Prospects for developing a molecular cure for thalassemia.

The delineation of the thalassemia syndromes started in the late 1950s with the distinction of alpha and beta thalassemias and the initial studies that pointed out to the considerable genetic heterogeneity of these disorders. Molecular investigations, however, had to wait for the development of recombinant DNA and molecular cloning techniques in the 1970s. The thalassemias were the first human disorders to be delineated at the DNA level and indeed, by the mid 1980s their molecular pathology had been determined. Investigators in this field always hoped that the molecular understanding of thalassemias would provide clues for the development of molecular therapies and perhaps cures. This hypothesis, however, was proven wrong; no clues for therapies have come from the molecular analysis of the large number of thalassemia mutations that have so far been characterized. Hopes for a molecular cure are now resting on the development of gene therapy. Special for beta thalassemia and the other beta chain hemoglobinopathies is a form of therapy that has been revealed to us by nature, i.e. the alleviation of the beta chain deficiency by the production of fetal hemoglobin in the patient’s red cells. Prospects for the development of gene therapy for beta thalassemia syndromes. Gene therapy for thalassemia has been an early goal of investigators working in this field. Indeed, we were thinking about gene therapy even before the molecular understanding of the control of globin genes had reached a stage that could make this goal realistic. The ultimate goal of gene therapy of thalassemia is to correct the mutant globin gene in the patient’ hemopoietic cells. Gene correction, however, requires technologies which are still at a very early stage of development. The current goal of gene therapy for thalassemia is to add in the stem cells of the patients a normal b globin gene or a fetal globin gene (gene addition). In order to be successful, gene additions require that the therapeutic beta or gamma globin gene (1) is transferred to a large number the patient’s pluripotent repopulating stem cells; (2) it is regulated like a normal globin gene; (3) it is expressed at the very high level characteristic of the beta globin genes of normal individuals. During the last 20 years a major effort has been directed towards the development of vectors that fulfill these requirements. For about 15 years retroviral vectors were used because of the very good understanding of the retroviral biology, and because they integrate into the cell’s genome. Stable integration of therapeutic genes into the patient’s stem cells is an absolute requirement for gene therapy of thalassemia. However, retroviral vectors (oncoretroviral vectors) require cell division for integration and this is a disadvantage because the vast majority of hemopoietic stem cells are at rest. Systematic studies of globin gene transfer using retroviral vectors started in the mid 1980s but the results were disappointing because globin gene expression was inconsistent and very low. The discovery of the major regulatory element of the b locus, the locus control region (LCR) brought new impetus in the field and a large effort was made in several laboratories to produce new globin gene vectors containing the beta or gamma globin genes and components of the LCR that hopefully would have stimulated high globin gene expression. The initial excitement, however, was followed by new disappointment because these vectors were incapable of achieving therapeutic levels of globin gene expression; they were also unstable. The field was resurrected with the introduction of lentiviral vectors. These vectors are superior to oncoretroviral vectors because (1) they do not require cell division to enter the cell’s nucleus and to integrate into the cell’s genome. (2) They allow the incorporation in the vector of larger regulatory sequences from the locus control region;