Human CD34+ hematopoietic cells transduced by retrovirus-mediated interferon alpha gene maintains regeneration capacity and engraftment in NOD/SCID mice

To achieve long-term expression of human interferon alpha-5 (IFNa) gene in the bone marrow (BM) hematopoietic microenvironment, replication-deficient retroviral vector LSN-IFNa was used to deliver the IFNa gene into human BM CD34+ cells. After fibronectin-facilitated transduction, a fraction of CD34+ cells was plated in methylcellulose medium with or without G418 to assess transduction efficiency and the effect of IFNα gene transfer on colony formation. Colony-forming assay in the presence of G418 (400 mg/mL) revealed that 41% CFU-GM colonies are G418 resistant after infection with LSN-IFNa retrovirus. There was no significant difference in CFU-GM/BFU-E colony formation among IFNa gene-transduced CD34+ cells, control vector (LXSN) transduced-CD34+ cells and nontransduced CD34+ cells. Another portion of CD34+ cells was grown in liquid medium to measure IFNa production. RIA revealed that IFNa gene-transduced CD34+ cells produced 72.2 ± 15.4 U/mL (106cells/24hours) of IFNa compared with 8.3 ± 2.1 U/mL and 4.3 ± 1.2 U/mL in LXSN-transduced or nontransduced CD34+ cells, respectively. The remaining portion of transduced CD34+ cells was transplanted into immunodeficient (NOD/SCID) mice to allow analysis of long-term expression of IFNa. Transplantation of 1×106 CD34+ cells into sublethally irradiated NOD/SCID mice showed that IFNa and neor mRNA were detectable in engrafted mouse BM cells for up to 6 months. We conclude that continual local expression of IFNa in transduced CD34+ cells does not impair either CD34+ cell growth and differentiation or engraftment and long-term survival in NOD/SCID mice.