Adeno-associated virus 2-mediated transduction and erythroid lineage-restricted expression from parvovirus B19p6 promoter in primary human hematopoietic progenitor cells

Human parvovirus B19 gene expression from the viral p6 promoter (B19p6) is restricted to primary human hematopoietic cells undergoing erythroid differ entiation. We have demonstrated that expression from this promoter does not occur in established human erythoid cell lines in the context of a recombi nant parvovirus genome (Ponnazhagan et al. J Virol 69:8096-8101, 1995). How ever, abundant expression from this promoter can be readily detected in pri mary human bone marrow cells (Wang et al. Proc Natl Acad Sci USA 92:12416-1 2420, 1995; Ponnazhagan et al. J Gen Virol 77:1111-1122, 1996). In the pres ent studies, we investigated the pattern of expression from the B19p6 promo ter in primary human bone marrow-derived CD34(+) HPC undergoing differentia tion into myeloid and erythroid lineages. CD34(+) cells were transduced wit h recombinant adeno-associated virus 2 (AAV) vectors containing the beta-ga lactosidase (lacZ) gene under the control off the following promoters/enhan cers: the cytomegalovirus promoter (vCMVp-lacZ), B19p6 promoter (vB19p6-lac Z), B19p6 promoter with an upstream erythroid cell-specific enhancer elemen t (HS-2) from the locus control region (LCR) from the human beta-globin gen e cluster (vHS2-B19p6-lacZ), and the human beta-globin gene promoter with t he HS-2 enhancer (vHS2-beta p-lacZ). Transgene expression was evaluated eit her 48 h after infection or following erythroid differentiation in vitro fo r 3 weeks. Whereas high-level expression from the CMV promoter 48 h after i nfection diminished with time, low-level expression from the B19p6 grid the beta-globin promoters increased significantly following erythroid differen tiation. Furthermore; in HPC assays, there was no significant difference in the level of expression from the CMV promoter in myeloid or erythroid cell -derived colonies. Expression from the B19p6 and the beta-globin promoters, on the other hand, was restricted to erythroid cell colonies. These data f urther corroborate that the B19p6 promoter is erythroid cell-specific and s uggest that the recombinant AAV-B19 hybrid vectors may prove useful in gene therapy of human hemoglobinopathies in general and sickle cell anemia and beta-thalassemia in particular.