Chronic Granulomatous Disease (CGD) is an inherited primary immunodeficiency
disorder with an incidence of ∼1:200,000 live births. This disease is caused by mutations
in any of the genes encoding subunits of the NAPDH oxidase, the enzyme responsible
for pathogen killing. Mutations in the p47 subunit of the NADPH oxidase
cause the most common form of autosomal recessive CGD, a disorder amenable to
haematopoietic stem cell (HSC) gene therapy. To this aim, I have developed and
tested in animal models a self-inactivating lentiviral vector containing a codon optimized
p47phox transgene under the transcriptional control of a myeloid promoter
(pCCLChim-p47).
The pCCLChim-p47 vector was able to induce high expression of the p47phox protein
and restoration of NADPH-oxidase in all models tested. In gene therapy-treated
mice levels of NADPH-oxidase activity were comparable to those found in WT animals.
The percentage of functional neutrophils remained stable over time in primary
and secondary transplants, suggesting that the vector is not prone to epigenetic inactivation.
In addition, gene corrected animals were protected against the Salmonella
pathogen with a reduction of CFU in the organs analyzed compared to KO and
Mock mice. Furthermore, gene transfer with the test vector pCCLChim-p47 had no
negative impact on the viability and proliferation of lineage negative cells, suggesting
the absence of a general vector- or transgene-related toxicity. Transduction with
the vector of human and mouse HSCs did not show adverse effects in the animals.
Probing that the lentiviral vector pCCLChim-p47 has a very low in vitro and in vivo
genotoxic risk, even at high vector copies per cell. Overall, our experiments paved
the way to a clinical development of the pCCLChimp47 vector.