Alpha interferon (IFN-α) gene transfer into hematopoietic stem cells

Abstract

Gene transfer or gene therapy has advantages in the treatment of a variety of disorders due to its selective expression within specific mammalian cells. Interferon-a (IFN-α) has been used in the management of leukemia, but its diverse adverse activities with multiple potential side effects, possibly unrelated to therapeutic targets, may negatively influence the ability of IFN-α to treat this disorder. Therefore, we examined the ability of adenovirus (Ad)IFN-α gene construct to transfect normal (CD34+ cells) and chronic myelogenous leukemia (CML) bone marrow mononuclear cells (BMMNC) and the transient overexpression of IFN-α in these cells. Ad-cytomegalovirus (CMV) promoter-driven IFN-α (AdCMV-IFN-α) at multiple doses was assessed to transfect highly purified CD34+ cells in liquid culture, and optimal transduction of CD34+ cells was achieved using 120 plaque-forming units. Flow cytometric determinations revealed that there was no significant difference in cell viability for the 4- or 24-h transfection periods. Immunoassay of IFN-α produced by CD34+ cells shows that IFN-α levels increased several-fold in transfected cells. Transient expression of the IFN-α gene did not suppress proliferation of CD34+ progenitors as indicated by BFU-E or colony-forming units granulocyte, macrophage (CFU-GM) growth. Reverse transcriptase/polymerase chain reaction analysis of RNA from CD34+-harvested CFU-GM progenitor cells demonstrated transient IFN-α mRNA expression. Similarly, CML BMMNC were transfected with AdCMV-IFN-α under similar conditions as described for CD34+ cells. BMMNC exposed to adenovirus for 24- and 48 h were found to express IFN-α at a substantial level. These in vitro data suggest that adenovirus-mediated gene transfer of IFN-α into hematopoietic stem cells can be achieved and that the IFN-α gene can be translated into its specific mRNA in CD34 progenitor cells.