Retroviral Transfer and Long-Term Expression of the Adrenoleukodystrophy Gene in Human CD34+Cells

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Abstract
Adrenoleukodystrophy (ALD) is a demyelinating disease of the central nervous system that results from a genetic deficiency of ALDP, an ABC protein involved in the transport of very long-chain fatty acids (VLCFAs). The cloning of the ALD gene and the positive effects of allogeneic bone marrow transplantation support the feasibility of a gene therapy approach. We report the retroviral transfer of the ALD cDNA to peripheral blood and bone marrow CD34+ cells from control donors and ALD patients. Prestimulation of these cells with cytokines, followed by infection with the M48-ALD retroviral vector, resulted in 20% transduction efficiency (4–40%) and expression of the vector-encoded ALDP in 20% of CD34+ cells (7.3–50%). Long-term culture (LTC) of transduced CD34+ cells from two ALD patients showed efficient transduction (24–28%) and stable expression (25–32%) of ALDP in derived clonogenic progenitors at 3 weeks of culture. The expression of ALDP in CFU cells derived from 5 and 6 weeks of LTC confirmed the effective transduction of LTC-initiating cells. Expression of ALDP was observed in CD68+ CFU-derived cells, suggesting that monocyte-macrophages, the target bone marrow cells in ALD, were produced from transduced progenitor cells. VL-CFA content was corrected in LTC and CFU-derived cells in proportion to the percentage of transduced cells, indicating that the vector-encoded ALDP was functional. Although not efficient yet to allow a clinical perspective, these results demonstrate the feasibility of ALD gene transfer into CD34+ cells of ALD patients. Adrenoleukodystrophy (ALD), a severe demyelinating disease of the central nervous system, can be corrected by allogeneic bone marrow transplantation. Because of the limitations of this approach, ALD gene transfer into patient hematopoietic stem cells followed by autologous transplantation could be a valuable alternative. We report the retroviral transfer of the human ALD cDNA into CD34+ cells from peripheral blood or bone marrow from control donors and ALD patients. Expression of vector-encoded ALDP and correction of the biochemical defect of ALD is demonstrated in long-term bone marrow culture (LTC) and monocyte-macrophages derived from progenitor cells of two ALD patients.