RETROVIRAL TRANSFER OF A HUMAN BETA-GLOBIN DELTA-GLOBIN HYBRID GENE LINKED TO BETA-LOCUS-CONTROL REGION HYPERSENSITIVE SITE-2 AIMED AT THE GENE-THERAPY OF SICKLE-CELL DISEASE

Human gamma-globin and delta-globin chains have been previously identi fied as strong inhibitors of the polymerization of hemoglobin S, in co ntrast to the beta-globin chain, which exerts only a moderate antisick ling effect. However, gamma-globin and delta globin are normally expre ssed at very low levels in adult erythroid cells, in contrast to beta- globin. We report the design of a beta-globin/delta-globin hybrid gene , beta/delta-sickle cell inhibitor 1 (beta/delta-SCI1) and its transdu ction by retrovirus-mediated gene transfer. The beta/delta-SCI1-encodi ng gene retains the overall structure of the human beta-globin gene, w hile incorporating specific amino acid residues from the delta chain p reviously found responsible for its enhanced antisickling properties. To achieve high expression levels of beta/delta-SCI1 in adult erythroc ytes, the hybrid gene was placed under the transcriptional control of the human beta-globin promoter and the DNase I hypersensitive site 2 o f the human beta locus control region. High-titer retroviruses were ge nerated, and stable proviral transmission was achieved in infected cel ls. The mRNA expression levels of the beta/delta-SCI1 gene in infected , dimethyl sulfoxide-induced murine erythroleukemia cells approached 8 5% of the endogenous murine beta(maj)-globin mRNA on a per gene basis, evidence that high gene expression levels were achieved in adult eryt hroid cells. Further evaluation of this strategy in transgenic animal models of sickle cell disease should assess its efficacy for the gene therapy of human patients.