This project will test the feasibility of using a new gene transfer device to transfect primary human blood progenitors, in vitro. The genetically modified blood precursor cells will be reintroduced by autologous transfusion and permitted to undergo terminal maturation in vivo. The transfected precursor cells will therefore serve as an effective blood substitute for patients with blood abnormalities. They will test this new device and approach using human sickle cell disease as a model. Each of the current sickle cell treatment procedures (heterologous transfusion, chemical inducers of fetal hemoglobin, and retroviral gene transfer) have significant deficiencies. For example, the possibility of infection with blood borne pathogens such as HIV haunts heterologous transfusion. The chemical agents used to induce the fetal globin gene have various side effects, and retroviral gene transfer, although efficient, suffers in the case of globin gene transfer from low expression and the ever present possibility of the formation of recombinant viruses. In Phase I, they are combining three new technologies to provide a novel approach to blood disorders. These are: (a) the ability to culture purified primary human erythroblasts, (b) particle-mediated DNA transfer (gene gun), and (c) a proprietary assay for transfection of erythroid cells which permits one to quantify and optimize the absolute efficiency of gene transfection. This approach will permit us, in Phase II or III, to introduce a fetal globin gene, in a construction which promotes high expression, into primary erythroid cultures of a sickle cell patient. These cells, expanded in culture, could be reintroduced into the patient ---i.e., autologous transfusion.
