Adenovirus and Retrovirus Mediated Interferon Alpha Gene Transfer into CD34+ Cells Maintains Regeneration Capacity and Enhances Adhesion Molecules in K562 Cells

Abstract

Background

Systemic administration of interferon-α (IFN-α) results in cytogenetic remissions and enhanced survival in a significant percentage of patients with chronic mylogenous leukemia (CML) and lymphoma. However, this treatment is associated with deleterious toxic effects. Gene transfer of the IFN-α gene into hematopoietic progenitors represents a novel strategy to deliver high concentrations of IFN-α to a local area.

Methods

We compared the effect of the transfer of the IFN-α gene on the cell growth and differentiation of several CD34+ cells in culture and in a NOD/SCID animal model, using adenovirus and retrovirus constructs.

Results

Transient local expression of the IFN-α gene using an adenovirus vector was associated with normal proliferation of CD34+ progenitors as measured by a colony forming unit of granulocyte-macrophage (CFU-GM) growth. Flow cytometric determination revealed that there was no significant difference in viability of these cells for 24-hour transduction periods. Reverse transcriptase/polymerase chain reaction (RT-PCR) analysis of RNA from CD34+ harvested CFU-GM progenitors demonstrated expression of IFN-α mRNA; radioimmunoassay (RIA) revealed that transduced cells secreted substantial levels of IFN-α protein. Furthermore, we constructed a retroviral vector in which IFN-α cDNA was driven by a viral LTR promoter to evaluate the effect of permanent IFN-α gene expression on cell growth. Retroviral packaging cells PA317 with high titers of retrovirus were produced and used to infect CD34+ and K562 cells. RIA showed that IFN-α-transduced CD34+ cells (with the aid of fibronectin fragment CH-296) produced approximately 400 units of IFN-α protein compared to CD34+ cells, or cells transduced with empty vector. IFN-α transduced CD34+ generated similar numbers of CFU-GM colonies as compared to control CD34+ cells. Engraftment of CD34+ cells transduced with IFN-α gene in NOD/SCID mice was successful for the first 30 days. Additionally, we studied the effect of local IFN-α expression on the cellular adhesion molecules, VLA-4, Mac-1, ICAM-1, and L-selectin in K562 cells, and human umbilical endothelial vein cells. K562 cells transduced with the IFN-α gene expressed a significantly elevated level of VLA-4, Mac-1, and ICAM-1.

Conclusion

We conclude that expression of the IFN-α gene using retrovirus vectors results in an adequate localized expression of IFN-α mRNA and protein.