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Conference on the Cause and Treatment of Facioscapulohumeral Muscular Dystrophy


May 8-9, 2000

Sponsored by:

The National Institute of Arthritis and Musculoskeletal and Skin Diseases
The National Institute of Neurological Disorders and Stroke
The NIH Office of Rare Diseases
The FSH Society, Inc.
The Muscular Dystrophy Association of America (MDA)

Table of Contents

Meeting Summary

Day 1 

Facioscapulohumeral muscular dystrophy (FSHD) is the third most common genetic disease of skeletal muscle. It affects approximately one in 20,000 persons. It is inherited in an autosomal dominant way, so that a single copy of the defective DNA may cause disease. Researchers have identified the mutation associated with more than 95% of cases. Though the nature of the DNA mutation is known, it has not yet been possible to identify a gene or mechanism that causes the disease.

The DNA mutation consists of deletions of copies of whole repeat units, each called D4Z4, located near the short end of chromosome 4, identified by the term 4q35. Each repeat unit, called D4Z4, consists of 3.3 kilobases of DNA, meaning that it is a strand of more than three thousand individual nucleotides. The number of such repeats in a person unaffected by FSHD ranges from twelve to one hundred, disease severity increases as the number of repeats decreases below ten. During the conference, researchers discussed the clinical criteria of FSHD and reported new clinical observations. Clinicians presented the results of a one-year double-blind clinical trial using albuterol, a drug that normally increases muscle mass. The results of this study in FSHD patients were largely negative, except for slightly improved grip strength and preservation of skeletal muscle mass. Another group assessed the safety and efficacy of muscle strength training in FSHD patients, finding that moderate exercise is safe and dynamic strength training may provide limited gain of dynamic strength specific to the muscles trained. Conferees identified several clinical areas where questions remain, such as determining the incidence of cardiac involvement and tongue and throat weakness in FSHD patients. A further important question is whether males and females have a different relationship between the size of the genetic deletion and clinical manifestations of the disease.

Molecular diagnosis of FSHD is complicated because there is a region of DNA on chromosome 10 which is almost completely identical to the region on chromosome 4 associated with the disease. However, improved biochemical analysis detects slight differences in the DNA sequence between the regions on chromosomes 4 and 10. Researchers looking at different patient populations and unaffected individuals found that genetic events in reproduction frequently result in exchange of genetic material between the similar regions of the two chromosomes. It was suggested that the high frequency of new occurrences of the disease may result from this exchange of DNA between chromosomes 4 and 10. The conferees also addressed the question of the size of the deletion and disease diagnosis, i.e., the smallest region found in asymptomatic family members or control populations. There was general agreement that fewer than eight repeats are uniquely associated with clinical symptoms of FSHD, while more than twelve are relatively rare among FSHD patients. New data presented continue to support the correlation between the size of the DNA deletion and disease severity. Lastly, researchers showed that they could improve molecular diagnostics using a technology called pulsed-field gel electrophoresis, which shows more clearly differences in DNA composition.

There is one family with members who have a clinical phenotype that is virtually indistinguishable from that of FSHD, but whose disease is not linked to the region of chromosome 4 associated with disease in 95% of FSHD patients. This family alone has more than 40 affected individuals and researchers are using it in their search for a second locus for FSHD.

Though no genes have been found in the site of the FSHD genetic defect (4q35), scientists continue to look for genes closer to the center of chromosome 4. They have found a region with the genes ANT and ALP and a newly characterized gene, SMT7. It is likely there are other genes in this area. Expression of the known genes (ANT, ALP, and SMT7) was shown to be significantly higher in skeletal muscle biopsy samples from FSHD patients compared to controls. Two genes lying closest to the repeats are FRG1 and FRG2. Characterizations of the expression patterns of these two genes, and the roles of their putative gene products, are being carried out. Researchers are planning on using a mouse model in which it will be possible to cause a higher than normal production of the gene products from genes such as ANT, ALP, and SMT7, which are near the FSHD region of human chromosome 4.

There are several lines of investigation in progress which focus on the D4Z4 repeats themselves. One group has identified a sequence of DNA, which they call DUX4, in the 4q35 region associated with FSHD. These researchers believe that DUX4 may in FSHD patients cause production of a protein that does not appear in unaffected individuals. The DUX4 cDNA sequence and gene product are being studied in cellular and animal models. Another investigator is studying the role of the repeats in chromatin formation (the local chromosome structure based on the DNA strand and associated substances) and repression of adjacent gene transcription, using a fruit fly (Drosophila) model in which one can construct strands of genetic material containing different copy numbers and different regions of the 3.3 kilobase repeats. A different approach is based on the observation that chemical attachment of methyl groups to DNA (methylation) is a normal process which can affect the transcription of genes. Studies indicate that the D4Z4 repeats are highly methylated in cells from unaffected people. It is possible that the deletions in the FSHD region cause a decrease in methylation, which may affect the packing of material attached to the DNA in this region of the chromosome, and disrupt gene expression.

Differential gene expression studies of muscle and regenerative muscle cells (myoblasts) from FSHD patients and controls are now providing the first directions for experiments to explore pathogenetic mechanisms that occur as a consequence of deletion of D4Z4 repeats on chromosome 4. Researchers are studying several proteins, including histone acetyl transferase (a protein that adds acetyl groups to histones, which appear vital for cellular viability), extracellular matrix proteins, and enzymes involved in cell response to the presence of highly reactive oxidative molecules, such as peroxide, which produce oxidative stress. In studies comparing myoblasts (muscle regenerative cells) from people with and without muscle diseases, researchers found that undifferentiated myoblasts from FSHD patients have a unique early sensitivity to highly reactive oxidative molecules, which provides the first biochemical hallmark of the disorder.

Future studies along the many complementary research avenues described at the conference will almost certainly prove fruitful in shedding light on FSHD from the clinical and molecular points of view, clarifying the relationship between genotype and phenotype.

Day 2  

Drs. Richard Lymn, NIAMS, and Giovanna Spinella, NINDS, opened the second day of the meeting with participant introductions and the background for continuing discussion of FSHD research needs and opportunities. The purpose of the brainstorming session was to build on current knowledge about FSHD and identify research approaches to understanding the riddles of this disease. NIAMS and NINDS invited several researchers expert in areas related to FSHD to bring additional insights into possible approaches to future investigations.

The discussion ranged over possible directions for future research. Major topic areas were approaches to determining pathogenesis, approaches to therapy, and resources needed to move research forward.

Recommendations for future directions, organized by topic, are listed below:

Molecular Processes

Characterize the molecular pathogenesis of FSHD; elucidate the role of the repeats associated with the disease as well as what causes their deletion. FSHD is associated with deletions of copies of whole repeat units from the end (subtelomeric) region of chromosome 4q. The deletion appears to result in global dislocation of gene expression. If the entire region is removed, there are birth defects, but no specific defects on skeletal muscle. Individuals appear to require the existence of 11 or fewer repeat units to be at risk for FSHD.

Determine the relationship between repeat length and its effect on the degree to which disease is manifested (penetrance). Determine also whether the loss of certain repeats is always associated with FSHD clinical expression, since there may be specificity in chromosomal transactions and the resulting development of disease.

Determine the gene sequence and whether the repeats are acting as suppressors or insulating units. The region containing the site associated with at least 95% of FSHD cases is composed almost entirely of the 3.3 kilobase repeats, which are not translated. Few genes have been found near the multi-repeat locus, including FRG1 and FRG2 in adjoining regions, and they do not appear to be related to development of disease. This suggests that FSHD may result from alterations in the chromatin structure, the local chromosome structure based on the DNA strand and associated substances. In particular, the data suggest that you need a minimum number of repeats in order to have a compact heterochromatin structure, which is the genetically inert form of chromosome packing. Lack of the compact structure may lead to disease through an unknown mechanism.

Clarify how similarity of regions on chromosomes 4 and 10 may relate to FSHD. There is a region on chromosome 10 that appears to be largely identical (95% homology) to that on chromosome 4 at the FSHD locus (4q35). Studies of affected and unaffected populations show that there is a high amount of exchange between the homologous regions on chromosomes 4 and 10. Although disease has never been associated with alterations on chromosome 10, the frequency of such exchanges may be related to the high proportion of new (i.e., non-familial) cases of FSHD encountered.

Tissue Changes

Characterize changes in muscle as the disease develops. This would be facilitated by non-invasive ways of looking at the muscle and microvasculature in affected and non-symptomatic regions. Studies using improved imaging techniques would provide better assessment of patient muscle, including vasculature, before the development of clinical symptoms. Confirmatory biopsies would be useful, though they are limited by the difficulty of the procedure in affected people and risk-benefit concerns.

Determine basis of differential involvement of muscles, reflected by the regional pattern of disease. Comparison of muscle groups might show the cause of relative specificity of affected muscles. Comparing expression patterns of working copies of individual genes (ribonucleic acid or RNA) and protein in affected and non-affected muscle will provide insights into alterations occurring as the disease progresses.

Explore the role of inflammation in FSHD. While FSHD has been described as the most inflammatory form of muscular dystrophy, there is no evidence that disease severity is lessened by administration of the anti-inflammatory drug prednisone. It is necessary to explore the relationship between inflammatory cells, muscle cell death, and blood vessels. The inflammatory response may affect vasculature, since hearing loss and retinal vasculopathy are widespread in FSHD patients.

Study properties of muscle cells derived from affected tissue. Cells cultured from FSHD muscle show increased sensitivity to oxidative stress. This needs to be followed up by studies verifying that this occurs in vivo and establishing how this cellular phenotype develops.

Possible Therapies

It was speculated that it may become possible to repair the disease locus by selected and targeted addition of 3.3 kb repeats to the disease locus on chromosome 4. Such targeted gene therapy might prevent development of the FSHD phenotype, but the practical feasibility of such an approach is as yet unknown.

Another approach to explore is the modification of cultured FSHD regenerative muscle cells that would reverse their higher sensitivity to oxidative stress. Such cultured cells, with better ability to respond to oxidative stress, might then be used for treatment of patients. It would be valuable to prevent the reintroduced cells from again developing increased sensitivity.

Population-Based Studies

Establish patient registries and recruit additional families for study. Increase the number of studies on the relationships between genotype and phenotype. This requires accurate and robust genotyping studies comparing disease severity within families. It is difficult to obtain financial support to establish genotype information. There are privacy issues associated with doing this on a broad scale, but establishing a central registry would help.

Determine if a nonstandard locus produces FSHD. There is one family where members have a disease that has characteristics of FSHD, but no defect has been found at the location of chromosome 4 associated with at least 95% of known cases. It is important to characterize the gene defects in this family and see if this provides a better understanding of FSHD disease processes.

Resources

Create new animal models. Understanding of FSHD would be improved by general availability of a good animal model that has genetic defects and phenotypes that are similar to the human disease. Once a strong colony is established, mice should be readily available to groups with expertise in many areas of muscle biology.

Facilitate use of differential gene and protein expression techniques which can serve as the basis of additional FSHD models, as well as provide directions for new therapeutic approaches.

Promote development and use of non-invasive imaging techniques, such as MRI, in order to analyze the state of tissue in affected and non-affected muscle and during disease progression.

Enhance formation of clinical and basic research consortia. Such consortia could initiate clinical and basic studies, and conduct clinical trials. One specific suggestion was to create centers that would develop and test animal models, establish patient registries, and provide tissue and genotypic analysis.

Last updated April 15, 2011