Antioxidants Help ERT Efficacy in Pompe, Early Study Suggests

Yedida Y Bogachkov PhD avatar

by Yedida Y Bogachkov PhD |

Share this article:

Share article via email
stem cell gene therapy | Pompe Disease News | illustration of mice and test tubes

Antioxidants can increase the effectiveness of enzyme replacement therapy (ERT) for Pompe disease caused by oxidative stress, the damage in cells or tissues by reactive oxygen molecules, according to a study in cells and mice.

The findings also showed that enhancing autophagy — the destruction of damaged or redundant components within cellsreduces oxidative stress and increases ERT efficacy.

The study, “Correction of oxidative stress enhances enzyme replacement therapy in Pompe disease,” was published in the journal EMBO Molecular Medicine.

Pompe disease is caused by a mutation in the GAA gene, which codes for acid alpha-glucosidase, a protein within the lysosome — a cell’s recycling center. It is responsible for the breakdown of a large sugar called glycogen into glucose, which is then used by the body for energy production.

People with Pompe have a toxic accumulation of glycogen in their cells. ERT, a synthetic version of the GAA protein, is the standard Pompe treatment, but its efficacy decreases over time in some patients and is not the same in all tissues.

Studies have suggested that Pompe and other conditions associated with lysosomal impairment also are characterized by changes in autophagy, oxidative stress, and other cellular processes. In fact, research has indicated that autophagy impairment, which leads to the accumulation of cellular processes targeted for destruction, impacts ERT effectiveness.

Hypothesizing that the resulting increase of oxidative stress further limits ERT benefits, researchers in Italy used a mouse model of Pompe disease and measured oxidative stress within skeletal muscles, heart, and liver.

Results showed higher levels of reactive oxygen molecules and of oxidative damage in lipids (fat molecules) within the muscles of these mice, in comparison with control animals.

Compared with control cells, oxidative stress also was increased in fibroblasts — connective tissue cells — from patients with different types of Pompe disease, including classic infantile-onset, non-classic infantile-onset, and late-onset disease. Specifically, reactive molecules and damage to lipids were lower in fibroblasts from the late-onset group compared with the other patients.

“The results obtained in cells were reflective of the in vivo [in animals] findings, thus indicating that cultured cells, such as fibroblasts, are a reliable tool for further studies and for in vitro [in the lab] pharmacological manipulation of the oxidative stress pathway,” the scientists wrote.

Cells from Pompe mice had abnormal mitochondria — the structures responsible for producing energy — compared with control mice, the researchers found. Differences also were found regarding the presence of autophagy markers in mitochondria, which the scientists said were compatible with less efficient degradation of mitochondria in Pompe cells.

The researchers subsequently evaluated the effects of activating autophagy on oxidative stress. Using both cell starvation and pharmacological (molecules) to stimulate autophagy in fibroblasts from Pompe patients, the team found that autophagy activation led to a significant reduction in oxidative stress, compared with cells under normal conditions. In contrast, using bafilomycin to block autophagy further increased oxidative stress.

This shows that, in Pompe disease, “the autophagic pathway is impaired, but not totally blocked,” the investigators wrote.

In control fibroblasts, autophagy activation had a lesser effect than in Pompe cells, but blocking this process with bafilomycin substantially increased oxidative stress.

To investigate whether increased oxidative stress would affect GAA activity correction by ERT, the scientists administered alglucosidase alfa — an approved ERT marketed as Lumizyme by Sanofi Genzyme — into Pompe fibroblasts. They found that more oxidative stress in cells led to less efficient correction of GAA activity.

The researchers then found that a chemical stress inducer, sodium arsenite, caused a dose-dependent reduction in GAA activity while also impairing the amount of therapeutic enzyme taken up by the cells. This resulted in abnormal GAA maturation.

Using antioxidants reduced cellular stress and, given in combination with the therapeutic enzyme, improved correction of GAA activity compared with ERT alone. The best-performing antioxidant was N-acetylcysteine (NAC), which also was better at improving ERT delivery to lysosomes.

Autophagy activation increased GAA activity when Pompe fibroblasts were treated with ERT.

M6PR, a cell membrane protein necessary for the transport of enzymes into the lysosome, was found at lower levels in Pompe fibroblasts than in control cells. However, using antioxidants such as NAC significantly increased the amount of M6PR at the membrane. Enhancing autophagy also raised M6PR levels at the cell surface, but to a lesser extent than the antioxidants.

The team then found that, in the Pompe mouse model, antioxidants — NAC and idebenone, an approved therapy for Leber hereditary optic neuropathy — significantly increased the efficiency of ERT in enhancing GAA activity in tissues. The antioxidants also led to a general trend in glycogen decrease within the tissues, as opposed to ERT alone. A statistically significant decrease was seen in the quadriceps muscle (thigh).

“We show that factors related to recipient tissues, in addition to the intrinsic properties of the recombinant [therapeutic] enzyme, influence the response to ERT and possibly the outcome of patients,” the investigators wrote.

“Our work provides proof-of-concept evidence that reducing oxidative stress, one of the secondary abnormalities observed in PD [Pompe disease], is a strategy that may be pursued to enhance the effects of ERT,” they added. “As antioxidants drugs are approved for human therapy and show in general good safety profiles, their use would represent a convenient and safe option as adjunctive treatments for PD.”