No abstract

IntroductionX-linked severe combined immunodeficiency (SCID-X1) is caused by loss-of-function mutations in the ␥ c gene (also known as IL2RG), 1,2 which encodes the common ␥-chain (␥ c ) subunit for the IL2, IL4, IL7, IL9, IL15, and IL21 receptors and is required for signal transduction with these cytokines. 3 Human SCID-X1 is characterized by lack of T, natural killer (NK) cells, and nonfunctional B cells and is fatal early in life from progressive infections if left untreated. Allogeneic stem cell transplantation is the current standard of care and can be curative, particularly when a matched sibling donor is available. 4 However, most patients lack a matched sibling donor and therefore receive transplants using haploidentical grafts obtained from a parent. These patients do significantly less well than those receiving matched sibling transplants, both in terms of overall survival and in terms of persistent immunodeficiency. [5][6][7] Most experts agree that better treatment is desirable for SCID-X1 patients who lack a matched sibling donor. [8][9][10] Gene therapy is one of several approaches that is being developed as an alternative to haploidentical allogeneic transplantation. [11][12][13][14][15] Pioneering studies done in Europe using vectors derived from the mouse Moloney leukemia virus (MLV) have proven that this approach can be effective because of restoration of T-cell numbers and function as well as humoral immunity. [11][12][13][14] However, the development of T-cell leukemia in 5 of 20 of these patients has uncovered a serious, unanticipated side effect associated with these gene therapy vectors. [16][17][18] Comprehensive molecular analyses of the leukemia cases have identified vector insertions into a small number of T-cell proto-oncogenes, most particularly LMO2, leading to its aberrant and high-level expression. [16][17][18] Further analysis of these integration sites has revealed that most, if not all, cases of gene activation were the result of the strong enhancer elements located in the U3 region of the MLV long-terminal repeat (LTR). Clearly, new vectors for SCID-X1 should not contain these MLV LTR enhancer elements to achieve better safety. This can be accomplished using self-inactivating (SIN) vectors that contain deletions in the U3 region of the LTR and is possible using either MLV or HIV-based vectors.We chose to use lentiviral vectors because they are more efficient for transducing quiescent human hematopoietic stem cells (HSCs), 19 and a recent study has suggested that, when an internal LTR enhancer is present, SIN lentiviral vectors require 10-fold higher copy numbers than an SIN-MLV vector to cause tumors in a tumor-prone mouse model, 20 presumably because of the known differences in integration site profile between HIV versus MLV vectors. 21 Another important consideration is the recent development of a stable packaging system for producing SIN lentiviral vectors. 22 This eliminates the need for using transient transfection methods with multiple plasmid DNAs to produce lentiviral...