Functional requirements for phenotypic correction of murine beta-thalassemia: implications for human gene therapy.

As initial human gene therapy trials for beta-thalassemia are contemplated, 2 critical questions important to trial design and planning have emerged. First, what proportion of genetically corrected hematopoietic stem cells (HSCs) will be needed to achieve a therapeutic benefit? Second, what level of expression of a transferred globin gene will be required to improve beta-thalassemic erythropoiesis? These questions were directly addressed by means of a murine model of severe beta-thalassemia. Generation of beta-thalassemic mice chimeric for a minority proportion of genetically normal HSCs demonstrated that normal HSC chimerism levels as low as 10% to 20% resulted in significant increases in hemoglobin (Hb) level and diminished extramedullary erythropoiesis. A large majority of the peripheral red cells in these mice were derived from the small minority of normal HSCs. In a separate set of independent experiments, beta-thalassemic mice were bred with transgenic mice that expressed different levels of human globins. Human gamma-globin messenger RNA (mRNA) expression at 7% of the level of total endogenous alpha-globin mRNA in thalassemic erythroid cells resulted in improved red cell morphology, a greater than 2-g/dL increase in Hb, and diminished reticulocytosis and extramedullary erythropoiesis. Furthermore, gamma-globin mRNA expression at 13% resulted in a 3-g/dL increase in Hb and nearly complete correction of red cell morphology and other indices of inefficient erythropoiesis. These data indicate that a significant therapeutic benefit could be achieved with expression of a transferred globin gene at about 15% of the level of total alpha-globin mRNA in patients with severe beta-thalassemia in whom 20% of erythroid precursors express the vector genome.