Petra D. Cravens

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.

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.

Decrease in the Numbers of Dendritic Cells and CD4+ T Cells in Cerebral Perivascular Spaces Due to Natalizumab

OBJECTIVE To extend our studies on the prolonged and differential effect of natalizumab on T lymphocyte numbers in the cerebrospinal fluid, we investigated the number and phenotypes of leukocytes and the expression of major histocompatibility complex (MHC) classes I and II in cerebral perivascular spaces (CPVS). We hypothesized that natalizumab reduces the number of antigen presenting cells in CPVS. DESIGN A case-control study in which inflammatory cell numbers in the CPVS of cerebral tissue were assessed by immunohistochemical staining. SUBJECTS A patient with multiple sclerosis (MS) who developed progressive multifocal leukoencephalopathy (PML) during natalizumab therapy. Controls included location-matched cerebral autopsy material of patients without disease of the central nervous system, patients with MS not treated with natalizumab, and patients with PML not associated with natalizumab therapy. RESULTS The absolute number of CPVS in the patient with MS treated with natalizumab was significantly lower than in the control groups owing to extensive destruction of the tissue architecture. The expression of MHC class II molecules and the number of CD209+ dendritic cells were significantly decreased in the CPVS of the patient with MS treated with natalizumab. No CD4+ T cells were detectable. CONCLUSIONS Our observations may explain the differential and prolonged effects of natalizumab therapy on leukocyte numbers in the cerebrospinal fluid.

Dendritic cells, chemokine receptors and autoimmune inflammatory diseases

Dendritic cells (DC) have been implicated in the induction of autoimmune diseases and have been identified in lesions associated with several autoimmune inflammatory diseases. Since DC are regarded as the professional antigen‐presenting cell (APC) of the immune system and the only APC capable of activating naïve T cells, they are likely to play a significant role in breaking tolerance of self‐reactive lymphocytes and in supporting autoimmune responses in these diseases. A number of studies have revealed that small molecular weight chemotactic proteins known as chemokines are present within the autoimmune lesions and may contribute to the recruitment not only of DC populations, but also of immune cells such as T cells, B cells, neutrophils and monocytes into the site, and to the formation of organized lymphoid tissue structures within the target organ. The focus of this review will be a discussion of the role of chemokines in the recruitment of DC in human autoimmune inflammatory disorders, specifically the trafficking of DC into the inflammatory sites and the subsequent migration of differentiated DC from the inflammatory sites into the draining lymph nodes. Once DC are properly positioned within the lymph nodes, circulating antigen specific naïve T cells can interact with DC and become activated, clonally expanded and stimulated to undergo differentiation into antigen‐experienced memory T cells. Subsequent reactivation of memory T cells that enter the autoimmune lesions by DC present in the inflammatory lesion is thought to play a central role in tissue inflammation.

Immunologic, clinical, and radiologic status 14 months after cessation of natalizumab therapy

Objective: Natalizumab is a humanized recombinant monoclonal antibody against very late activation antigen-4 approved for the treatment of patients with multiple sclerosis (MS). A phase II study failed to demonstrate a difference between natalizumab treatment groups and the placebo group with regard to gadolinium enhancing lesions on MRI 3 months after discontinuation of therapy. The objective of this study was to assess clinical MS disease activity, surrogate disease markers on MRI, immunologic parameters in peripheral blood and CSF, as well as safety in patients with MS after discontinuation of natalizumab therapy. Methods: This study is a longitudinal and serial cross-sectional assessment, in which 23 patients who were treated with natalizumab in the context of two phase III clinical trials were originally enrolled. A subgroup of patients was followed over 14 months. The annual relapse rate, neurologic disease progression assessed by the Expanded Disability Status Scale, disease surrogate markers on MRI, cellular and humoral immune markers in peripheral blood and CSF, and adverse events of the drug were monitored. Results: With regard to clinical disease activity, neuroimaging, and immune responses, the majority of patients in our cohort were stable. Decreased lymphocyte cell numbers and altered cell ratios returned to normal 14 months after cessation of natalizumab. No infectious complications were observed. Conclusion: This is the first long-term follow-up of patients who discontinued natalizumab. We did not observe a clinical, radiographic, or immunologic rebound phenomenon after discontinuation of natalizumab therapy.

Natalizumab and progressive multifocal leukoencephalopathy: What are the causal factors and can it be avoided?

Natalizumab (Tysabri) was the first monoclonal antibody approved for the treatment of relapsing forms of multiple sclerosis (MS). After its initial approval, 3 patients undergoing natalizumab therapy in combination with other immunoregulatory and immunosuppressive agents were diagnosed with progressive multifocal leukoencephalopathy (PML). The agent was later reapproved and its use restricted to monotherapy in patients with relapsing forms of MS. Since reapproval in 2006, additional cases of PML were reported in patients with MS receiving natalizumab monotherapy. Thus, there is currently no convincing evidence that natalizumab-associated PML is restricted to combination therapy with other disease-modifying or immunosuppressive agents. In addition, recent data indicate that risk of PML might increase beyond 24 months of treatment.

Memory B cells from a subset of treatment-naïve relapsing-remitting multiple sclerosis patients elicit CD4(+) T-cell proliferation and IFN-γ production in response to myelin basic protein and myelin oligodendrocyte glycoprotein.

Recent evidence suggests that B‐ and T‐cell interactions may be paramount in relapsing‐remitting MS (RRMS) disease pathogenesis. We hypothesized that memory B‐cell pools from RRMS patients may specifically harbor a subset of potent neuro‐APC that support neuro‐Ag reactive T‐cell proliferation and cytokine secretion. To test this hypothesis, we compared CD80 and HLA‐DR expression, IL‐10 and lymphotoxin‐α secretion, neuro‐Ag binding capacity, and neuro‐Ag presentation by memory B cells from RRMS patients to naïve B cells from RRMS patients and to memory and naïve B cells from healthy donors (HD). We identified memory B cells from some RRMS patients that elicited CD4+ T‐cell proliferation and IFN‐γ secretion in response to myelin basic protein and myelin oligodendrocyte glycoprotein. Notwithstanding the fact that the phenotypic parameters that promote efficient Ag presentation were observed to be similar between RRMS and HD memory B cells, a corresponding capability to elicit CD4+ T‐cell proliferation in response to myelin basic protein and myelin oligodendrocyte glycoprotein was not observed in HD memory B cells. Our results demonstrate for the first time that the memory B‐cell pool in RRMS harbors neuro‐Ag specific B cells that can activate T cells.

Depletion of B Lymphocytes From Cerebral Perivascular Spaces by Rituximab

BACKGROUND Rituximab is a recombinant chimeric monoclonal antibody against CD20, a molecule expressed on cells of the B-cell lineage. A phase 2 clinical trial recently provided strong evidence of the beneficial effects of rituximab in patients with relapsing-remitting multiple sclerosis. We and other investigators previously demonstrated that rituximab therapy depletes B lymphocytes from peripheral blood and cerebrospinal fluid of patients with relapsing-remitting multiple sclerosis. OBJECTIVE To determine the effect of rituximab on the presence of B cells in cerebral perivascular spaces. Design, Setting, and Patients Case report from a tertiary academic medical center. Cerebral white matter from autopsy material of a patient with gastrointestinal mantle-cell lymphoma who developed progressive multifocal leukoencephalopathy following rituximab therapy was evaluated by immunohistochemistry. Location-matched brain sections of patients with multiple sclerosis not treated with rituximab, patients without central nervous system disease, and patients with progressive multifocal leukoencephalopathy not associated with rituximab were used as controls. MAIN OUTCOME MEASURES Assessment of the number of B lymphocytes in cerebral perivascular spaces in a patient with gastrointestinal mantle-cell lymphoma treated with rituximab, patients with multiple sclerosis, patients with progressive multifocal leukoencephalopathy not associated with rituximab, and healthy control subjects. RESULTS We were unable to detect B cells in cerebral perivascular spaces of the patient who developed progressive multifocal leukoencephalopathy following rituximab therapy 8 months after her last dose. In contrast, B cells were detectable in all control brain tissues. CONCLUSIONS To our knowledge, this is the first report to show B-lymphocyte depletion from brain tissue following rituximab therapy. A reduction in B-cell numbers may be an important contributing factor in the pathogenesis of central nervous system infections.

Silencing Nogo-A Promotes Functional Recovery in Demyelinating Disease

To determine if suppressing Nogo‐A, an axonal inhibitory protein, will promote functional recovery in a murine model of multiple sclerosis (MS).