Mouse Study Alerts Potential Gene Therapy Issues in Female Patients

Marta Figueiredo PhD avatar

by Marta Figueiredo PhD |

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A gene therapy-like approach that provided a full-length MECP2 — the gene mutated in most cases of Rett syndrome — to cells safely and effectively lessened disease-specific symptoms in a mouse model of a milder form of Rett, a study shows.

However, female mice treated with this approach showed signs of motor-related adverse events associated with excessive, toxic levels of MeCP2, the protein coded by the MECP2 gene and missing or defective in Rett patients.

These early findings add to the large body of data highlighting the fine balance of protein levels that MECP2-directed gene therapy strategies have to achieve to be both safe and effective.

They also suggest that the brain circuit regulating motor function is particularly sensitive to high MeCP2 levels in females, and emphasize the importance of including female mice in preclinical studies of Rett syndrome, the team noted.

The study, “Safety and efficacy of genetic MECP2 supplementation in the R294X mouse model of Rett syndrome,” was published in the journal Genes, Brain, and Behavior.


Affecting females almost exclusively, Rett syndrome is caused mostly by mutations in the MECP2 gene, which is located on the X chromosome (one of the sex chromosomes). The MeCP2 protein regulates other genes’ activities and is involved in nerve cell function and communication.

MeCP2 levels are tightly regulated, as both deficiency and overproduction lead to cellular toxicity and disease. Notably, excessive MeCP2 levels mainly due to MECP2 gene duplication lead to a neurodevelopmental disorder called MECP2 duplication syndrome, known as MDS.

While Rett syndrome and MDS share motor features and other symptoms, MDS affects males almost exclusively and is also associated with an impaired immune system.

“Given this narrow therapeutic window of MECP2 dosage, interventions for RTT [Rett syndrome] that supplement full-length MeCP2 protein must do so without eliciting excess MeCP2 levels and MDS-like adverse effects,” the researchers wrote.

Achieving this fine balance of MeCP2 levels through gene therapy — which works by delivering a healthy copy of the disease-causing gene to cells — is even more challenging due to Rett-associated mosaicism in female patients.

Mosaicism, which refers to the presence of groups of cells in the body that are genetically distinct, is caused by X-chromosome inactivation, a process that only occurs in females due to the presence of two X chromosomes (one from each biological parent).

X-chromosome inactivation leads to the random “turn-off” of one of the X chromosomes in each cell, meaning that female Rett patients typically show a mosaic pattern of cells producing a working MeCP2 protein, and cells with a defective or missing protein.

Now, researchers at Vanderbilt University evaluated the effects of supplementing a full-length, working MECP2 gene to cells in a mouse model of a milder form of Rett syndrome. The mice carried a common MECP2 mutation in Rett patients, called R294X, that leads to the production of a shorter, but somewhat working MeCP2 protein, resulting in milder symptoms.

This type of mutation was chosen as it may “confer higher risk for MDS-like adverse effects with MeCP2 supplementation,” the researchers wrote.

The team found that providing a healthy MECP2 gene to cells largely rescued behavioral, respiratory, and motor deficits in both male and female mice.

However, MDS-like motor coordination and gait deficits — difficulties in walking — were observed exclusively in females, “suggesting that the underlying motor circuitry is particularly sensitive to MeCP2 dosage in females,” the scientists wrote.

In agreement, the therapeutic approach was associated with a normalization of full-length MeCP2 levels in male mice, but also with about 1.5 times higher-than-normal levels in female mice, “consistent with random X-chromosome inactivation,” the team added.

These findings suggest that gene therapies delivering a full-length MECP2 gene to patients with a R294X mutation may be safe and effective in the rarely affected males, but “careful consideration of risk for adverse motor effects may be warranted for girls and women with RTT,” the researchers wrote.

The data also emphasize “the notion that MeCP2 dosage can have sex-specific effects” and “the importance of using female mice in preclinical studies for RTT,” the team concluded.