Lineage- and stage-restricted lentiviral vectors for the gene therapy of chronic granulomatous disease

Abstract

Insertional mutagenesis represents a serious adverse effect of gene therapy with integrating vectors. However, although uncontrolled activation of growth-promoting genes in stem cells can predictably lead to oncological processes, this is far less likely if vector transcriptional activity can be restricted to fully differentiated cells. Diseases requiring phenotypic correction only in mature cells offer such an opportunity, provided that lineage/stage-restricted systems can be properly tailored. In this study, we followed this reasoning to design lentiviral vectors for the gene therapy of chronic granulomatous disease (CGD), an immune deficiency due a loss of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in phagocytes, most often secondary to mutations in gp91 phox . Using self-inactivating HIV1-derived vectors as background, we first expressed enhanced green fluorescent protein (eGFP) from a minimal gp91 phox promoter, adding various natural or synthetic transcriptional regulatory elements to foster both specificity and potency. The resulting vectors were assessed either by transplantation or by lentiviral transgenesis, searching for combinations conferring strong and specific expression into mature phagocytic cells. The most promising vector was modified to express gp91 phox and used to treat CGD mice. High-level restoration of NADPH activity was documented in granulocytes from the treated animals. We propose that this lineage-specific lentiviral vector is a suitable candidate for the gene therapy of CGD.

Physiological regulation of transgene expression by a lentiviral vector containing the A2UCOE linked to a myeloid promoter

“…This approach should reduce the likelihood of transcriptional interference with cellular genes in hematopoietic stem cells, while maintaining full therapeutic benefit for the patients. 10,11 However, variegation of gene expression and promoter inactivation may occur, as gene-transduced cells in CGD lack a selective advantage over non-transduced cells and therefore selection for transgene-expressing cells, as is the case for X-linked severe-combined immunodeficiency, adenosine deaminase-deficient and Wiskott --Aldrich syndrome, is not expected to occur.…”

Development of gene therapy for blood disorders: an update

“…57 In an effort to achieve greater safety, lineage-and stage-restricted lentiviral vectors are being developed for treatment of CGD. 58,59 Other vector systems are also being considered, although they are in much earlier stages of development.…”

Gene therapy: too much splice can spoil the dish

“…Properly tailored, it could similarly serve to destabilize harmful fusion transcripts in cells particularly susceptible to transformation, namely stem cells and early precursors. In situations in which only differentiated cells require phenotypic correction for disease to be prevented, the safety margins of integrating gene therapy vectors could thus be significantly increased by combining stage- and lineage-specific promoters, to avoid protooncogene activation in stem cells and early precursors (17), and sequences targeted by miRNAs expressed in these cells, in which they would promote the degradation of dangerous cellular-viral fusion transcripts ( Figure 1C and ref. 18).…”