A self-inactivating lentiviral vector for SCID-X1 gene therapy that does not activate LMO2 expression in human T cells

To develop safer and more effective vectors for gene therapy of X-linked severe combined immunodeficiency (SCID-X1), we have evaluated new self-inactivating lentiviral vectors based on the HIV virus. The CL20i4-hγ_c-Revgen vector contains the entire human common γ chain (γ_c) genomic sequence driven by the γ_c promoter. The CL20i4-EF1α-hγ_cOPT vector uses a promoter fragment from the eukaryotic elongation factor alpha (EF1α) gene to express a codon-optimized human γ_c cDNA. Both vectors contain a 400-bp insulator fragment from the chicken β-globin locus within the self-inactivating long-terminal repeat. Transduction of bone marrow cells using either of these vectors restored T, B, and natural killer lymphocyte development and function in a mouse SCID-X1 transplantation model. Transduction of human CD34⁺ bone marrow cells from SCID-X1 patients with either vector restored T-cell development in an in vitro assay. In safety studies using a Jurkat LMO2 activation assay, only the CL20i4-EF1α- hγ_cOPT vector lacked the ability to transactivate LMO2 protein expression, whereas the CL20i4-hγ_c-Revgen vector significantly activated LMO2 protein expression. In addition, the CL20i4-EF1α- hγ_cOPT vector has not caused any tumors in transplanted mice. We conclude that the CL20i4-EF1α-hγ_cOPT vector may be suitable for testing in a clinical trial based on these preclinical demonstrations of efficacy and safety.