Michael K. Racke

Immune surveillance in multiple sclerosis patients treated with natalizumab.

Our objective was to test whether natalizumab, an antibody against very late activating antigen (VLA)‐4, interferes with central nervous system immune surveillance as assessed by leukocyte cell numbers and cellular phenotypes in cerebrospinal fluid (CSF) and peripheral blood.

Peroxisome Proliferator-Activated Receptor-γ Agonist 15-Deoxy-Δ12,1412,14-Prostaglandin J2 Ameliorates Experimental Autoimmune Encephalomyelitis

Peroxisome proliferator-activated receptors (PPAR) are members of a nuclear hormone receptor superfamily that includes receptors for steroids, retinoids, and thyroid hormone, all of which are known to affect the immune response. Previous studies dealing with PPAR-γ expression in the immune system have been limited. Recently, PPAR-γ was identified in monocyte/macrophage cells. In this study we examined the role of PPAR-γ in experimental autoimmune encephalomyelitis (EAE), an animal model for the human disease multiple sclerosis. The hypothesis we are testing is whether PPAR-γ plays an important role in EAE pathogenesis and whether PPAR-γ ligands can inhibit the clinical expression of EAE. Initial studies have shown that the presence of the PPAR-γ ligand 15-deoxy-Δ12,14-PGJ2 (15d-PGJ2) inhibits the proliferation of Ag-specific T cells from the spleen of myelin basic protein Ac1–11 TCR-transgenic mice. 15d-PGJ2 suppressed IFN-γ, ΙL-10, and IL-4 production by both Con A- and myelin basic protein Ac1–11 peptide-stimulated lymphocytes as determined by ELISA and ELISPOT assay. Culture of encephalitogenic T cells with 15d-PGJ2 in the presence of Ag reduced the ability of these cells to adoptively transfer EAE. Examination of the target organ, the CNS, during the course of EAE revealed expression of PPAR-γ in the spinal cord inflammatory infiltrate. Administration of 15d-PGJ2 before and at the onset of clinical signs of EAE significantly reduced the severity of disease. These results suggest that PPAR-γ ligands may be a novel therapeutic agent for diseases such as multiple sclerosis.

T-bet Regulates the Fate of Th1 and Th17 Lymphocytes in Autoimmunity

IL-17-producing T cells (Th17) have recently been implicated in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), an animal model for the human disease multiple sclerosis. However, little is known about the transcription factors that regulate these cells. Although it is clear that the transcription factor T-bet plays an essential role in the differentiation of IFN-γ-producing CD4+ Th1 lymphocytes, the potential role of T-bet in the differentiation of Th17 cells is not completely understood. In this study, therapeutic administration of a small interfering RNA specific for T-bet significantly improved the clinical course of established EAE. The improved clinical course was associated with suppression of newly differentiated T cells that express IL-17 in the CNS as well as suppression of myelin basic protein-specific Th1 autoreactive T cells. Moreover, T-bet was found to directly regulate transcription of the IL-23R, and, in doing so, influenced the fate of Th17 cells, which depend on optimal IL-23 production for survival. We now show for the first time that suppression of T-bet ameliorates EAE by limiting the differentiation of autoreactive Th1 cells, as well as inhibiting pathogenic Th17 cells via regulation of IL-23R.

T-bet is essential for encephalitogenicity of both Th1 and Th17 cells.

The extent to which myelin-specific Th1 and Th17 cells contribute to the pathogenesis of experimental autoimmune encephalomyelitis (EAE) is controversial. Combinations of interleukin (IL)-1β, IL-6, and IL-23 with transforming growth factor β were used to differentiate myelin-specific T cell receptor transgenic T cells into Th17 cells, none of which could induce EAE, whereas Th1 cells consistently transferred disease. However, IL-6 was found to promote the differentiation of encephalitogenic Th17 cells. Further analysis of myelin-specific T cells that were encephalitogenic in spontaneous EAE and actively induced EAE demonstrated that T-bet expression was critical for pathogenicity, regardless of cytokine expression by the encephalitogenic T cells. These data suggest that encephalitogenicity of myelin-specific T cells appears to be mediated by a pathway dependent on T-bet and not necessarily pathway-specific end products, such as interferon γ and IL-17.

Peroxisome Proliferator-Activated Receptor α Agonists as Therapy for Autoimmune Disease

Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor superfamily. PPARγ ligands, which include the naturally occurring PG metabolite 15-deoxy-Δ12,14-PGJ2 (15d-PGJ2), as well as thiazolidinediones, have been shown to have anti-inflammatory activity. The PPARα agonists, gemfibrozil, ciprofibrate, and fenofibrate, have an excellent track history as oral agents used to treat hypertriglyceridemia. In the present study, we demonstrate that these PPARα agonists can increase the production of the Th2 cytokine, IL-4, and suppress proliferation by TCR transgenic T cells specific for the myelin basic protein Ac1–11, as well as reduce NO production by microglia. Oral administration of gemfibrozil and fenofibrate inhibited clinical signs of experimental autoimmune encephalomyelitis. More importantly, gemfibrozil was shown to shift the cytokine secretion of human T cell lines by inhibiting IFN-γ and promoting IL-4 secretion. These results suggest that PPARα agonists such as gemfibrozil and fenofibrate, may be attractive candidates for use in human inflammatory conditions such as multiple sclerosis.

The utility of MRI in suspected MS Report of the Therapeutics and Technology Assessment Subcommittee of the American Academy of Neurology

Advancements in imaging technologies and newly evolving treatments offer the promise of more effective management strategies for MS. Until recently, confirmation of the diagnosis of MS has generally required the demonstration of clinical activity that is disseminated in both time and space. Nevertheless, with the advent of MRI techniques, occult disease activity can be demonstrated in 50 to 80% of patients at the time of the first clinical presentation. Prospective studies have shown that the presence of such lesions predicts future conversion to clinically definite (CD) MS. Indeed, in a young to middle-aged adult with a clinically isolated syndrome (CIS), once alternative diagnoses are excluded at baseline, the finding of three or more white matter lesions on a T2-weighted MRI scan (especially if one of these lesions is located in the periventricular region) is a very sensitive predictor (>80%) of the subsequent development of CDMS within the next 7 to 10 years. Moreover, the presence of two or more gadolinium (Gd)-enhancing lesions at baseline and the appearance of either new T2 lesions or new Gd enhancement on follow-up scans are also highly predictive of the subsequent development of CDMS in the near term. By contrast, normal results on MRI at the time of clinical presentation makes the future development of CDMS considerably less likely.

Distinct roles for B7-1 (CD-80) and B7-2 (CD-86) in the initiation of experimental allergic encephalomyelitis.

The activation and differentiation of T cells require both antigen/MHC recognition and costimulatory signals. The present studies examined the role of B7-1 (CD80) and B7-2 (CD86) costimulation in the prototypic autoimmune disorder, experimental allergic encephalomyelitis (EAE). In adoptively transferred EAE, in vitro activation of myelin basic protein (MBP)-specific lymph node cells was inhibited by the combination of anti-CD80 plus anti-CD86, but not individually. However, in actively induced disease, one injection of anti-CD80 significantly reduced disease, while anti-CD86 exacerbated disease. Interestingly, one injection of CTLA-4Ig suppressed disease, while multiple injections resulted in enhanced disease. Thus, the costimulation provided by B7-1 molecules appears to be important for the development of encephalitogenic T cells. The enhanced disease caused by multiple injections of CTLA-4Ig or a single injection of anti-CD86 suggests an inhibitory function for CD86 interaction with its counterreceptors CD28 and CTLA-4 in EAE. Alternatively, these results are consistent with an essential timing requirement for the coordinated interaction of B7 and CD28 family receptors, and that disruption of this critical timing can have opposing results on the outcome of an immune response.

Glatiramer acetate (Copaxone) therapy induces CD8+ T cell responses in patients with multiple sclerosis

Glatiramer acetate (GA; Copaxone) is a random copolymer of glutamic acid, lysine, alanine, and tyrosine that is used therapeutically in patients with multiple sclerosis (MS). To investigate the mechanism of the drugs immunomodulatory effect, we used immunophenotypic approaches to characterize the precise nature of GA-induced T cell responses. We demonstrate here that healthy individuals and untreated MS patients exhibit prominent T cell proliferative responses to GA. However, these responses are different in distinct subsets of T cells. Whereas GA-induced CD4(+) T cell responses are comparable in healthy individuals and MS patients, CD8(+) T cell responses are significantly lower in untreated MS patients. Treatment with GA results in upregulation of these CD8(+) responses with restoration to levels observed in healthy individuals. Both CD4(+) and CD8(+) GA-specific responses are HLA-restricted. GA therapy also induces a change in the cytokine profile of GA-specific CD4(+) and CD8(+) T cells. This study provides the first direct immunophenotypic evidence, to our knowledge, of GA-specific CD8(+) T cell responses and their upregulation during the course of therapy, which may suggest a role for these responses in the immunomodulatory effects of the drug.

Therapeutic Induction of Regulatory, Cytotoxic CD8+ T Cells in Multiple Sclerosis

In the setting of autoimmunity, one of the goals of successful therapeutic immune modulation is the induction of peripheral tolerance, a large part of which is mediated by regulatory/suppressor T cells. In this report, we demonstrate a novel immunomodulatory mechanism by an FDA-approved, exogenous peptide-based therapy that incites an HLA class I-restricted, cytotoxic suppressor CD8+ T cell response. We have shown previously that treatment of multiple sclerosis (MS) with glatiramer acetate (GA; Copaxone) induces differential up-regulation of GA-reactive CD8+ T cell responses. We now show that these GA-induced CD8+ T cells are regulatory/suppressor in nature. Untreated patients show overall deficit in CD8+ T cell-mediated suppression, compared with healthy subjects. GA therapy significantly enhances this suppressive ability, which is mediated by cell contact-dependent mechanisms. CD8+ T cells from GA-treated patients and healthy subjects, but not those from untreated patients with MS, exhibit potent, HLA class I-restricted, GA-specific cytotoxicity. We further show that these GA-induced cytotoxic CD8+ T cells can directly kill CD4+ T cells in a GA-specific manner. Killing is enhanced by preactivation of target CD4+ T cells and may depend on presentation of GA through HLA-E. Thus, we demonstrate that GA therapy induces a suppressor/cytotoxic CD8+ T cell response, which is capable of modulating in vivo immune responses during ongoing therapy. These studies not only explain several prior observations relating to the mechanism of this drug but also provide important insights into the natural immune interplay underlying this human immune-mediated disease.

The mechanism of action of glatiramer acetate treatment in multiple sclerosis

Objective: Glatiramer acetate (formerly known as copolymer 1) is the major noninterferon immunomodulatory agent used in the treatment of relapsing-remitting multiple sclerosis. Its mechanism of action over the past 40 years has evolved with our understanding of the immune response. Methods: We review the various mechanisms that have been proposed for this random polymer over the years, with emphasis on recent methods that utilize modern immunologic techniques. Results: Studies describing processes such as immune deviation and effects on regulatory T cells and antigen-presenting cells are presented. Conclusions: Effects of glatiramer acetate on the immune response have evolved as our technical abilities and knowledge of the immune response itself have developed.