If you follow Rett research closely (or maybe even loosely), this will be a drug name you’re familiar with. My Google analytics tells me it’s one of the most popular searches currently bringing people to this blog so I know you’re out there and looking for information about this potentially groundbreaking new medication and the implications it may have for Rett syndrome.

NNZ-2566 is a medication being developed by the small pharma company Neuren. Neuren is a company in New Zealand which has offices in Australia and the United States. They have special interests in areas of neuroscience where there are gaps in development of treatments. Neuren currently has a growing portfolio on traumatic brain injury, neurodegenerative disorders and neurodevelopment disorders.

There are very few human clinical trials actually going on for Rett syndrome and NNZ-2566 is one of them. The trials are happening in America as we speak.

[What]

So the first question I aim to answer for you is “what is it?” We have a protein prevalent in our bodies called IGF-1. Its function is to help cells grow. It is an important component in the actions of the growth hormone in all of our bodies. In the brain, it helps cells to divide and make synaptic connections from neuron to neuron. However, people with Rett syndrome have issues with their IGF-1 because MeCP2 affects its expression. Now, where NNZ-2566 comes into this mix is this: by looking at the IGF-1 protein and which bits of the whole package could be beneficial in treating Rett, scientists have found that the first 3 amino acids of IGF-1 are what’s needed.

IGF-1 does not readily pass from our circulation into the brain. However, a small piece of IGF-1, called a tripeptide, is able to pass from the blood and into the brain. NNZ-2566 is a synthetic version of the tripeptide that has been modified so that it can be absorbed orally, passing from the digestive tract into the circulation and then into the brain.  Evidence from the laboratory suggests that the tripeptide is the part of IGF-1 that seems to be effective in treating Rett syndrome. A trial of NNZ-2566 is underway in adolescents and adults with Rett Syndrome at Baylor College of Medicine and the University of Alabama. A trial of IGF-1 in young girls is underway at Boston Children’s Hospital.

[How]

NNZ-2566 is delivered as an oral, strawberry flavored liquid whereas IGF-1 has to be injected.

[Contraindications]

Just like any drug, IGF-1 isn’t suitable for everyone. When growth hormones are given after puberty, they can cause continued bone growth so there’s a window of time when this could be administered as a treatment. However, NNZ-2566, which is a peptide and not the whole protein, does not appear to have those same issues. The clinical trial with NNZ-2566 is designed to show that it is suitable for teenagers and adults with Rett. This means NNZ-2566 could have the potential to be suitable for anyone with Rett at any period of their life, something that no other suggested treatment we’ve ever seen can claim.

[The Benefits]

You may remember a while back, I posted about gene therapy and the post said that, while treatment with vectors is an interesting idea, there are great challenges. That these methods were like using a shotgun when what we need is a sniper rifle.

Well, IGF-1 is like a sniper rifle. Through the current trials, scientists are hoping to deliver more growth enhancers into the central nervous system which gets right to the central problem immediately, bypassing the need to deliver a new gene with a vector and hope it is expressed to correct the defective MeCP2.

But of course, I’ve already mentioned that IGF-1 needs to be injected while NNZ-2566 is an oral medication to hopefully deliver the exact same results and be potentially suitable for  any age.

[The Challenges]

Like every current suggested method for the treatment of Rett, there are a few challenges to face.  The fundamental challenge with NNZ-2566 is that it is new drug, and new drugs have to go through a lot of rigorous testing before they can be used on all age groups.  Since NNZ-2566 was developed by Neuren for traumatic brain injury, it has had a sizable amount of safety testing already completed and has been shown to be generally safe and well tolerated by adult human subjects. However, this fact does not remove the need for safety evaluation in Rett patients.

[Current Studies]

NNZ-2566 is currently being studied in human clinical trials for both Rett syndrome and traumatic brain injury. Both conditions have something in common in that there are weak synapses in the brain. A study in Fragile X syndrome is scheduled to begin shortly.

Phase 1 (the safety tests) have been completed. We know that it’s a well-tolerated drug.

Phase 2A began in May 2013 and hopes to end in September this year. These are currently being undertaken at Baylor College of Medicine with Drs. Dan Glaze and Jeff Neul and at the University of Alabama – Birmingham with Dr. Alan Percy. This trial consists of 60 participants between the ages of 16-45.

The Phase 2 study examines the safety profile, looking for unexpected adverse changes, deterioration or anything that wouldn’t be expected in the natural progression of Rett syndrome. This is where the ongoing Natural History Study adds value. Unless we study Rett in the long run and learn what happens and the effects we can expect, it’s difficult to measure the effectiveness of drugs we want to trial for Rett. The data extracted during the Natural History Study gives us baselines of what to expect so that during these types of trials, we can know what would normally have been expected and gives scientists a greater base of knowledge for use when monitoring such trials.

This phase is also measuring efficacy (effectiveness).

Based on the tests with mouse models, we’re hoping to see improvements in breathing and cardiac function, and decreases in seizures. The patients are also being observed for changes in many other behaviors.

[The Next Step]

The next step for NNZ-2566 will be proof of concept studies in the pediatric population. Remember, the youngest person in the current study is 16 years old because the FDA guidelines are such that a drug usually cannot be tested on juveniles until the trial has first been successful with adults (when adult patients are available).

[Funding]

This is where partnerships are incredibly important. IRSF started the Baylor study with a $600,000 ANGEL grant to get it going, but this doesn’t even come close to covering the cost. Partnership with Neuren is extremely important to bring more resources to the table for these trials.

[Considering a Trial?]

It’s very important to fully understand the implications of putting your child into a clinical trial. Please view this document from IRSF on safety in Rett Syndrome trials.

This post was made possible by contributions from:

• Steve Kaminsky, Ph.D. – Chief Science Officer, IRSF
• Janice Ascano, Ph.D. – Manager of Grants and Research, IRSF
• Nancy Jones, Ph. D. – Senior Director, Clinical Development and Medical Affairs at Neuren Pharmaceuticals

Note: This post may be easier to understand if you read this one first.

In the summer, RSRT published an announcement entitled “Statins Suppress Rett Symptoms in Mice“. You know me, always here to help break these things down in plain English. I know that title looks pretty bold and exciting. And it is! But possibly not for the reasons you think.

[The Study]

The study was carried out in Monica Justice’s lab. It was funded by NIH, RSRT, IRSF, and the Autism Science Foundation.

[The Mice]

In this study, male mice with Mecp2 mutations were inundated with the carcinogen ENU to force mutations in other genes. The mice were then observed to see which ones were doing better with their Rett symptoms. Five particular mice appeared better than the others. In this paper they describe one of these mice; the one that had a mutation in a gene within the cholesterol metabolism pathway. When that gene (called ‘Sqle‘) was mutated, the Rett mice did better.

[Statins]

From this study, it would seem that modifying cholesterol pathways by mimicking the outcome of a Sqle mutation may be a positive thing for girls with Rett. One cannot simply mutate SQLE in humans. So this is where statins come in.   Statins are a group of drugs that modify cholesterol pathways. This is definitely not a case of going and telling your Dr. your daughter should have statins. Now, statins need to be studied even further in mice with Mecp2 mutations. What statin? What dosage? What are the long term effects?  THEN, clinical trials should be undertaken in order to answer those questions in humans.

[Backing up…]

But first, let’s back up and discuss why experiements like this are being done and address a couple questions you may have.

Firsly, why are these experiments usually done on male mice? Good question! Although we know that Rett syndrome happens almost exclusively to females, we know of a few cases where a male is diagnosed. However, it’s a far worse ordeal for the male genome than the female, so males don’t usually live long. Because of the exponentially increased severity of Rett in males, it’s faster and easier to do these experiments on male mice because the success or failure is faster to determine.

To quote Steve Kaminsky of IRSF, “The phenotype is much easier to follow in male mice. They die much sooner, so if you can extend their life, BOOM you have a result. It would take a lot more work to observe changes in females. We scientists are always looking for the cleanest test tube and male mice are that for us.”

A second question you may be asking is why are we looking at other genes when we already know the gene that causes Rett? There are three ways we can seek to address Rett syndrome:

1. Fix the mutated gene MECP2
2. Identify the partner genes
3. Hit the downstream targets or pathways of the MeCP2 protein.

These sorts of studies address that second point. This study sought to address the possibility that, although we can’t yet reverse Rett, perhaps we can find other genes with a relationship with MECP2 that can aid in the recovery of symptoms. The challenge is understanding what pathways we can work with to get around MECP2 mutations. In this study, it would appear that modifying cholesterol pathways may help circumvent the MECP2 problem.

What this paper shows is that cholesterol metabolism has a partnership with MECP2 in some way shape or form. That partnership isn’t known at this point. There are certain parts of the pathways that crossover or are partners with the MECP2 pathways. By using statins, you can change some of the patterns of cholesterol pathways, perhaps them enhancing the MECP2 pathways. Essentially building a serogate carrier to help MECP2 do what it does.

Steve Kaminsky explains it this way, “You may not need a 100% gene reversal or modification to treat Rett syndrome. If we can make incremental increases in the synapses found in Rett with different methods, different cocktails of drugs, each different treatment may result in a percentage of recovery. Like this: what if we only needed to recover the MECP2 gene 60% in order for a girl or woman with Rett to function with speech, hand use…all the things that are difficult at present. What if one compund (like a statin) could recover 10%, 15%  or 20%? And another drug the same? Each different treatment may result in a percentage of recovery and work together to treat the symptoms of Rett syndrome.”

However, there’s a caution here.  We all know Rett syndrome manifests with a range of severity of different symptoms.  Therefore, not all forms of Rett will be helped by a treatment such as this.  At this time, we cannot predict who will benefit from Statins.

[How to take it]

So how do we take this announcement that statins may arrest/treat/cure some of the symptoms of Rett syndrome? We believe that this paper tells us that:

• There’s a relationship between the cholesterol pathway and MECP2
• The study also reveals that there are four other genes that have these similar partner relationships (this announcement is focused on the Sqle gene, the one related to cholesterol).

Genetic mutations aren’t good things to have (as we all know so well). But in this case, it seems that the Sqle gene being mutated in a mouse with a mutation on Mecp2, that’s a good thing. What would happen to someone with only a mutation in Sqle and nowhere else is another story. Janice Ascano, PhD, manager of grants and research for IRSF, put it in perspective really well when she told us, “Basically, it’s a case of two wrongs make a right.”

We’re not going to go try to mutate SQLE in people with Rett. But there are compounds out there (statins) that downregulate those other pathways. Downregulating Sqle may create a more even playing field for the two genes. And we’re definitely not going to go and start administering statins to girls with Rett. Steve Kaminsky said it oh-so-well when he told us, “People think ok where do I sign up? You don’t suddenly start administering a statin as a result of a mouse study. Initial discovery now needs to be translated into preclinical data that makes sense to the FDA. Simply making this observation doesn’t mean that we can immediately go and deliver it to a Rett patient in a safe way. This is a great discovery that has a tail of translational research tacked onto it. Now to use that discovery data to complile preclinical data to take to the FDA. That process can take some time. What’s good about this story is that there are a lot of statins so we can move backwards and start looking at drugs already approved and start the translational research of testing it in a mouse. People take statins every day to help control their metabolism.”

[What now?]

Well, as said previously, this just means that there’s an open door to exploring the possibilities that statins may have other medicinal values than simply modifying cholesterol metabolism. We’ve discovered that this other gene affects something. Now to do all the homework that goes with that. An exciting thing to take away from this is the idea that other scientists who work with cholesterol could now jump into the Rett scene. They just need access to the mice.

This post was made possible by contributions from:

• Steve Kaminsky, Ph.D. – Chief Science Officer, IRSF
• Janice Ascano, Ph.D. – Manager of Grants and Research, IRSF
• Kori Coates, Executive Director, Cure Rett
• Paige Nues, Director of Family Support, IRSF

by Elizabeth Halford, with contributions from Steve Kaminsky, Ph.D., Janice Ascano, Ph.D., Paige Nues, Monica Coenraads and Kori Coates.

As of late, there’s been much buzz about gene therapy as a potential treatment  for Rett syndrome. And for good reason! Gene therapy isn’t new. In fact, it’s been used to treat a number of disorders, among them: retinal disease, acute lymphocytic leukemia (ALL), multiple myeloma and Parkinson’s disease.

Dr. Gail Mandel said in a recent interview with RSRT that one good reason to consider it for Rett syndrome is that “…the genetic studies pioneered by Adrian Bird showed that it’s a reversible disease, even in late symptomatic stages in mice. So that suggested that you could put back a good copy of MECP2 and possibly reverse or maybe stabilize or just improve by putting it back.”

Currently, there are tests going on to see exactly how we can whip the MECP2 gene into shape, change the biology of Rett syndrome and reverse its symptoms. Some exciting advancements have been made so today, I’d like to bring some clarity on the topic and tell you what I know (and more importantly, what I don’t yet know) about gene therapy.

[the first study]

In 2013, a paper published in Molecular Therapuetics first discussed the possibility of using a virus, called AAV9, to do gene replacement in Rett mice. Essentially, they used AAV9, an altered version of the common cold virus, to encapsulate healthy Mecp2 and sneak it into the brain cells. In this study* scientists Stuart Cobb, Steven Gray and their teams “demonstrated reversibility of RTT-like phenotypes in mice. This suggests that MECP2 gene replacement is a potential therapeutic option in patients.” and that “These results support the concept of MECP2 gene therapy for RTT.”

The study reported improvements in motor function, tremors, seizures and compulsive movements.  One symptom which didn’t respond to the therapy was abnormal breathing.

They concluded the paper by saying that this work shows, at “proof-of-concept level”, that MECP2 can be delivered at a tolerable level via AAV2/9 vectors (those viruses we mentioned before) to the brain of Mecp2-null mice. Longer-term studies in female mice are the next steps before any human application can be considered.

[the second study]

So that first study into gene therapy for Rett syndrome concluded that, yes, it is possible that a virus could deliver healthy MECP2. But it ended with the next step being the need to now test in female mice. Remember, even though Rett mainly happens in females, these studies usually start with male mice.

Many people in the Rett community will have seen, read or heard about recent announcements about gene therapy studies being successful in female mice. The paper, published in the Journal of Neuroscience, reports that the study**, led by Molecular biologist Gail Mandel, of Oregon Health and Sciences University, used a similar virus as the first study. The same results were seen as in the first study, including non-reversal of breathing problems. What was different about this particular study (other than the fact that it was done using female mice) is that a 10-15% uptake of MECP2 in the brain was achieved to get a reversal of symptoms as opposed to only 5% uptake in the first study.

This revelation very much lines up with something Dr. Steve Kaminsky told me recently when he said that “You may not need a 100% gene reversal or modification to treat Rett syndrome. Perhaps if we could make incremental increases with different methods and different cocktails of drugs. For example, what if MECP2 gene therapy could incrementally improve  function with movement and hand use? Then, what if a compound (like a statin) and a second drug could improve each by an additional 10%, 15% or 20%? Each different treatment may result in a percentage of recovery but together will work in synergy to treat the symptoms of Rett syndrome.”

These gene therapy studies are examples of discovery based research and there is still a lot of work before they can be transferred to the clinical environment.  So to this end we also need to be pursuing immediate clinical research to change the biology associated with Rett syndrome.  Monica Coenraads, Executive Director of RSRT said recently in an interview with Dr. Mandel that, “I say this to families often: we can work on downstream targets – BDNF, IGF-1, Chromatin remodeling, a whole long list of drugs. They’re not gonna cure Rett syndrome. They may improve one or two symptoms which, you know, because the girls have so many symptoms, we would take any improvement in any symptom. But if we really want to see the dramatic cure that everybody talks about, for that I think we need to address the underlying genetic problem.” In the same interview, she also tell us, ”I think it takes a lot of different perspectives and ideas and ways of looking at a problem because this is a complex protein and a complex problem.” To that end, she agrees that we should also be looking into other avenues for treatment of Rett. She told us, ”It’s important to pursue as many approaches as possible since we don’t know what will pan out” and that “Activating the silent MECP2, gene modifiers, downstream targets are all on the table. As is continued basic science to better understand what MECP2 does in the hopes that it will inform us on potential new strategies.”

[the challenge]

Although these experiments are super, there are substantial challenges with using these methods to deliver MECP2 to brain cells. One being that the virus hits the normal neuron and the mutant neuron. That’s like hunting with a shotgun. It’s going to hit everything. If AAV9 could be designed to recognize the mutant and leave the healthy neuron alone, that’s hunting with a sniper rifle. In the Rett patient, the body of the neuron is smaller. In these studies, it was shown that the body of the neuron grew, but that also happened in the healthy cells, not just the mutant ones. If the efficiency of uptake is increased, could we see over expression of MECP2 in the normal neuron and would that induce a MECP2 duplication syndrome in those cells? We just don’t know, and this will be a large challenge for any regulatory agency approving these therapies in the future.

We want to leave the healthy neuron alone and hit the mutant one. That’s science fiction right now. No one can do that. The viruses, as they are now, hit everything. And this is why gene therapy is so hard with Rett – half of the cells are normal, half are mutant.

Yes, gene therapy works for other disorders, but that’s because all the cells are affected so a shotgun is your weapon of choice. What the FDA would want to know before testing on humans is what are you doing to the pathology of the normal cells? As the data stands now, the FDA would say that we’re nowhere near ready. This is a whole new area for the FDA and because of the deaths that have been associated with gene therapy along the way, gene therapy has a bit of baggage that goes along with it. The Biologics Division at the FDA is very finely tuned and, according to Dr. Kaminsky, “we can’t even begin to talk to them until many of the basic challenges associated with using AAV vectors with Rett syndrome models have been worked out. So we still have a steep hill to climb, but it is worth climbing.”

[what next?]

Gail Mandel cautions at the end of the recent report that (according to this post from Autism Speaks) ”…important steps – including safety studies – remain before this gene therapy is ready for clinical trials with human volunteers.” That means more mice studies.

So now we’ve proven that:

1. Gene replacement using a virus is possible in male mice
2. Gene replacement using a virus is possible in female mice

What’s left to discover? Lots. And lots.

We are still at the fundamental basic stage of this science. Dr. Kaminsky tells us that “we are still 5-10 years away from having vectors that can do the things we’d like to do. In the meantime there are many other approaches to Rett that could have immediate effects on Rett biology. So we are targeting the downstream pathways that could bring immediate change for Rett patients and improve their quality of life.  I think all of this is great conversation. It’s wonderful research by  Stuart, Steven and Gail, but in the meantime we’re moving forward with known drugs to see if we can get incremental improvement in our girls.”

I personally believe in pursuing both paths. A dramatic cure would be ideal and there’s science working on that. But what happens when/if we put all our eggs into the gene therapy basket and the human clinical trial (which is many years away) possibly fails mid-study? Even in the final stages of trials, we often see failure. As in this heartbreaking story about a drug trial for Fragile X syndrome.  In the case of gene therapy trial failure, we would be taken backwards at least a decade if we didn’t have other studies in place. I support the science of looking at downstream targets, testing existing drugs and gene therapy. It’s all good.

[in conclusion]

I think the most inspirational view of gene therapy for Rett syndrome came to me in conversation with Dr. Kaminsky while preparing this post. He told me, “Gene therapy is a very distant star but hopefully we will be able to navigate to it. A number of scientists looking into this. A number of organizations are funding gene therapy, but it’s a distant star so we need to take multiple approaches. If we hang our hat on this one approach, a single failure means we’re taken back a decade. When playing chess, you have multiple ways you can move across the board with multiple strategies in place. With these two papers we are light years closer to that distant star.”

* The work was supported by grants from the International Rett Syndrome Foundation (SJG), from the Medical Research Council (SRC; grant no. G0800401) and from the Rett Syndrome Association Scotland (SRC/MESB)

** The work was supported by the Rett Syndrome Research Trust, the National Institutes of Health (to G.M.), the Wellcome Trust (Grants #077224 and #091580), Medical Research Council, UK (Grant #G0800401), Action Medical Research in association with the Henry Smith Charity, and the R.S. MacDonald Charitable Trust (Grant #SP4443 to A.B.)