The Connective Tissue and Diseases Section began studying inflammatory myopathies (polymyositis, dermatomyositis, and related diseases) some years ago in an attempt to understand the relationship of autoantibodies to autoimmune disease. At the time, these diseases seemed to offer the best example of autoimmune diseases associated with highly specific disease-related autoantibodies and evidence of a viral etiology. They are very uncommon and hence relatively less studied than other autoimmune diseases, and they are very debilitating and hence in need of improved therapy. In order to attract patients here to allow more detailed clinical, immunological, genetic, and viral studies, we began doing trials of therapy and have completed a number of such studies. They are among the very few published controlled trials in this difficult to treat family of illnesses. These trials have encompassed a variety of approaches to immunosuppression. The most recent trial - a pilot trial of the anti-thyroid drug methimazole - was undertaken to make use of an unexpected property of the drug which was discovered by Dr. Leonard Kohn of NIDDK and Dr. Dinah Singer of NCI: that it can down-regulate MHC. An analysis of this trial has been completed and submitted for publication. We have just begun a trial of the ANTI-TNF AGENT, INFLIXIMAB (REMICADE), a mouse-human chimeric antibody that has been approved for use in rheumatoid arthritis and inflammatory bowel disease. The rationale for carrying out the trial is the evidence, admittedly modest, that TNF is present in the active lesions in myositis biopsies. The trial is being carried out with financial support from the manufacturer, Centocor, under a clinical CRADA. The patient population is similar to that we have used in other studies - those who have had unsatisfactory responses to conventional immunosuppressive therapy. Defining diagnostic boundaries. Our referral clinic sees quite a number of patients who have been diagnosed as having myositis and having been treated unsuccessfully with standard therapy. A substantial proportion of these patients have either a demonstrable genetic disease (such as McArdle's, PFK deficiency, acid maltase deficiency, one of the many types of limb girdle dystrophy) or a presumptive genetic disease awaiting precise diagnosis (undiagnosable dystrophy, undiagnosable vacuolar myopathy, undiagnosable channelopathy.) We have joined with Eric Hoffman's lab in the Center for Genetic Medicine at the Children's National Medical Center to attempt to use gene expression to sharpen diagnosis. Our first study has been an attempt to separate dysferlin deficiency (which causes Miyoshi myopathy and type IIb limb girdle dystrophy) from myositis. We chose this problem because inflammation is present in dysferlin deficiency. The analysis of gene expression in dysferlin deficiency, including a comparison to Duchenne Muscular Dystrophy, is largely done and is being prepared for submission. The major finding is that a number of membrane remodeling genes are up-regulated, consonant with and substantially extending published histologic findings. We used Affymetrix U95A and MuscleChip microarrays to generate 14 expression profiles from 5 patients with dysferlin deficiency. By comparing these profiles to profiles of normal muscle (n=22) and of muscle from patients with Duchenne Muscular Dystrophy (n=26) using both Affymetrix and GeneSpring software we identified 106 genes with dysregulation specific to dysferlin deficiency. Seventeen of these genes are involved in membrane regulation, with six genes known to participate directly in the regulation of vesicle formation or trafficking, including RAD, Rab5C, EPSIN, and arfaptin1. We have confirmed altered expression of both vesicle trafficking proteins as well as membrane signaling molecules by QMF-RT-PCR and immunohistochemistry, and have observed altered plasma membrane expression and/or localization of a number of proteins important for membrane signaling and stability. Analysis of the effects on inflammatory genes is still in process. Future plans: We will begin by analyzing gene expression in a group of biopsies from patients with very early myositis in whom there is negligible visible inflammation. Our plan is to determine whether or not expression studies, particularly of inflammatory genes, can be used to separate primary inflammatory muscle disease (myositis) from diseases that can closely resemble it.
