Correction of murine models of
beta-thalassemia has been achieved through high-level
globin lentiviral vector gene transfer into mouse hematopoietic stem cells (HSCs). However, transduction of human HSCs is less robust and may be inadequate to achieve therapeutic levels of genetically modified erythroid cells. We therefore developed a double gene lentiviral vector encoding both human
gamma-globin under the transcriptional control of erythroid regulatory elements and methylguanine
methyltransferase (MGMT), driven by a constitutive cellular promoter. MGMT expression provides cellular resistance to
alkylator drugs, which can be administered to kill residual untransduced, diseased HSCs, whereas transduced cells are protected. Mice transplanted with beta-thalassemic HSCs transduced with
a gamma-globin/MGMT vector initially had subtherapeutic levels of red cells expressing
gamma-globin. To enrich
gamma-globin-expressing cells, transplanted mice were treated with the
alkylator agent 1,3-bis-chloroethyl-1-nitrosourea. This resulted in significant increases in the number of
gamma-globin-expressing red cells and the amount of
fetal hemoglobin, leading to resolution of
anemia. Selection of transduced HSCs was also obtained when cells were
drug-treated before
transplantation. Mice that received these cells demonstrated reconstitution with therapeutic levels of
gamma-globin-expressing cells. These data suggest that MGMT-based
drug selection holds promise as a modality to improve gene therapy for
beta-thalassemia.