IN-VIVO SELECTION OF RETROVIRALLY TRANSDUCED HEMATOPOIETIC STEM-CELLS

One of the main impediments to effective gene therapy of blood disorde rs is the resistance of human hematopoietic stem cells to stable genet ic modification. We show here that a small minority of retrovirally tr ansduced stem cells can be selectively enriched in vivo, which might b e a way to circumvent this obstacle. We constructed two retroviral vec tors containing an antifolate-resistant dihydrofolate reductase cDNA t ranscriptionally linked to a reporter gene. Mice were transplanted wit h transduced bone marrow cells and then treated with an antifolate-bas ed regimen that kills unmodified stem cells. Drug treatment significan tly increased the percentage of vector-expressing peripheral blood ery throcytes, platelets, granulocytes, and T and B lymphocytes. Secondary transplant experiments demonstrated that selection occurred at the le vel of hematopoietic stem cells. This system for in vivo stem-cell sel ection provides a means to increase the number of genetically modified cells after transplant, and may circumvent an substantial obstacle to successful gene therapy for human blood diseases.