Boosting a key brain protein could help treat Rett syndrome

"Rett syndrome is a rare genetic neurodevelopmental condition that causes a regression in development, typically after 6 to 18 months of normal growth, leading to severe impairments in motor skills, speech and communication," said corresponding author Dr. Huda Zoghbi, director of the Duncan NRI, Distinguished Service Professor at Baylor, and a Howard Hughes Medical Institute investigator. "The disorder primarily affects girls; about 1 in 10,000 live births."

How MECP2 Mutations Disrupt Brain Function

Rett syndrome results from loss of function mutations in the MECP2 gene. This gene plays a critical role in the brain because it regulates the activity of many other genes involved in neurological processes. When the gene is altered, the resulting MeCP2 protein may be missing entirely or unable to function normally. In some cases, mutant forms of MeCP2 are produced in smaller amounts or have reduced ability to bind DNA, which is essential for carrying out its role in controlling gene activity.

Experiments in mouse models have shown that Rett syndrome symptoms can be reversed under certain conditions. When healthy MeCP2 protein is introduced into the brains of these animals, their symptoms improve. Researchers have also found that increasing the amount of a partially functional mutant MeCP2 protein can lead to improvements in survival, movement, and breathing problems in mice.

"This is important because about 65% of patients with Rett syndrome have partially functional MeCP2 that either has decreased DNA binding or is less abundant than normal," said first author Harini Tirumala, graduate student of molecular and human genetics in the Zoghbi lab. "Working with mouse models and cells derived from patients with Rett syndrome, our study provides proof of concept that increasing the levels of mutant MeCP2 in patients with the condition could provide therapeutic benefit."

Understanding MECP2 Protein Variants

Developing treatments that adjust MeCP2 levels is challenging because the brain requires the protein to stay within a narrow range. Too little MeCP2 leads to Rett syndrome, while excessive amounts cause another neurological disorder known as MECP2 Duplication Syndrome. Achieving the right balance has been a major obstacle for therapy development.

"We knew from previous studies that the brain normally produces two slightly different versions of the MeCP2 protein, known as E1 and E2," Zoghbi said. "These versions come from the same gene, which is processed one way to produce E1 and a different way for E2."

A useful way to picture this process is to think of the gene as a recipe for building the protein. The instructions contain four components: e1, e2, e3 and e4. To make the MeCP2 E1 protein, cells combine e1, e3 and e4. To produce MeCP2 E2, cells include all four components, meaning the e2 segment appears only in the E2 version. The brain produces both proteins, but E1 is the more abundant form.

"We also knew that there have been no reports of Rett syndrome patients carrying mutations on E2 protein. Only mutations that disrupt E1 protein cause the condition," Tirumala said. "Studies in mice support this observation."

"Altogether, we knew that MeCP2-E2 differs from MeCP2-E1 by a single ingredient in the gene, is less abundant than E1, is not associated with Rett syndrome and is not needed for MeCP2 function in the brain," Tirumala said. "This led us to hypothesize that guiding brain cells to skip the e2 ingredient would promote the production of more MeCP2-E1 protein in patients with Rett syndrome and improve disease outcomes. We tested our hypothesis in mice and in cells derived from patients with Rett syndrome."

Increasing MeCP2 Protein in Experiments

To test the idea, scientists first removed the e2 segment from the normal Mecp2 gene in mice and examined how this affected protein levels and neurological function. The change significantly increased MeCP2 production.

"We were pleased to find that this approach led to 50% to 60% increase of MeCP2 protein in normal mice," Tirumala said.

The team then applied the same strategy to cells taken from patients with Rett syndrome who carry MECP2 mutations that reduce protein levels and activity. By deleting the e2 component from the mutant gene, the researchers evaluated how the cells responded.

"We were excited to see that deleting ingredient e2 enhanced MeCP2 production," Tirumala said. "Importantly, depending on the severity of the mutation, these cells recovered part or all of their normal structure, their normal electrical activity and their ability to regulate the levels of other genes."

Testing a Possible Therapeutic Approach

Researchers also explored whether a drug could be used to block the e2 segment and boost MeCP2 production.

"We tested the value of morpholinos to enhance the production of MeCP2 protein in mice," Tirumala said. "Morpholinos are synthetic molecules designed, in this case, to prevent the production of MeCP2-E2 protein by blocking the access to the e2 ingredient," Tirumala said. "It was exciting to see that our morpholinos significantly increased MeCP2 protein in mice."

"Our work lays the foundation and provides preclinical evidence for a therapeutic approach for Rett syndrome that increases MeCP2 and confers functional improvement," Zoghbi said. "Although morpholinos themselves are not an option because of their toxicity, similar strategies, like antisense oligonucleotide therapies already used in other conditions, could potentially be developed for Rett syndrome."

Study Authors and Funding

Additional contributors to the study include Li Wang, Yan Li, Sameer S. Bajikar, Ashley G. Anderson, Wei Wang, Alexander J. Trostle, Mahla Zahabiyon, Aleksandar Bajic, Jean J. Kim, Hu Chen and Zhandong Liu. All were affiliated with Baylor College of Medicine and Duncan NRI during the research, although some have since moved to institutions such as Stanford University, University of Virginia and UT Southwestern Medical Center -- Dallas.

The research was supported by the National Institutes of Health (grants 5R01NS057819, P30 CA125123 and S10OD028591), the Howard Hughes Medical Institute, National Institute of Neurological Disorders and Stroke (F32NS122920), the Henry Engel Fund and the Eunice Kennedy Shriver National Institute of Child Health and Human Development (P50HD103555).