Lentiviral gene therapy rescues p47phox chronic granulomatous disease and the ability to fight Salmonella infection in mice

Andrea Schejtman,Walmir Cutrim Aragão-Filho,Marta Zinicola,Maren Weisser,Adrian J Thrasher,Siobhan O Burns,David C Thomas,Hubert B Gaspar,Claire Booth,Antonio Condino-Neto,Simon Clare,Giorgia Santilli

doi:10.1038/s41434-020-0164-6

Abstract

Chronic granulomatous disease (CGD) is an inherited primary immunodeficiency disorder characterised by recurrent and often life-threatening infections and hyperinflammation. It is caused by defects of the phagocytic NADPH oxidase, a multicomponent enzyme system responsible for effective pathogen killing. A phase I/II clinical trial of lentiviral gene therapy is underway for the most common form of CGD, X-linked, caused by mutations in the gp91phox subunit of the NADPH oxidase. We propose to use a similar strategy to tackle p47phox-deficient CGD, caused by mutations in NCF1, which encodes the p47phox cytosolic component of the enzymatic complex. We generated a pCCLCHIM-p47phox lentiviral vector, containing the chimeric Cathepsin G/FES myeloid promoter and a codon-optimised version of the human NCF1 cDNA. Here we show that transduction with the pCCLCHIM-p47phox vector efficiently restores p47phox expression and biochemical NADPH oxidase function in p47phox-deficient human and murine cells. We also tested the ability of our gene therapy approach to control infection by challenging p47phox-null mice with Salmonella Typhimurium, a leading cause of sepsis in CGD patients, and found that mice reconstituted with lentivirus-transduced hematopoietic stem cells had a reduced bacterial load compared with untreated mice. Overall, our results potentially support the clinical development of a gene therapy approach using the pCCLCHIM-p47phox vector.

Highlights

Chronic granulomatous disease (CGD) is an inherited disorder of blood phagocytic cells that renders patients susceptible to certain bacterial and fungal infections and proneThese authors contributed : Adrian J
We propose to use a parallel lentiviral vector for the treatment of p47phox-deficient CGD (p47phox CGD). p47phox CGD accounts for ~25% of all CGD cases in western countries its frequency rises in regions with high degree of consanguinity [13,14,15]
The disease is caused by mutations in the NCF1 gene [16] encoding p47phox, the cytoplasmic component that, upon phosphorylation and translocation to the membrane, coordinates the assembly of the cytosolic subunits of NADPH oxidase resulting in the activation of the enzymatic complex. p47phox CGD has been historically associated with a milder phenotype compared with X-CGD as the majority of patients have residual superoxide production; infections are still a major cause of mortality and morbidity and patients still suffer from gut disease and inflammation [18]

Summary

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Chronic granulomatous disease (CGD) is an inherited disorder of blood phagocytic cells that renders patients susceptible to certain bacterial and fungal infections and prone. A lentiviral gene-therapy trial for the X-linked form of CGD has shown stable reconstitution of NADPH oxidase activity in six out of eight patients with >1-year follow up [11]. The disease is caused by mutations in the NCF1 gene [16] (a deletion of GT at the start of the exon 2 accounts for almost 84% of cases [17]) encoding p47phox, the cytoplasmic component that, upon phosphorylation and translocation to the membrane, coordinates the assembly of the cytosolic subunits of NADPH oxidase resulting in the activation of the enzymatic complex. We tested the pCCLCHIM-p47phox lentiviral vector for its ability to restore p47phox expression and NADPH oxidase function in human p47phox-deficient cells and in a murine model of the disease. Our results demonstrate that lentiviral gene therapy may be a valuable alternative to HSCT for patients with p47phox CGD lacking a suitable donor

Materials and methods

Results and discussion

Compliance with ethical standards