Long-term expression of gamma-globin mRNA in mouse erythrocytes from retrovirus vectors containing the human gamma-globin gene fused to the ankyrin-1promoter

Gene therapy for patients with hemoglobin disorders has been hampered by th e inability of retrovirus vectors to transfer globin genes and their cis-ac ting regulatory sequences into hematopoietic stem cells without rearrangeme nt. In addition, the expression from intact globin gene vectors has been va riable in red blood cells due to position effects and retrovirus silencing. We hypothesized that by substituting the globin gene promoter for the prom oter of another gene expressed in red blood cells, we could generate stable retrovirus vectors that would express globin at sufficient levels to treat hemoglobinopathies. Recently, we have shown that the human ankyrin (Ank) g ene promoter directs position-independent, copy number-dependent expression of a linked gamma -globin gene in transgenic mice. We inserted the Ank/(A) gamma -globin gene into retrovirus vectors that could transfer one or two c opies of the Ank/(A)gamma -globin gene to target cells. Both vectors were s table, transferring only intact proviral sequences into primary mouse hemat opoietic stem cells. Expression of Ank/(A)gamma -globin mRNA in mature red blood cells was 3% (single copy) and 8% (double copy) of the level of mouse alpha -globin mRNA, We conclude that these novel retrovirus vectors may be valuable for treating a variety of red cell disorders by gene replacement therapy including severe beta -thalassemia if the level of expression can b e further increased.