A team of researchers at Baylor College of Medicine and the Duncan Neurological Research Institute (Duncan NRI) at Texas Children's Hospital reports in Science Translational Medicine a potential new approach to 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 , Distinguished Service Professor at Baylor, director of the Duncan NRI and a Howard Hughes Medical Institute investigator. "The disorder primarily affects girls; about 1 in 10,000 live births."
Rett syndrome is caused by loss-of-function mutations in the MECP2 gene, which is key for normal brain function as it modulates the levels of several genes regulating neurological functions. These mutations either lead to loss of the protein or encode a defective protein that cannot fulfill its normal function. Some of the disease-causing mutant MeCP2 proteins are less abundant and/or have decreased DNA binding, an essential function of this protein.
Mouse models of Rett syndrome show that the disorder is reversible – introducing normal MeCP2 protein in the brains of these mice reverses the symptoms. Importantly, researchers have shown that increasing the levels of a mutant MeCP2 protein that retains a little function also improves symptoms, including survival, motor coordination and respiratory abnormalities 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 how the MECP2 gene works prompts a new idea
It's not easy to develop therapeutics that modulate MeCP2 abundance. Too little MeCP2 causes Rett syndrome, yet too much MeCP2 causes a different neurological disorder, MECP2 Duplication Syndrome. Reaching this delicate balance has made it challenging to develop safe and effective treatments.
"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."
Think of a gene as a recipe for a protein. The recipe for MeCP2 has four ingredients: e1, e2, e3 and e4. To make the MeCP2-E1 protein, cells only combine ingredients e1, e3 and e4. To make MeCP2-E2, cells combine all four ingredients, making ingredient e2 unique to this version of the protein. The brain produces both versions, but E1 predominates.
"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."
First, the researchers genetically deleted ingredient e2 from the normal Mecp2 gene in mice and assessed the effect on the protein's abundance and its neurological function. "We were pleased to find that this approach led to 50% to 60% increase of MeCP2 protein in normal mice," Tirumala said.
The researchers then applied the same approach to cells derived from patients with Rett syndrome carrying MECP2 mutations that reduce the abundance and activity of the protein. They deleted ingredient e2 from this mutant MECP2 gene and assessed the effect on the protein's abundance and the characteristics of these cells. "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."
Finally, the team assessed the therapeutic potential of this approach. Would a drug that blocks access to ingredient e2 increase abundance of the MeCP2 protein?
"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."
Other contributors to this work 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. The authors were all affiliated with Baylor College of Medicine, and Duncan NRI when they contributed to this work, but some have graduated and are now affiliated with Stanford University, University of Virginia and UT Southwestern Medical Center – Dallas.
This project was funded 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).
