Nonmyeloablative conditioning regimen to increase engraftment of gene-modified hematopoietic stem cells in young rhesus monkeys.

Immune responses to transgene products may lead to rejection of transduced cells, limiting successful gene therapy for genetic diseases. While moderate dosages of chemotherapeutic agents such as busulfan may increase hematopoietic stem cells (HSC) engraftment, they are not immune suppressive and do not abrogate immune responses to transgene products. Studies focused on nonmyeloablative conditioning with busulfan ± fludarabine in a clinically relevant monkey model to induce immune suppression to allow cells expressing a foreign transgene product to persist. Bone marrow CD34(+) HSC were transduced in two equal fractions using simian immunodeficiency virus (SIV)-based lentiviral vectors carrying a nonexpressed DNA sequence tag (NoN) and the green fluorescent protein (GFP) reporter gene. Post-transplant there was no evidence of elimination of cells containing the potentially immunogenic GFP gene; several recipients had stable persistence of cells, and no differences were detected with fludarabine, which was rapidly cleared. Antibodies and cellular immune responses to GFP developed in recipients with the highest levels of GFP-marked cells, although these cells were not eliminated. These studies establish a clinically relevant pediatric primate model to assess the effects of conditioning regimens on the engraftment of transduced HSC and the immune responses to cells expressing a foreign gene product.