Expression of Biologically Active Human Factor IX in Human Hematopoietic Cells after Retroviral Vector-Mediated Gene Transduction

Publisher Website
Google Scholar
Abstract
Gene therapy is a potential treatment for hemophilia, wherein cells transduced with a normal factor IX gene could provide a continuous in vivo source of circulating factor IX. In this study, we examined the potential use of hematopoietic cells as a target for factor IX gene therapy. Human myeloid leukemia cells (HL-60) were transduced by retroviral vectors carrying a normal human factor IX cDNA under control of either the Moloney murine leukemia virus long terminal repeat (MoMuLV LTR) (LIXSN), the SV40 promoter (LNSVIX), or a cytomegalovirus (CMV) promoter (LNCIX). Factor IX production was measured in the transduced cells both in the uninduced state and after induction of granulocytic differentiation [with dimethylsulfoxide (DMSO)] or monocytoid differentiation [with phorbol myristic acetate (PMA)]. Transcription of factor IX from the MoMuLV LTR was seen in all cells, with a two-fold increase upon differentiation. Induction with PMA led to an 8- to 15-fold increase in factor IX transcripts from an internal CMV promoter. No factor IX transcripts from the internal SV40 promoter were detected. Immunoreactive factor IX protein was identified by Western blot from induced HL-60 cells transduced by either LIXSN or LNCIX. Factor IX production by HL-60 cells transduced by LNCIX ranged from 38–93 ng/106 cells/24 hr following induction of monocytic differentiation. The factor IX antigen titer was directly related to factor IX coagulant titer (r = 0.98; p < 0.001). These data indicate that human myelomonocytic cells are capable of performing the necessary post-translational modifications to produce functional factor IX. These studies suggest the potential of autologous transplantation of factor IX-transduced bone marrow cells for gene therapy of hemophilia B. Hemophilia B results from genetic deficiency of factor IX. In this study, Hao et al. explore the possibility of using hematopoietic cells as the source of factor IX protein after gene therapy. The results demonstrate that some of the vectors that were examined produce significant amounts of factor IX, which possesses functional clot-promoting activity. Thus, gene therapy for hemophilia B may be performed using bone marrow cells.