Rett Syndrome: MeCP2-deficiency and the Potential for Reversibility

Rett Syndrome: MeCP2-deficiency and the Potential for Reversibility

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Dr. James Eubanks, PhD
Senior Scientist, Krembil Research Institute
Division Head, Genetics and Development Program, UHN
Professor, Department of Surgery, University of Toronto
Cross-appointed in Institute of Medical Science and Department of Physiology

By: Parnian Pardis

Imagine your daughter playing in the sandbox one day, laughing and having fun with her brother. Only two days later, your whole world turns upside down: suddenly, your daughter is unable to walk or communicate properly. The onset of Rett syndrome has occurred, a rare disorder which several University of Toronto (UofT) researchers believe may be reversible.

One in every 10,000 girls suffer from this X-linked progressive neurodevelopmental disorder, for which symptoms do not arise until 6-18 months of age.1 Girls are suddenly confronted with developmental stagnation and the onset of symptoms including, but not limited to: loss of communication and motor skills; abnormal hand or eye movements; seizures; and irregular heartbeats.1

While Rett syndrome was recognized in 1983 and mutations in the gene encoding methyl-CpG binding protein 2 (MeCP2) were identified as its cause in 1999,2 this disorder does not yet have a cure nor an effective treatment. Dr. James Eubanks, a Senior Scientist at the Krembil Research Institute of the University Health Network, wants to fill this gap.

Dr. Eubanks received his Doctoral degree at the University of California (UC), San Diego in 1991. Following his postdoctoral training at the Salk institute and Duke University, Dr. Eubanks came to Toronto and was hired as an assistant professor at the University of Toronto in 1994. He now serves as Division Head of the Genetics and Development Program at the Krembil Research Institute and a Professor in the Department of Surgery at the University of Toronto. Dr. Eubanks also holds cross-appointments in the Institute of Medical Science and Department of Physiology.

Dr. Eubanks was interested in the influence of genes and gene products that have a significant function in early brain development. He used MeCP2 as a reference sample while investigating the potential role of specific methyl DNA-binding factors in influencing brain sensitivity in epilepsy and stroke. The discovery in 1999 that identified MecP2 as cause of Rett syndrome2 led Dr. Eubanks to focus on understanding the impact of MeCP2 on brain development and function, as well as the consequences of its absence on brain activity.

To paraphrase a colleague, Dr. Janine LaSalle from UC Davis, Dr. Eubanks likened the brain to a huge orchestra and MeCP2 to its conductor. Members of the orchestra may be able to play their instruments when the conductor is absent, but we cannot actually hear the melody. So, we can’t appreciate the music. MeCP2 thus governs nervous system activity to generate a homeostatic balance and keep the system in normal tolerance range. With this understanding, it is clear that future advancements targeting MeCP2 deficiency in mice are crucial to improve the quality of lives of patients with Rett syndrome, and ideally, to eradicate the syndrome as a whole.

Dr. Eubanks and his team have started to think about how to rescue the phenotype

of MeCP2-deficient mice. Fortunately, a colleague in Scotland had generated a number of mutant mouse lines that either 1) completely lacked MeCP2, or 2) expressed an allele to allow a silent MeCP2 to be reactivated using a Cre-lox system. To identify whether therapeutic improvement was possible in patients with Rett syndrome, Dr. Eubanks’ team devised strategies to reactivate the non-functional MeCP2 gene in these genetic models at very symptomatic stages of development, and conducted subsequent behavioral analyses. “To our pleasure, and somewhat surprise, the mice got a lot better. Dramatically better,” said Dr. Eubanks.

Videos of a mouse prior to, and three months after, MeCP2 reactivation demonstrated exactly this improvement. Pre-treatment, the mouse moved with difficulty, experienced seizures and
tremor, and had trouble breathing. Three months later, the diagnosis of that same mouse was remarkably improved. The mouse still experienced a hobble while walking, but could now move freely and lived on average three times longer than its untreated litter mates. The previously attenuated cognitive abilities in this mouse had recovered to levels found in wild-type mice. Previous groups in Scotland had shown positive outcomes following reintroduction of the non-functional MeCP2 gene at less symptomatic stages, but this finding was the first indication that the problems associated with several neural systems could be rescued and set the foundation for why Dr. Eubanks thinks Rett Syndrome is reversible.

“We should be able to develop something that can have a similar effect in patients as what we see in mice. The unfortunate part is that the strategies used in mice have no clinical applicability—you can’t make that system in patients. So we have to come up with something different, and that’s what we’re doing now,” Dr. Eubanks explained.

There are several paths to make these findings clinically relevant. Ideally, researchers can identify the cure, the “moonshot” drug that can restore MeCP2 functionality. Dr. Eubanks acknowledges the possibility of this based on the type of mutation present. For example, in the case of a nonsense mutation, a mistake in the genetic sequence for the MECP2 results in a premature termination codon and halts protein synthesis. However, effective drugs will trick the cell to skip over a nonsense mutation and reach the end of the normal protein coding sequence. Searching for a drug with high fidelity and low toxicity, Dr. Eubanks and Principal Investigator, Dr. Alan Kozikowski, from StarWise Therapeutics, have received a Small Business Innovation research grant to experiment with this further.

The second approach is pharmacological therapy—targeting deficits that have already been identified as a consequence of MeCP2 deficiency. Dr. Natasha Shulyakova did exactly this. During her doctoral training with Dr. Eubanks, she questioned whether absence of MeCP2 causes increased oxidative stress in the neural system, and whether too much reactive oxygen species has progressive deteriorative consequences. In vitro, she was able to show that an antioxidative strategy implemented to improve mitochondrial efficiency decreases the level of oxidative stress and helps restore diminished ATP levels.

Taking her findings one step further, this same antioxidative “cocktail” significantly improved various performance indications in MeCP2-deficient mice.

“They had better ambulatory activity, their balance was a bit better, and their social interactive behavior improved. Their epilepsy activity didn’t change, so it didn’t help everything, but it helped a number of these properties,” remarked Dr. Eubanks. Gathering interest, Dr. Shulyakova and Dr. Eubanks worked with a team to take her results and put them into a clinical trial. Two years later, having completed all regulatory requirements and manufacturing the necessary cocktail, recruitment for this clinical trial commenced in April of 2018.

Finally, gene editing, gene therapy, and X chromosome reactivation can play a large role with further developments in technology. Coupling these techniques, mutations have been corrected in cultured cells. The difficulty that remains is translating these findings in vivo. Ultimately, “you can’t do it all—you’d like to do it all—but you really have to do what you think is best and most likely to give returns now. And let basic science identify the things that will give returns tomorrow,” says Dr. Eubanks.

Funding from CIHR, Ontario Brain Institute, Epilepsy Canada and the Canadian CDKL5 Network allows Dr. Eubanks to actively pursue the ideals of the Institute of Medical Science—bench to bedside. While he may have a lot of ideas, Dr. Eubanks humbly recognizes students as the driving force for his ongoing studies. “Often, it’s the job of the student to show what isn’t going to work… something that may not be recognized is how important the ‘it didn’t work’ turns out to be for me sitting here. It’s frustrating for the student, but in many ways [it’s] essential for sculpting what is the right direction,” says Dr. Eubanks.


  1. Chahrour M, Zoghbi HY. The story of Rett syndrome: from clinic to neurobiology. Neuron. 2007;56(3):422-437.
  2. Amir RE, Van den Veyver IB, Wan M, et al. Rett syndrome is caused by mutations in X-linked MECP2, encoding methyl-CpG-binding protein 2. Nat Genet. 1999;23(2):185-188.