In vivo expansion of gene-modified hematopoietic cells by a novel selective amplifier gene utilizing the erythropoietin receptor as a molecular switch

✍ Scribed by Takeyuki Nagashima; Yasuji Ueda; Yutaka Hanazono; Akihiro Kume; Hiroaki Shibata; Naohide Ageyama; Keiji Terao; Keiya Ozawa; Mamoru Hasegawa

• Publisher: John Wiley and Sons
• Year: 2004
• Tongue: English
• Weight: 314 KB
• Volume: 6
• Category: Article
• ISSN: 1099-498X
• DOI: 10.1002/jgm.467

✦ Synopsis

Abstract

Background

In vivo expansion of gene‐modified cells would be a promising approach in the field of hematopoietic stem cell gene therapy. To this end, we previously developed a selective amplifier gene (SAG), a chimeric gene encoding the granulocyte colony‐stimulating factor (G‐CSF) receptor (GCR), as a growth‐signal generator and the hormone‐binding domain of the steroid receptor as a molecular switch. We have already reported that hematopoietic cells retrovirally transduced with the SAG can be expanded in a steroid‐dependent manner in vitro and in vivo in mice and nonhuman primates. In this study, we have developed a new‐generation SAG, in which the erythropoietin (EPO) receptor (EPOR) is utilized instead of the steroid receptor as a molecular switch.

Methods

Two EPO‐driven SAGs were constructed, EPORGCR and EPORMpl, containing the GCR and c‐Mpl as a signal generator, respectively. First, to compare the steroid‐driven and EPO‐driven SAGs, Ba/F3 cells were transduced with these SAGs. Next, to examine whether GCR or c‐Mpl is the more suitable signal generator of the EPO‐driven SAG, human cord blood CD34^+^ cells were transduced with the two EPO‐driven SAGs (EPORMpl and EPORGCR). Finally, we examined the in vivo efficacy of EPORMpl in mice. Irradiated mice were transplanted with EPORMpl‐transduced bone marrow cells followed by administration of EPO.

Results

The EPO‐driven SAGs were shown to induce more rapid and potent proliferation of Ba/F3 cells than the steroid‐driven SAGs. The EPORMpl induced more efficient EPO‐dependent proliferation of the human cord blood CD34^+^ cells than the EPORGCR in terms of total CD34^+^ cell, c‐Kit^+^ cell, and clonogenic progenitor cell (CFU‐C) numbers. In the transplanted mice the transduced peripheral blood cells significantly increased in response to EPO.

Conclusions

The new‐generation SAGs, especially EPORMpl, are able to efficiently confer an EPO‐dependent growth advantage on transduced hematopoietic cells in vitro and in vivo in mice. Copyright © 2003 John Wiley & Sons, Ltd.