Journal
GENE THERAPY
Volume 19, Issue 8, Pages 872-876Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/gt.2012.37
Keywords
SCID-X1; self-inactivating; gamma-retrovirus; GMP; process development; clinical-grade
Categories
Funding
- Cincinnati Children's Research Foundation
- German Academic Exchange Service (DAAD)
- European Union
- German Ministry for Research (BMBF)
- Medical Research Council
- Wellcome Trust
- Great Ormond Street Children's Charity
- Great Ormond Street Hospital Childrens Charity [V1259] Funding Source: researchfish
- Medical Research Council [G0501969] Funding Source: researchfish
- MRC [G0501969] Funding Source: UKRI
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Patients with X-linked severe combined immunodeficiency (SCID-X1) were successfully cured following gene therapy with a gamma-retroviral vector (gRV) expressing the common gamma chain of the interleukin-2 receptor (IL2RG). However, 5 of 20 patients developed leukemia from activation of cellular proto-oncogenes by viral enhancers in the long-terminal repeats (LTR) of the integrated vector. These events prompted the design of a gRV vector with self-inactivating (SIN) LTRs to enhance vector safety. Herein we report on the production of a clinical-grade SIN IL2RG gRV pseudotyped with the Gibbon Ape Leukemia Virus envelope for a new gene therapy trial for SCID-X1, and highlight variables that were found to be critical for transfection-based large-scale SIN gRV production. Successful clinical production required careful selection of culture medium without pre-added glutamine, reduced exposure of packaging cells to cell-dissociation enzyme, and presence of cations in wash buffer. The clinical vector was high titer; transduced 68-70% normal human CD34(+) cells, as determined by colony-forming unit assays and by xenotransplantation in immunodeficient NOD. CB17-Prkdc(scid)/J (nonobese diabetic/severe combined immunodeficiency (NOD/SCID)) and NOD. Cg-Prkdc(scid) Il2rg(tm1Wjl)/SzJ (NOD/SCID gamma (NSG))) mice; and resulted in the production of T cells in vitro from human SCID-X1 CD34(+) cells. The vector was certified and released for the treatment of SCID-X1 in a multi-center international phase I/II trial.
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