Efficient gene transfer to hematopoietic progenitor cells using SV40-derived vectors

We used recombinant SV40 (rSV40)-derived vectors to deliver transgenes to h uman and simian hematopoietic progenitor cells in culture, and in vivo afte r transduction ex vivo. rSV40 are highly efficient vectors that are made in very high titers. They infect almost all cells, whether resting or dividin g. Two rSV40s were used: SV(HBS), carrying hepatitis B surface antigen as a marker; and SV(Aw) carrying IN#33, a single chain Fv antibody against HIV- 1 integrase. CD34(+) cells derived from human fetal bone marrow (HFBM) and rhesus macaque bone marrow were transduced once with SV(HBS) without select ion. On average 60% of colonies derived from transduced CD34(+) cells carri ed and expressed HBsAg, as assessed by PCR and immunochemistry. Transgene c arriage persisted following differentiation of transduced rhesus CD34(+) ce lls into T lymphocytes. In an effort to increase the percentage of gene-mar ked cells, three sequential treatments of CD34(+) cells were done using SV( Aw), without selection. Two weeks later >95% of colonies expressed IN#33. U nselected SV(Aw)-transduced CD34(+) cells from HFBM were transplanted into sublethally irradiated SCID mice. Bone marrow harvested 3 months later show ed that >50% of bone marrow cells expressed IN#33. This is comparable with the percentage of human cells in these animals' bone marrow as judged by im munostaining for human CD45. The stability and longevity of transduction in this setting suggests that rSV40 vectors integrate into the cellular genom e. This possibility was supported by finding that PCR of genomic DNA using primer pairs with one cellular and one viral primer yielded PCR products on ly in transduced, but not control, cells. These PCR products hybridized wit h an SV40 DNA fragment. Thus, rSV40 vectors transduce normal human and prim ate bone marrow progenitor cells effectively without selection, and maintai n transgene expression in vivo following reimplantation. Such high efficien cy transduction may be useful in treating diseases of CD34(+) cells and the ir derivatives.