Lentiviral Gene Therapy Combined with Low-Dose Busulfan in Infants with SCID-X1

Abstract

Background: Allogeneic hematopoietic stem-cell transplantation for X-linked severe combined immunodeficiency (SCID-X1) often fails to reconstitute immunity associated with T cells, B cells, and natural killer (NK) cells when matched sibling donors are unavailable unless high-dose chemotherapy is given. In previous studies, autologous gene therapy with γ-retroviral vectors failed to reconstitute B-cell and NK-cell immunity and was complicated by vector-related leukemia.

Methods: We performed a dual-center, phase 1-2 safety and efficacy study of a lentiviral vector to transfer IL2RG complementary DNA to bone marrow stem cells after low-exposure, targeted busulfan conditioning in eight infants with newly diagnosed SCID-X1.

Results: Eight infants with SCID-X1 were followed for a median of 16.4 months. Bone marrow harvest, busulfan conditioning, and cell infusion had no unexpected side effects. In seven infants, the numbers of CD3+, CD4+, and naive CD4+ T cells and NK cells normalized by 3 to 4 months after infusion and were accompanied by vector marking in T cells, B cells, NK cells, myeloid cells, and bone marrow progenitors. The eighth infant had an insufficient T-cell count initially, but T cells developed in this infant after a boost of gene-corrected cells without busulfan conditioning. Previous infections cleared in all infants, and all continued to grow normally. IgM levels normalized in seven of the eight infants, of whom four discontinued intravenous immune globulin supplementation; three of these four infants had a response to vaccines. Vector insertion-site analysis was performed in seven infants and showed polyclonal patterns without clonal dominance in all seven.

Conclusions: Lentiviral vector gene therapy combined with low-exposure, targeted busulfan conditioning in infants with newly diagnosed SCID-X1 had low-grade acute toxic effects and resulted in multilineage engraftment of transduced cells, reconstitution of functional T cells and B cells, and normalization of NK-cell counts during a median follow-up of 16 months. (Funded by the American Lebanese Syrian Associated Charities and others; LVXSCID-ND ClinicalTrials.gov number, NCT01512888.).

Figure 1. Vector Marking in Blood and Marrow Cells after Gene Therapy.

Panels A through D show the vector copy number (VCN) per cell in peripheral‑blood T‑cell (CD3+), B‑cell (CD19+), NK‑cell (CD3−CD56+), and myeloid‑cell (CD14+/CD15+) lineages, respectively, as sorted by flow cytometry, in Patients 1 through 8 at various time points after cell infusion. Panel E shows the results of cells obtained by bone marrow aspiration at 4 and 12 months after gene therapy. The mean vector copy number per cell of selected CD34+ and CD19+ cells is shown. In addition, the mean vector copy number per cell measured in pooled myeloid colony‑forming unit (CFU) assays after 14 days of culture is shown. The vector copy number is shown as the mean vector genome copy per cell in the sorted populations and was determined by quantitative polymerase chain reaction with the use of a standard curve derived from a transduced single‑copy clonal cell line.

Figure 2. Immune Reconstitution after Gene Therapy

Panels A through F show absolute numbers of peripheral‑blood immune‑cell subsets, as determined by means of standard flow cytometry, over time after cell infusion. Dotted lines indicate values for T‑cell counts that are defined in the protocol as representing clinically significant reconstitution at 52 weeks after gene therapy. Panel G shows the quantity of DNA T‑cell–receptor excision circles (TRECs) in peripheral‑blood mononuclear cells, with a dotted line indicating the lower limit of the normal range. The values above the hash marks on the y axis range from 201 to 40,000.