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) Gene Transfer into CD34+ Cells and CML Mononuclear Cells
a Departments of Pharmacology and Medicine, New York Medical College, Valhalla, New York, USA;
b The Institute of Medical Science, The University of Tokyo, Tokyo, Japan;
c University of Rostock, Division of Hematology and Oncology, Rostock, Germany
Key Words. Gene transfer • 
interferon • Leukemia • Stem cells
Dr. Nader G. Abraham, Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.
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-
(IFN-) 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- to treat this disorder. Therefore, we examined the ability of adenovirus (Ad)-IFN- gene construct to transfect normal (CD34+ cells) and chronic myelogenous leukemia (CML) bone marrow mononuclear cells (BMMNC) and the transient overexpression of IFN- in these cells. Ad-cytomegalovirus promoter driven IFN- (AdCMV-IFN-) 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 h or 24 h transfection periods. Immunoassay of IFN- produced by CD34+ cells shows that IFN- levels increased several fold in transfected cells. Transient expression of the IFN- 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- mRNA expression. Similarly, CML BMMNC were transfected with AdCMV-IFN- under similar conditions as described for CD34+ cells. BMMNC cells exposed to adenovirus for 24 h and 48 h were found to express IFN- at a substantial level. This in vitro data suggest that Ad-mediated gene transfer of IFN- into hematopoietic stem cells can be achieved and that the IFN- gene can be translated into its specific mRNA in CD34 progenitor cells.
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