while virtually no colonies appeared with 10⁻⁷ M estrogen or without cytokines. These results suggest that influences of the endogenous estrogen can be almost eliminated by using the GCRTmR/Tm or ΔGCRTmR/Tm system to expand gene-modified hematopoietic stem/progenitor cells. Copyright © 1999 John Wiley & Sons, Ltd.

We have developed a novel system for expansion of gene‐modified hematopoietic stem/progenitor cells to overcome the low efficiency of current gene transfer methodology. This system involves ‘selective amplifier genes’, that encode fusion proteins between the granulocyte colony‐stimulating factor receptor (GCR) and the hormone‐binding domain of estrogen receptor (ER). Hematopoietic progenitors expressing the chimeras showed estrogen‐responsive growth in a controllable manner. However, endogenous estrogen may activate the fusion proteins in vivo, depending on the hormonal status of the subjects.We replaced ER with a mutant receptor (TmR) which specifically binds to 4‐hydroxytamoxifen (Tm), to overcome limitations with wild‐type ER. Interleukin‐3 (IL‐3)‐dependent Ba/F3 cells and hematopoietic progenitor cells transduced with the resultant fusion proteins (GCRTmR and ΔGCRTmR) were examined for ligand‐inducible growth.GCRTmR‐ and ΔGCRTmR‐expressing Ba/F3 showed IL‐3‐independent growth in response to Tm, while the cells were unresponsive to estrogen at concentrations up to 10−7–10−6 M. Furthermore, murine bone marrow cells transduced with GCRTmR and ΔGCRTmR formed colonies in methylcellulose medium in response to Tm, while virtually no colonies appeared with 10−7 M estrogen or without cytokines.These results suggest that influences of the endogenous estrogen can be almost eliminated by using the GCRTmR/Tm or ΔGCRTmR/Tm system to expand gene‐modified hematopoietic stem/progenitor cells. Copyright © 1999 John Wiley & Sons, Ltd.