Efficient adenoviral vector transduction of human hematopoietic SCID-repopulating and long-term culture-initiating cells

This article presents our studies on the adenoviral transduction efficiency, level of transgene expression, cell cycle status, and multilineage reconstitution ability of human CD34(+) hematopoietic cells transduced under proliferating and survival growth conditions. Bone marrow and umbilical cord blood CD34(+) cells were cultured in serum-free medium under survival conditions with thrombopoietin (Tpo) alone, or under proliferating conditions with Tpo, c-Kit ligand (KL), and Flt3 ligand (FL). Adenoviral vectors carrying the enhanced green fluorescent protein (EGFP) gene under the control of the PGK-1 promoter were used to transduce CD34(+) cells. Approximately 10% of CD34(+) cells were EGFP(+) under both culture conditions. In contrast, up to 50% of CD34(+) CD38(-) cells were EGFP(+), whereas a maximum of 8% of CD34(+) CD38(high) cells were EGFP(+) (p < 0.001). Both colony-forming unit cells (CFU-C) and 5-week long-term culture-initiating cells (LTC-ICs) were efficiently transduced. Under survival conditions, a substantial fraction of transduced CD34(+) cells remained quiescent. The nondividing CD34(+) EGFP(+) cells contained LTC-ICs capable of reconstituting longterm culture for as long as 10 weeks. CD34(+) EGFP(+) cells also retained the ability to engraft and multilineage-reconstitute NOD/SCID mice. These observations demonstrate that primitive human hematopoietic progenitor cells can be efficiently transduced by adenoviral vectors.