With second-generation self-inactivating (SIN) vectors, multiple SCID-X1 patients have successfully reconstituted T-cell immunity in the absence of early leukemic events 15, 16, 17 with a follow-up of up to 7 years.
Constitutive activation of the transgene 8, the choice of vectors 9 and specific details of the gene therapy procedure have all been proposed as factors contributing to the risk of leukemia and myelodysplastic syndrome that occurred in several trials for primary immunodeficiency disorders (PIDs) including SCID-X1 10, 11, chronic granulomatous disease (CGD) 12, 13 and Wiskott–Aldrich Syndrome (WAS) 14. It is concerning that patients developed leukemia from insertional oncogenesis both early and late, 15 years after transplantation of retroviral-based engineered cells 7. While most patients treated with first generation of gene therapy survived and benefited from the therapy, a substantial fraction (>25%) of patients developed leukemia from insertional oncogenesis 4, 5, 6.
This strategy has resulted in both successes and setbacks. Using integrating viral vectors, such as gamma-retroviral and lentiviral vectors, extra copies of a functional IL2RG gene are semi-randomly integrated into the genome of SCID-X1 patient-derived CD34 + HSPCs. Gene therapy is an alternative therapy to allo-HSCT. The importance of achieving gene correction in long-term hematopoietic stem cells (LT-HSCs) to achieve sustained clinical benefit is demonstrated by the waning of a functional immune system in patients who do not derive their immune system from LT-HSCs with a wild-type IL2RG gene. Because of the selective advantage of lymphoid progenitors expressing normal IL2RG, however, only a small number of genetically corrected hematopoietic stem and progenitor cells (HSPCs) are needed to reconstitute T-cell immunity 2, 3. Although allogeneic hematopoietic cell transplant (allo-HCT) is considered the standard of care for SCID-X1, it holds significant risks due to potential incomplete immune reconstitution, graft versus host disease (GvHD) and a decreased survival rate in the absence of an human leukocyte antigen (HLA)-matched sibling donor 1. Without early treatment, affected male infants die in the first year of life from infections. The gene encodes a shared subunit of the receptors for interleukin-2 (IL-2), IL-4, IL-7, IL-9, IL-15, and IL-21.
X-linked sSevere cCombined iImmunodeficiency (SCID-X1) is a primary immune deficiency disorder (PID) caused by mutations in the IL2RG gene on the X chromosome. In sum, our study provides specificity, toxicity and efficacy data supportive of clinical development of genome editing to treat SCID-Xl. We achieve high levels of targeting frequencies (median 45%) in CD34 + HSPCs from six SCID-X1 patients and demonstrate rescue of lymphopoietic defect in a patient derived HSPC population in vitro and in vivo. As measures of the lack of toxicity we observe no evidence of abnormal hematopoiesis following transplantation and no evidence of off-target mutations using a high-fidelity Cas9 as a ribonucleoprotein complex. Using a CRISPR-Cas9/AAV6 based strategy, we achieve up to 20% targeted integration frequencies in LT-HSCs. Here we describe an approach for X-linked sSevere cCombined iImmunodeficiency (SCID-X1) using targeted integration of a cDNA into the endogenous start codon to functionally correct disease-causing mutations throughout the gene. Gene correction in human long-term hematopoietic stem cells (LT-HSCs) could be an effective therapy for monogenic diseases of the blood and immune system.
0 kommentar(er)
