In June the 10th Annual Spinal Muscular Atrophy Research Group Meeting was held in Montreal, Canada. Over 160 researchers and clinicians attended. Scientific sessions were held on the following topics: Clinical Trials, Outcome Measures, Stem Cell Therapies, SMN Functions, and Drug Discovery Programs. Over 80 presentations were given. A detailed overview of each session is provided below.

We would like to take this opportunity to highlight several particularly relevant research developments separately. These include the development of novel clinical trial outcome measures for both Type I and ambulatory Type III patients, which will allow future trials to enrol additional SMA patient types. On the basic research front, one the most interesting highlights was new data presented by multiple labs indicating that SMN has a specific function in neurons and that the role of SMN in the cell body is likely different than its role in motor axons. This is important because delineating and understanding the importance of the specific role of SMN in axons will facilitate more strategic design of drug discovery screens. Drs. Hans Keirstead and Doug Kerr also gave exciting talks on stem cell therapies for SMA. Dr. Kerr showed that mice motor neurons derived from mouse stem cells send axons out of the spinal column into the periphery when injected into the spinal cord. These motor axons can form functional connections on muscles, which provided clinical improvement in mice with motor neuron disease. Moreover, Dr. Keirstead reported good progress on developing stem cell-derived motor neurons for human trials. Finally, there were a number of talks focusing on drug discovery efforts to find novel small molecule therapies for SMA. Talks on this topic were given by representatives of the labs of deCODE Chemistry, Christina Brahe, Brunhilde Wirth, Brent Stockwell, the NINDS SMA project, and Trophos. The initiative on up-regulating SMN levels in SMA that is closest to the clinic was given by deCODE Chemistry.

Several SMA researchers presented detailed research updates at the Family and Professional Conference in San Diego this July. Talks were given by Dr, Jill Jarecki on FSMA- funded Research Activities, Dr. Hans Keirstead on Stem Cell Therapies for SMA, Dr. Klemens Hertel on RNA-Mediated Therapies for SMA, Dr. Arthur Burghes on Basic Research News form Montreal, Dr. Ching Wang on the Hydroxyurea Clinical Trial, and Kathy Swoboda on Project Cure and the Carni-Val Clinical Trial. The summary of this session will be included in the next issue of the FSMA “Research Compass”.

Clinical Trials

A number of randomised clinical trials are currently ongoing along with some open label studies. The EUROSMART study being run in Germany, Israel, Italy, Poland, Spain, Turkey has now been completed. Dr. Basoglu from Ankara presented the results of this randomised Spinal Muscular Trial comparing acetyl-L-carnitine to placebo in Type II and III patients above 4 years old (Ages 4- 57 years old, n=59 female, n=51 male, Type II n=52 and Type III n=58) . Though some data presented suggested that acetyl-L-carnitine might have helped in the delay on deterioration of the patients no overall significant benefit was shown. Both groups (placebo and medication) showed a mild improvement in muscle strength but a decrease in lung function was observed in both groups. The study had an overall dropout of 14 patients. Though no significant benefit was shown these data are important for other studies ongoing, especially the study of sodium valproate in children over 2 years of age. In all studies with sodium valproate there is a need to protect the patient from loss of carnitine, so the EUROSMART data is very relevant to future studies, as carnitine needs to be co-administered.

Dr. Swoboda reported on two Project Cure SMA trials assessing sodium valproate in SMA patients greater than 2 years of age. In an open label study (n=40 patients), Project Cure SMA assessed tolerability and toxicity of sodium valproate. It was shown that valproate was well tolerated in these children, when given in combination with Carnitine. Only four children dropped out of the study: two due to excessive weight gain and two due to lack of perceived benefit after 6 months. The modified Hammersmith scale was used to assess efficacy. An average increase of 6 points was observed after 12 months, and a 3-point increase is viewed as a clinically relevant change. This data was further assessed by dividing children into two groups: children over 5 years of age and children under 5 years of age. Overall, both groups considered together improved on the modified Hammersmith Score in motor function as discussed above. However, the most notable improvement was seen in children less than 5 years of age. 93% of children under 5 showed a significant improvement in Hammersmith score, while only 37% of children over the age of 5 showed a significant improvement.

While the open label nature of this study does not allow definitive conclusions to be drawn, the encouraging results suggested that a randomised placebo controlled study was warranted. Currently, Project Cure SMA is conducting a randomised placebo controlled study to test a combined regimen of Valproate and Carnitine. This is multi-center study with testing sites in Utah lead by Dr. Swoboda, Ohio lead by Dr. Kissel, Wisconsin by Dr Schroth, in Maryland by Dr. Crawford, Michigan by Dr. Acsadi, and Montreal by Dr. D’Anjou. This study consists of two distinct cohorts. The first is for children ages 2 to 8 years with non-ambulatory Type II SMA, who can site independently for 3 seconds. This portion of the study is randomised double blind and placebo controlled. The study duration is for 12 months. For the first 6 months children will be divided into placebo and drug treatment groups. For the second 6 months, all children will receive drug. Sixty children are needed in this cohort. The second cohort is for children ages 3 to 17 years with SMA Type III who can stand independently without braces or other support for 2 second. All patients in cohort 2 receive drug, and 30 children are needed for this cohort. Enrolment of 20 patients is still needed over both cohorts at the Ohio, Maryland, Wisconsin, Michigan, and Montreal sites. The target date for the completion of enrolment is October 2006 with study completion in October of 2007. Please see www.projectcuresma.org for more details on how to contact the study coordinator at each site if you are interested in participating.

Dr. Swoboda also reported results on a few individual patients receiving sodium phenyl butyrate in patients younger than 2 years old in an open label trial. From this experience, it has been learned that it is quite difficult to administer in adequate amount of PBA. However, a few individual experiences have been promising. Therefore, attempts are being made to seek a formal very small-scale study with the support of the company that manufactures the product for another rare disorder. Alternatives are also being investigated e.g. new formulation so that the medicine is easier to take, although this is not available yet.

Dr Ching Wang reported on two other clinical trials he is leading at Stanford University to test the safety and efficacy of Hydroxyurea in patients living with SMA. Both of these trials are double blind randomised placebo trials with a ratio of 2 to 1 patients receiving drug versus placebo. The first study is being conducted in Type I SMA patients. Dr. Wang currently has 14 patients enrolled in this study and needs 4 more to complete enrolment. The second study involves Type II and severe Type III patients. Enrolment is complete for the Type II/ III trial with 24 patients enrolled at this point in time. The outcome measures being assessed in the study are Gross Motor Function Measurement, Time Test, and MUNE. This study is currently ongoing, and full analysis is not yet available. In addition, the study has not yet been unblinded (i.e. the patient receiving Hydroxyurea versus placebo remains unknown). However, the data suggest that the safety profile is reasonable. In a few patients a drop in the white blood cell count was noted, but was temporary. In the interim analysis, SMN transcript and protein levels were also assessed. A responder group with increased SMN levels was reported, although it unclear at this point whether these patients have received drug as they study is still blinded. This study is continuing over the next several months and final results regarding efficacy in the functional motor tests will be presented upon completion.

Professor Wirth then discussed the activity in Germany to develop a biomarker test using blood samples. This is part of the development of a clinical trials protocol to study sodium valproate in children under 2 years of age to be performed in Europe. Initially 10 SMA carriers were tested; they were given valproate for 3 months. Blood samples were taken before medication and on achievement of correct blood level of valproate. This study showed (n=7/10) that mRNA levels increased. However, 3 carriers did not show a change. Developing such a biomarker is crucial for early clinical trials in SMA.

A protocol is currently being designed, which originally was going to be a placebo-controlled study across three or four countries in the EU. However, on review with clinical experts in some EU countries it was agreed that the first study should be open label and probably designed to be country specific. Once there is information available on this study it should be possible to consider future European studies. There are currently discussions ongoing with the European Medicines Agency (EMEA) to ensure that all safety aspects of the study have been considered. The German experts are leading the development of the protocol at this moment and information will be provided on the German patient organisation website.

The development of an effective assay of mRNA or protein levels remains a critical goal of the research team, in addition to the ongoing work of Professor Wirth’s research into a blood assay, Professor Morris from Centre for Inherited Neuromuscular Diseases, UK described an ELISA assay kit that is currently being developed. This is able to detect protein levels that interact with SMN.

Outcome Measures

A number of the clinical experts discussed the development of more accurate scales for the evaluation changes in Type I, II and III patient. The main issue is that at this time there is an emphasis on the normal development of a child that does not allow for accurate measurement of changes in SMA children. In Type I children Professor Finkel discussed a collaborations between US, Italy and the UK to merge the most valuable scales (CHOP INTEND) to be able to detect the fine changes seen in Type I children, this has been specially developed to not stress the child being assessed but allow a precise evaluation of changes in physical development.

Then Professor Crawford then described the limitation of some of the current assessments used to follow Type II and severe Type III children. He noted that the demand to follow rigid manoeuvres at specific times does not allow the child to fully express their development stage and may even cause them not to participate. He has identified a series of manoeuvres that allow the clinician to rapidly see how the child is compensating for their disability. This scale is still under development but generated much interest at the meeting.

Dr. Krosschell then indicated that the current Hammersmith scale has limitations on the assessment of partially ambulatory children, this leads to a ceiling effect. They have developed a modified Hammersmith Function Motor Scale (MHFMS) to limit the ceiling effect in stronger Type II and Type III children. This theme was further developed by Dr. Kaufmann who explain how the Gross Motor Function Test (GMFT) is at times both time consuming and fatiguing. Because the Hammersmith Functional Motor scale (HFMS) is not fully developed for ambulatory children and has a ceiling effect she added 13 items on the test to allow better assessment of Type III children and allows them to being included in clinical studies. The recommendation was to use timed assessments to allow adequate comparison over separate visits to the hospital.

Basic Research

Stem Cells – Professor Doug Kerr gave the latest development from studies being carried out in stem cells, these are derived from the basic cell type that then differentiate to produce different specific cells e.g. muscle, nerve or heart cells. By taking the cell and adding specific growth factors and selection treatments it is possible to develop motor neurone specific cell lines that are able to grow beyond the myelin sheath towards a chemically induces stimulation. This has been used in rat models to help overcome viral born infections leading to disability in the lower limbs. Though not a full model of SMA this does help to show that in a damaged spinal cord it is possible to partially repair the neuronal link to the muscles. Unfortunately this involves multiple injections with a variety of potent agents that cannot be used in humans. He also developed motor neurons from embryonic stem cells from Smn-knockout mice to assess their defects in axon outgrowth more systematically.. The motor neurons developed very well, but the axonal outgrow was reduced. In a Petri dish the axons reached muscle cells, but after two days all connections were gone, supporting a role for SMN in motor neurons.

Dr. Keirstead next explained his work regarding the development of human motor neurons from human embryonic stem cells, which are obtained from in vitro fertilization procedures. He has developed a protocol to generate a very pure population of motor neurons (about 95%) from stem cells. His procedure has been developed and tested under strict government controls in compliance with the quality requirements of the FDA. This will allow human testing to advance at a much faster pace. In addition, Dr. Keirstead demonstrated during his talk that his lab has generated motor neurons that both look and behave like normal motor neurons. For example, they express genes known to be found only in motor neuron cells opposed to any other cell type. More importantly, he demonstrated that these motor neurons function like normal motor neurons. For example, Dr. Keirstead played a moving showing the motor neurons causing muscles to actively twitch when co-cultured together in a Petri dish. Dr. Keirstad’s ultimate goal is to produced clinically relevant cell population for therapeutic development in SMA, which can be used for motor neuron replacement

SMN Function – It is essential to understand the critical function of SMN in motor neurons in order to gain a better understanding of SMN disease pathology. Motor neurons are very special cells because they have cell bodies located in the spinal cord, but also must send out very long extensions called axons. These axons must grow all the way to the muscles to in order to control muscle movement, in many cases up to several feet in length.

SMN is known to have a very critical function in the cell body of every single cell type in the human body (specifically in scrip assembly and biogenesis). Dr. Dreyfuss presented data discussing the understanding of SMN function in the cell body of all cell types. He discussed a complicated series of interactions of many proteins with SMN, and showed how this large complex has a critical role RNP biogenesis. The SMN complex, comprised of SMN protein and Gemins, binds to Sm proteins and to the snRNAs to allow for the controlled RNA protein interactions needed RNP biogenesis. Dr Renvoise, who has demonstrated that SMN functional domains govern its intracellular localization and residency in gem/Cajal bodies, further developed this. Professor Pellizzoni showed that Gemin8 is essential for the architecture and function of the SMN complex in RNP biogenesis.

Although SMN clearly has a critical role in the cell bodies of every cell in the human body as discussed above, it is not fully understood why just one cell type, motor neurons, are affected in patients having SMA. One proposal is that SMN has a specialized function needed in motor axons. Another idea is because motor neurons are very specialized cells they could be more sensitive to the loss of the SMN in the cell body (snRNP assembly) than all the other cells types. While researchers are divided about this, it has been known for several years that SMN is found in both the cell body and the axons of motor neurons, suggesting a specialized axonal function is feasible. Some of the most exciting new data in Montreal focused on this controversy. Several presentations were given showing that SMN likely has a distinct function in the in motor axons than in the cell body SMN in the cell body, and that SMN forms a distinct complex in motor neurons than in the cell body. Representatives of the Beattie lab, the Sendtner lab, the Morris lab, the Cote lab, the Acsadi lab, the Kothary lab, Burghes lab, Androphy lab, Bassell lab, the Claus lab, and Battaglia labs gave these presentations.

Looking for Future Treatment – A number of talks focusing on drug discovery efforts to find novel small molecule therapies for SMA were presented in Montreal. Talks on this topic were given by representatives of the labs of deCODE Chemistry, Stephan Stamm Elliot Androphy, Christina Brahe, Brunhilde Wirth, Brent Stockwell, the NINDS SMA project, and Trophos.

Professor Stamm, University of Erlangen, Germany presented a paper on the use of Tra2-beta 1 phosphorylation dependent regulation of SMN2 exon 7, as a model to search for potential new treatments approaches. He has identified a series of small molecules, which by inhibiting phosporylation of tra2-beta1, cause inclusion of exon 7 and increase SMN2 gene expression. Further work has to be performed before the molecules can be considered in animal models.

A presentation by Dr Brahe, Institute of Medical Genetics, Catholic University, Rome discussed laboratory studies on salbutamol (a Beta2 agonist), which has been used in small clinical study in the UK to treat Type III patients. As a known treatment for asthma, it has an anabolic effect but also, though only with a short half-life, is an upregulator with effect on full length SMN protein production. There was discussion on the clinical experience with salbutamol or similar treatments and it is possible these only have a short term effect, however, a few patients have benefited from their use. The view was that placebo controlled clinical trials are necessary to be able to understand if there is a real benefit from the use of salbutamol rather that just a result of the stimulant effect.

We then received feedback from Institute of Human Genetics, Cologne where Dr. Riessland showed that a test compound (M344) from a new class of novel histone deacetylase inhibitors (HDAC) upregulates SMN 2 protein expression in fibroblasts from SMA patients, in tissue cultures. It was shown that this was due to significant delta 7 inclusion. M344 revealed a toxic cell effect at very high concentrations, so it may be a promising candidate for drug development. Another HDAC inhibitor, Trichostatin A was discussed by Dr. Sumner. Treatment of SMA mice at post natal day 5 (after onset of symptoms) led to 20% increase of survival i.e. 16 to 20 day.

Dr. Letso from Columbia University explained how they have developed a high throughput screen using luminescence to allow them to study multiple chemicals as possible future treatments. Dr. Meg Winberg, Director of Research of SMA Foundation explained how they are funding an animal testing facility to allow all rapid and consistent evaluation of medicines. This will be needed as more and more potential chemicals are being brought to a stage where pre-clinical testing can be completed prior to the start of clinical studies. Then Dr. Heemskerk from the US Government body, National Institute of Neurological Disorders and Stroke (NIH) explained the efforts of the NIH of developing pre-clinical testing facilities to support the development of potential treatments for SMA. A model treatment based on an known drug (indoprofen) is being studied by chemists to try and enhance the beneficial properties of the drug for possible treatment of SMA, while removing the known toxic effects and allow a better ability to cross the blood brain barrier.

Dr Bordet from the French Pharmaceutical Research Company Trophos, presented a neuroprotective compound, which is already in phase Ib trials for ALS (Amyotrophic Lateral Sclerosis). The compound TRO19622 promotes survival and blocks apoptosis, it is a cholesterol like compound that blocks cytochrome c and reduces caspase. Animal models in SMA are currently being considered.

Dr Singh and Dr Butchbach presented an update on the DECODE project. Dr Singh explained how they specially selected the characteristics of the specifically designed compound 156844, which is a quinazoline, to up regulate SMN levels. Then Dr Butchbach explained that this is very potent as with limited toxicity in vitro models for SMN expression. The product has been specifically designed to pass the blood brain barrier.

The FSMA/deCODE collaboration was initiated in 2003 as the continuation of a successful early phase drug discovery project at Vertex Pharmaceuticals. The deCODE chemistry-FSMA collaboration has focused on the optimization of a class of molecule called 2,4-diaminoquinazolines that was discovered in the high-throughput, cell-based assay developed at Vertex. During the past three years, deCODE chemistry has developed optimized analogues that have the vast majority of characteristic needed for a potential SMA drug. These compounds are highly potency in the cell-based assay, have excellent metabolic stability, show very efficient penetration of the blood-brain barrier, possess an attractive pharmacokinetic profile, and demonstrate the desired activity in SMA cellular models, including increasing the numbers of SMN containing gems structures in cells derived from SMA Type I patients to levels found in carrier cells. This is a very substantial increase in SMN protein levels. As the lead optimization phase of the projects comes to completion during the next 6 months and a final clinical candidate is selected, the focus of the collaboration will be to further assess the pharmaceutical properties of the clinical candidate in investigational new drug (IND) application-focused experiments. The IND package constitutes an extensive series of experiments required by the FDA to assess safety in animals before a drug can be tested to humans. With a novel compound like ours, human testing would commence in a Phase I trial, which is usually conducted in healthy volunteers. The goal of this first human study would be to assess drug safety and tolerability. If everything continues in positive manner, the anticipated timeline for IND filing with the FDA is the end of 2007 and Phase I testing can begin 30 days after that filing.
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Slides from the presentation available here