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Major limitations to gene therapy using HSCs are low gene transfer efficiency and the inability of most therapeutic genes to confer a selective advantage on the gene-corrected cells. One approach to enrich for gene-modified cells in vivo is to include in the retroviral vector a drug resistance gene, such as the P140K mutant of the DNA repair enzyme [O.sup.6]-methylguanine-DNA methyltransferase (MGMT *). We transplanted 5 rhesus macaques with [CD34.sup.+] cells transduced with lentiviral vectors encoding MGMT * and a fluorescent marker, with or without homeobox B4 (HOXB4), a potent stem cell self-renewal gene. Transgene expression and common integration sites in lymphoid and myeloid lineages several months after transplantation confirmed transduction of long-term repopulating HSCs. However, all animals showed only a transient increase in gene-marked lymphoid and myeloid cells after [O.sup.6]-benzylguanine (BG) and temozolomide (TMZ) administration. In 1 animal, cells transduced with MGMT * lentiviral vectors were protected and expanded after multiple courses of BG/TMZ, providing a substantial increase in the maximum tolerated dose of TMZ. Additional cycles of chemotherapy using 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) resulted in similar increases in gene marking levels, but caused high levels of nonhematopoietic toxicity. Inclusion of HOXB4 in the MGMT * vectors resulted in no substantial increase in gene marking or HSC amplification after chemotherapy treatment. Our data therefore suggest that lentivirally mediated gene transfer in transplanted HSCs can provide in vivo chemoprotection of progenitor cells, although selection of long-term repopulating HSCs was not seen., Introduction Retroviral vector--mediated gene transfer into HSCs has the potential to improve treatment for various genetic, malignant, and infectious diseases (1), (2). However, a critical limitation to stem cell gene [...]

The transcription factor homeobox B4 (HOXB4) is a promising agent capable of providing a growth advantage to genetically modified hematopoietic stem and progenitor cells (HSPCs). In this issue of the JCI, Zhang and colleagues overexpressed HOXB4 in HSPCs from large animals using retroviral vectors (see the related article beginning on page 1502). Two years after transplantation, most animals developed leukemia, a consequence of combined HOXB4 and deregulated protooncogene expression. These results highlight the risks of combining integrating vectors and growth-promoting genes for clinical applications., Hematopoietic stem cells and gene therapy Hematopoietic stem and progenitor cells (HSPCs) are ideal targets for permanent genetic correction of defects in any lineage of hematopoietic cells. Most clinical applications [...]