Mutations in a gene called MECP2 are the cause of Rett Syndrome. Restoration of adequate levels of MECP2 has been shown to undo the damage caused by a mutated copy of the gene, demonstrating the powerful reach of MECP2‘s influence as symptom after symptom disappeared in fully mature mouse models of Rett Syndrome. This astonishing breakthrough presents us with the urgent challenge of determining whether such results can be achieved in humans.
There are two primary approaches to reversing Rett Syndrome. The first is to understand the function of the MeCP2 protein and to design rational drugs to compensate for its deficit. The second approach is to identify the various outcomes of having an MeCP2 deficiency and screen for anything that ameliorates that outcome. RSRT is pursuing both approaches.
The following are the projects that RSRT is supporting financially as well as intellectually. To date almost $14 million have been committed to these projects.
There is a multitude of data suggesting that mice models of Rett have low levels of a neurotrophic factor called BDNF (brain derived neurotrophic factor). BDNF is a very important and complex protein that is implicated in a variety of disorders. Increasing BDNF in the Rett mice models, either genetically or pharmacologically is beneficial. An FDA approved drug for multiple sclerosis called copaxone (or Glatiramer Acetate) is known for increasing BDNF and therefore of interest in treating Rett.
RSRT has committed to funding an open label study of copaxone in two centers, the Tri-State Rett Syndrome Center at Children’s Hospital at Montefiore in the Bronx, under the supervision of Dr. Sasha Djukic, and at Sheba Medical Center in Ramat Gan in Israel under the supervision of Dr. Bruria Ben Zeev. Each center will give copaxone to ten individuals for 6 months. Below is a comparison of the two studies.
|Title||Pharmacological treatment of Rett Syndrome with Glatiramer Acetate (Copaxone)||An open-label exploratory study to investigate the treatment effect of glatiramer acetate (Copaxone) on girls with Rett Syndrome|
|Principal Investigator||Aleksandra Djukic, MD, PhD||Bruria Ben Zeev, MD|
|Location||Children’s Hospital at Montefiore, Bronx||Sheba Medical Center, Ramat Gan, Israel|
|Objectives||Primary: gaitSecondary: cognition, autonomic function, EEG, quality of life||Primary: EEG improvementSecondary: autonomic function, general behavior, communication, hand stereotyping, feeding, gastrointestinal|
|Study Size||10 girls – 10 yrs old and up||10 girls – 6 to 15 yrs old|
|Dose (injections)||Ramp up to 20 mg per day||Ramp up to 20 mg per day|
|Length of study||6 months||6 months|
While copaxone is not going to cure Rett Syndrome we hope that by increasing BDNF we will see improvements in symptoms. The trials are currently recruiting.
The MECP2 Consortium is a dynamic collaboration between the laboratories of three distinguished scientists, Adrian Bird, Michael Greenberg and Gail Mandel. Made possible by a $1 million gift by RSRT Trustee Anthony Schoener and his wife Kathy, the Consortium was formed to definitively determine how this complex protein, MeCP2, functions and exerts its powerful influence on the human brain. Deep knowledge of its structure and roles in neurological development and maintenance will inform the design of rational treatments for disorders caused by abnormalities in this protein. RSRT has invested $1.8 million in the Consortium thus far.
Three papers to date have been published through the effort of the Consortium. Two papers by the Bird and Greenberg labs deepen our knowledge of the MeCP2 protein and a paper from the Mandel lab opens the door to gene therapy as a treatment option.
Despite almost two decades of research into the molecular mechanisms of MeCP2 function, many questions are yet to be answered conclusively: Is MeCP2 just a methyl-CpG binding protein? Is it a multifunctional protein or primarily a transcriptional repressor? Does it act at specific sites within the genome or more globally? In which cell types is it functionally relevant? The Consortium is tackling these questions through a series of experiments, some of which are described below.
MeCP2 is associated with chromosomes and it is vital to have a detailed picture of exactly where it is bound in different cell types and under various conditions. Chromatin immunoprecipitation (ChIP) has been used to map binding sites genome-wide in whole brain. These studies showed that MeCP2 binds globally and tracks DNA methylation. However existing data does not allow detailed mapping of MeCP2 binding at specific loci due to low coverage. The Consortium is generating a high-resolution map of MeCP2 binding in neurons and glia using mice that express a MeCP2-GFP fusion protein. Experiments will be duplicated using GFP fusion mice with MeCP2 mutations in the methyl binding domain to assess the distribution of MeCP2 without the influence of DNA methylation.
The Consortium will explore how neuronal stimulation affects the binding profile of MeCP2 in the brain and whether the affinity of MeCP2 for methylated DNA changes upon phosphorylation. Mice who are unable to phosphorylate MeCP2 at specific sites will be used. Furthermore, the relationship between activity-dependent MeCP2 phosphorylation and DNA methylation will be explored.
A fundamental question that has received comparatively little attention is how the function of MeCP2 might impact on the cell biology of the nucleus, particularly with respect to nuclear architecture. It is well known that many transcriptional regulators function by interactions with nuclear matrix and associated proteins. This aspect has never been investigated in the context of Rett. The Consortium will determine whether loss of MeCP2 from neurons and glia alters the structure of the nucleus.
Increasing Levels of the MeCP2 Protein
Rett Syndrome is caused by a deficiency of MeCP2 protein. One approach to curing the disorder, therefore, is to restore normal levels. This may be accomplished in a number of ways including small molecule therapeutics (drugs) and/or biologics (gene therapy, protein replacement).
Some treatments may prove to be mutation specific (for example, drugs aimed at restoring normal function to an abnormally truncated protein or drugs that reconfigure misfolded proteins) while others will be more global in nature, such as activating the normal MECP2 gene on the silent X chromosome.
The following RSRT-funded projects are aimed at increasing levels of MeCP2.
A Chemical Genetic Approach for Unsilencing Mecp2
Ben Philpot, Ph.D., Bryan Roth, Ph.D., Terry Magnuson , Ph.D.
University of North Carolina at Chapel Hill
These investigators have successfully employed a high-throughput screen to identify compounds that unsilence the gene for Angelman Syndrome, UBE3A. They are now applying the same approach to Rett Syndrome. Using high throughput robotic equipment they are working their way through a library of 20,000 compounds. To learn more about this work we invite you to watch a video of Dr. Philpot describing this work. RSRT is investing $2.2 million in this project.
A High-Throughput Screen to identify compounds that reactivate the functional MECP2 allele in Rett Syndrome
Jeannie Lee, M.D., Ph.D.
Dr. Lee will develop an assay using mouse cells that can detect activation of the MECP2 gene. Utilizing the significant resources of The Broad Institute the assay will undergo a high-throughput screen to identify compounds able to turn on expression of the gene.
An Oligotherapeutics Approach to Treat Rett Syndrome
Jeannie Lee, M.D., Ph.D.
In 2013 RSRT funded a second project in the Lee lab. While the goal of the new award is the same – activate MECP2 – how she proposes to accomplish it is completely novel. The hypothesis of Dr. Lee’s approach rests on an observation that a group of proteins called Polycomb complexes working in concert with a certain type of RNA, called noncoding RNA (lncRNA) are relevant for keeping genes silent on the inactive X. Dr. Lee’s therapeutic strategy is to awaken the MECP2 gene by disrupting the binding that occurs between the lncRNA and the Polycomb complexes.
Identification of drugs or genes that activate the silent MECP2 gene
Antonio Bedalov, M.D., Ph.D.
Fred Hutchinson Cancer Research Center
The gene that is mutated in Rett Syndrome, MECP2, is located on the X chromosome. All females have two X chromosomes in every cell but one is inactivated. The goal of this project is to identify drugs/compounds and/or genes that will activate the silent MECP2 on the inactive X chromosome.
Reactivation of the Inactive X-linked MECP2 Gene as a Therapeutic Strategy for Rett Syndrome
Michael Green, M.D., Ph.D.
University of Massachusetts
We have identified factors that control X chromosome inactivation and based upon this information derived small molecule drugs that can reactivate the silenced MECP2 gene in cell culture. We will continue to study reactivation of the silenced MECP2 gene and, in particular, extend our cell culture results to mouse models of RTT.
Identification of the mechanisms that silence the MECP2 gene on the inactive X chromosome
Marisa Bartolomei, Ph.D.
University of Pennsylvania
A synergistic project to the project above this effort is also focused on activating the silent MECP2 gene on the inactive X. Dr. Bartolomei seeks to identify the molecular mechanisms that keep MECP2 silent in the hopes that these modifications can be reversed with drug(s). It is possible that activating the silent MECP2 may require a combination of loosening up the mechanisms that keep this gene silent in concert with activation drugs identified in the above project.
Evaluate the outcome of boosting MeCP2 levels in wildtype cells of female mice
Huda Zoghbi, M.D.
Baylor College of Medicine
Females with Rett Syndrome lack functional MeCP2 in approximately 50% of their neurons while 50% have normal (wildtype) MeCP2. Dr. Zoghbi will explore whether boosting MeCP2 levels in the wildtype cells might enhance the overall neural network activity even in the face of the 50% null cells. Encouraging results could set the foundation for a large scale screen to identify targets that can modulate MeCP2 levels. This knowledge will be critical for Rett Syndrome as well as the MECP2 duplication syndrome.
AAV9 Gene Therapy for RTT
Gail Mandel, Ph.D., Adrian Bird, Ph.D., Brian Kaspar, Ph.D.
Ohio State University, Oregon Health Science University
Since Rett Syndrome is a single gene disorder, gene therapy is potentially a viable approach to treatment. Recently the Kaspar lab has reported that Adeno-Associated Virus, serotype 9 (scAAV9), is able to cross the blood-brain-barrier achieving global distribution in the central nervous system when administered via the bloodstream. This collaborative project between Kaspar, a leading gene therapist, and Gail Mandel and Adrian Bird, prominent Rett researchers, will determine the clinical potential of scAAV9 in gene replacement strategies for Rett Syndrome. The Kaspar lab has already been successful in initial trials of SMN gene delivery with scAAV9 for treating a mouse model of Spinal Muscular Atrophy (SMA), completely rescuing the lethal phenotype seen in that disease model. The genetics of the Rett model are ideally suited to investigation of the clinical utility of scAAV9. To learn more about this work we invite you to watch a video of Dr. Mandel and her lab members describing this work.
Adeno-associated Virus-mediated Gene Transfer for the Treatment of Rett Syndrome
Ronald C. Crystal, M.D., Adrian Bird, Ph.D.,
Weill Cornell Medical College, University of Edinburgh
This project also seeks to develop a gene therapy approach to the treatment of Rett but with a different vector than the project outlined above. The vector used will be the new generation AAVrh10 which has shown remarkable gene expression throughout the central nervous system. The Crystal lab is one of only a few in the world that has conducted gene therapy clinical trials in humans. Complimenting the Crystal expertise at gene therapy and clinical trials is Adrian Bird, who brings his deep knowledge of MeCP2 and the Rett animal models. If gene therapy with AAVrh10 is successful in the mice Crystal’s past experience in clinical trial development will prove invaluable.
Alleviation of Specific Symptoms
The array of individual symptoms in Rett Syndrome is so significant that eliminating a single one may, in many cases, dramatically improve quality of life. Finding an FDA approved drug/compound or procedure which ameliorates a symptom (such as disordered breathing, extreme anxiety, seizures) would be the quickest and most cost effective route to clinical trial.
Evaluate the outcome of deep brain stimulation (DBS) on cognition in Rett mouse models
Huda Zoghbi, M.D.
Baylor College of Medicine
The use of DBS has revolutionized the treatment of Parkinson’s and is now also used for depression, OCD, Alzheimer’s and more recently in pediatric disorders such as dystonia and Tourette. The availability of Rett mouse models allows us the opportunity to explore potential benefits of this procedure for Rett. Encouraging data can be quickly moved to the clinic.
Therapeutic Approaches to Reversing Forebrain and Brainstem Abnormalities
David Katz, Ph.D.
Case Western Reserve University
This project will seek to determine whether NMDA receptor antagonists can reverse the imbalance of excitation vs inhibition present in the forebrain and brainstem of Rett mice. If so, certain drugs could be quickly moved into clinical trials. Watch a video interview with Dr. Katz.
Immune modulation as a new therapeutic approach for Rett Syndrome
Jonathan Kipnis, Ph.D.
University of Virginia
Recent data generated in the Kipnis lab demonstrates that immune system and T cells, in particular, are required for normal brain function. T cells, working through soluble factors (cytokines), control synaptogenesis, regulate adult neurogenesis, and affect levels of brain derived neurotrophic factor (BDNF). Depletion or malfunction of T cells correlates with reduced synaptogenesis, decreased levels of BDNF, and impaired brain function. Rett syndrome is associated with motor malfunction, reduced neurogenesis, and deficit in BDNF. Studies indicate that some aspects of T cell function are impaired in Rett patients. Therefore, Kipnis hypothesizes that a malfunction in adaptive immune system (T cells, in particular) in Rett patients contributes, at least in part, to some aspects of disease progression. Thus, if T cell function is improved, the disease can be attenuated and some symptoms might be partially ameliorated.
Learn more about this work.
In vivo screen of drugs and drug-like compounds
Andrew Pieper, M.D., Ph.D.
University of Texas Southwestern Medical Center in Dallas
This project tests FDA approved drugs and compounds of interest in mice models of Rett to identify Hits that improve disease-related signs in the animals. Compounds that alleviate the symptoms or progression of the disease in mice will form the basis of advanced drug discovery programs; approved drugs can be fast-tracked to clinical trials.
mGluR5 Signaling – Is it Implicated in Rett?
Mark Bear, PhD
Dr. Bear is a pioneering researcher whose thoughtful insight has dramatically advanced the understanding of Fragile X. His seminal work was featured in Forbes late last year.
Like Rett Syndrome, Fragile X is a single gene disorder, but caused by mutations in a gene called Fmr1. When Fmr1 is mutated, protein synthesis fails to shut down leading to an excess. Some years ago Dr. Bear proposed that compounds which can block a certain type of receptor, mGluR5 (which triggers the burst of synaptic protein synthesis) might counteract over-expression of protein and thereby cancel out the damaging effect of Fmr1 deficiency. His theory has proved correct and clinical trials of mGluR5 antagonists are currently ongoing at multiple pharmaceutical companies.
Dr. Bear has proposed that just as Fragile X is due to over-synthesis of proteins at the synapse, Rett may be due to under-expression of protein at the same locations. RSRT has committed funding to the Bear lab to first test this hypothesis and if proven correct to explore whether pharmacological manipulations of mGluR signaling will improve any of the mutant mice Rett-like symptoms. The in vivo mouse work will be performed in collaboration with Professor Adrian Bird.
In vivo evaluation of specific drugs in mouse models of MECP2 disorders
Huda Zoghbi, M.D.
Baylor College of Medicine
The Zoghbi lab identified MECP2 mutations as the cause of Rett Syndrome in 1999 and has been very active in the field since. This project is a thorough evaluation of specific drugs which will be tested in the various mouse models of MECP2 disorders: the knockout, the truncated MECP2 and the MECP2 over-expressing mouse lines. Instead of completely lacking the MECP2 gene as in the knockout mouse, the truncated MECP2 mouse expresses the first portion of the MECP2 protein. These animals have milder deficits with a much slower disease progression, and are therefore a model of milder forms of Rett Syndrome. Interestingly, too much MECP2 also causes a severe neurological disease, called MECP2 duplication syndrome. The MECP2 over-expressing mouse carries multiple copies of the MECP2 gene and is a relevant model of MECP2 duplication syndrome in humans. The goal is to speed any promising drugs/compounds to the clinic.
Vitamin D therapy for MeCP2 target Irak1/NF-κB dysregulation
In our previous studies, we identified a number of potential “target” genes of MeCP2 in these important cortical neurons. One of the over-expressed genes is Irak1, a component of the NFkB signaling pathway. We propose to investigate the therapeutic potential of one known inhibitor of NFkB signaling, Vitamin D , in Mecp2 mutant mice. Our proposed experiments will not only add to our understanding of molecular and pathological mechanisms of Rett syndrome, but will also potentially contribute to future therapeutic strategies via Vitamin D or alternative NFkB pathway drugs.
Effect of the serotonin 1a agonist F15599 on respiration and locomotion in a mouse model of Rett syndrome
John M Bissonnette, MD
Oregon Health and Science University
In ongoing experiments we find that the serotonin 1a agonist sarizotan effectively reduces apnea and corrects irregularity in Mecp2 deficient female mice. Sarizotan is a phase II drug that has been evaluated in clinical trials to treat L-dopa induced dyskinesia in Parkinson’s disease.
F15599 is a serotonin 1a agonist shown to be effective in rodent models of depression and cognition. It is an attractive candidate that may correct respiratory disorders without affecting locomotion in Rett mice.
Treatment of Osteoporosis in Murine Rett Syndrome Models: A Comparison of Zoledronic Acid vs Teriparatide on Osteoblast Function, Gene Expression and Bone Mass
Jay R. Shapiro, M.D.
Johns Hopkins University
Osteoporosis leading to fractures affects nearly 40% of young children and adults with Rett Syndrome. The pathogenesis of osteoporosis in Rett is not understood and no accepted treatment regimen exists. This project addresses both of these issues.
Novel insights in MeCP2 function suggests new therapeutic strategies
Stavros Lomvardas, Ph.D.
University of California San Francisco
This experiment leverages the discovery that MeCP2 deficient olfactory receptor neurons (ORN) have a very robust readout; they co-express molecules that are never expressed in the same neuron in wild type mice. Dr. Lomvardas will capitalize on this finding to screen for drugs that can reverse the deficit – in other words find drugs, using high throughput screens (HTS) that can turn off one of ectopically expressed molecular markers.
Identifying Target Genes and Genes that Modify MECP2 Mutations
The development of interventions aimed at genes that MECP2 controls is yet another potential avenue. Recent data, however, suggests that MECP2 may control thousands of genes. Furthermore, these genes may vary considerably depending on the tissue type. It will therefore be extremely challenging to develop treatments if thousands of genes need to be targeted. Nevertheless identifying these genes is the focus of a number of labs and progress in this area may reveal genes worth pursuing.
A very interesting route to consider is that of MECP2 modifier genes. It is likely that differences in the genetic make-up of an individual can modulate the impact of an MECP2 mutation. There are individuals who have common MECP2 mutations and normal X chromosome inactivation but who do not have Rett Syndrome. It is likely that these individuals are protected from their MECP2 mutation due to mutation(s) in other genes. Identifying these modifier genes could open up new avenues for treatment.
Identification of gene modifiers that ameliorate Rett symptoms
Monica Justice, Ph.D.
Baylor College of Medicine
An emerging field of interest is that of genetic modifiers that impact the severity of disease. Rett Syndrome is ideal for modifier screening for two reasons: a) it has been shown to be reversible in mice and b) some girls do not have Rett symptoms despite having common mutations in MECP2. This project aims to identify these advantageous modifiers using a forward genetic screen in mice. The knowledge of disease amelioration may provide novel avenues for therapeutic intervention. In a preliminary screen, five lines that carry inherited suppressors, which increase lifespan and decrease other RTT-related symptoms in mice, have been identified. As expected, most of the suppressors act at the level of chromatin remodeling, and may be difficult to target therapeutically. However, one modifier, Sqle, belongs to the cholesterol pathway and suggests statins may be helpful in Rett Syndrome.
Learn more about this work.
MECP2 Duplication Syndrome Fund at RSRT
Testing reversibility of the MECP2 Duplication/Triplication Syndrome
Huda Zoghbi, M.D.
Baylor College of Medicine
The dramatic reversal of Rett symptoms in mice described by Adrian Bird in 2007 opened the field to questions that now must also be explored in the MECP2 Duplication Syndrome. We know that in Rett, restoration of proper MeCP2 function in mice only days away from death brought them back to health. Would elimination of the influence of excess MeCP2 in the Duplication Syndrome have a similarly dramatic effect? Dr. Zoghbi’s experiment will yield an answer to this very important questions.
Gene Therapy Approach to Treating MECP2 Duplication Syndrome
Kevin Foust, Ph.D.
Ohio State University
The duplication syndrome is caused by having an extra copy of the MECP2 gene. In theory, reducing the amount of MeCP2 protein should improve the disease. Dr. Foust will use adeno-associated virus (AAV) to deliver RNA interference to lower the amount of MeCP2 protein. If successful the project will provide proof-of-concept data showing that MeCP2 reduction is a therapeutic option for patients.