Final Activity Report Summary - FOAMYCGD (Development of Foamy Virus Vectors for Gene Replacement in Chronic Granulomatous Disease)
Genetic disorders that affect the ability of an organism to fight infections from common pathogens result in life-long hospital admissions, impaired quality of life and a shortened survival. One such disorder is chronic granulomatous disease (CGD) that is caused by a single gene disorder which results in dysfunctional white blood cells. The disease can only be treated by transplantation of normal haematopoietic stem cells (HSC) but only about 25 % of the patients can find suitable donors.
For the rest, a possible therapeutic strategy would be the correction of their own cells with a vector that carries the correct copy of the missing gene. This gene therapy approach has been successfully applied to a number of patients with other immunodeficiencies and it was the aim of our lab to develop vectors that can deliver genes to HSC. For gene transfer vehicles we used a retroviral vector based on the non-pathogenic virus called FoamyVirus (FV). The virus has the ability for permanent integration in the DNA of HSC and it prefers to integrate in areas of the genome that our away from known genes.
Overall this vector system has the following features: 1) it is non-pathogenic to humans, 2) is incapable for replication, 3) it is documented that it can transfer genes to HSC of mouse and human origin and 4) it has a relatively safe integration profile. Our research strategy was to generate FV vectors with a correct copy of the gene and assay their performance in the model cell line and in the preclinical mouse model.
After testing two FV vectors we constructed that displayed low levels of protein expression, we synthesized a vector that was optimized to work in mammalian cells. In the model cell line that we use, this vector showed levels of expression that were 88 % of the normal human cells with a correct copy of the missing gene. Such levels would be compatible with a clinical benefit in treated patients. In addition, the corrected cells expressed the protein stably over two months of continuous culture in vitro indicating that the vector was stable and the presence of the vector was not cytopathic to the cells.
We then tested our vector in HSC from a mouse model where preliminary ex vivo data showed the animals who received corrected HSC to have successfully engrafted indicating that the ex vivo genetic manipulations did not affect the HSC's ability to home to a host. The phenotypic correction of the CGD disorder in the animal model will be assayed after an extended observation period of 6-12 months in order to confirm the safety of our approach prior to embarking for a human clinical trial.
For the rest, a possible therapeutic strategy would be the correction of their own cells with a vector that carries the correct copy of the missing gene. This gene therapy approach has been successfully applied to a number of patients with other immunodeficiencies and it was the aim of our lab to develop vectors that can deliver genes to HSC. For gene transfer vehicles we used a retroviral vector based on the non-pathogenic virus called FoamyVirus (FV). The virus has the ability for permanent integration in the DNA of HSC and it prefers to integrate in areas of the genome that our away from known genes.
Overall this vector system has the following features: 1) it is non-pathogenic to humans, 2) is incapable for replication, 3) it is documented that it can transfer genes to HSC of mouse and human origin and 4) it has a relatively safe integration profile. Our research strategy was to generate FV vectors with a correct copy of the gene and assay their performance in the model cell line and in the preclinical mouse model.
After testing two FV vectors we constructed that displayed low levels of protein expression, we synthesized a vector that was optimized to work in mammalian cells. In the model cell line that we use, this vector showed levels of expression that were 88 % of the normal human cells with a correct copy of the missing gene. Such levels would be compatible with a clinical benefit in treated patients. In addition, the corrected cells expressed the protein stably over two months of continuous culture in vitro indicating that the vector was stable and the presence of the vector was not cytopathic to the cells.
We then tested our vector in HSC from a mouse model where preliminary ex vivo data showed the animals who received corrected HSC to have successfully engrafted indicating that the ex vivo genetic manipulations did not affect the HSC's ability to home to a host. The phenotypic correction of the CGD disorder in the animal model will be assayed after an extended observation period of 6-12 months in order to confirm the safety of our approach prior to embarking for a human clinical trial.