Application of a human multidrug transporter (ABCG2) variant as selectable marker in gene transfer to progenitor cells

Stem cell-based gene therapy is often unsuccessful because of the relatively low number of genetically modified cells with repopulating capabilities. To provide a selective advantage for the modified cells we applied the human ABCG2 protein, a resident xenobiotic transporter in stem cells, as a selectable marker. This protein is active as a homodimer, and its relatively small cDNA is an advantage in gene therapy applications. In the present study a mutant form of ABCG2 (R482G), showing drug-pumping activity with an altered substrate specificity, was coexpressed with a therapeutic gene by using a bicistronic vector and an efficient retroviral transduction protocol. Expression of the gp91(phox) protein in human gp91phox knockout hematopoietic progenitor cells corrected the loss-of-function mutation responsible for human chronic granulomatous disease, whereas the mutant ABCG2 protein selectively protected the transduced cells against clinically applicable cytotoxic agents. Overexpression of ABCG2 did not affect hematopoietic cell maturation or the restoration of granulocyte function by gp91(phox). We suggest that the mutant ABCG2 protein is an ideal candidate for human stem cell protection and for use as a selectable marker in gene therapy.