Predictive cytokine biomarkers of clinical response to glatiramer acetate therapy in multiple sclerosis.

A prospective study of 62 patients with relapsing-remitting multiple sclerosis (RRMS) treated with Glatiramer acetate (GA) was conducted to evaluate the value of baseline and treatment-modulated cytokines in predicting the clinical response to the drug after 2years of therapy. There were 32 responders and 30 non-responders. GA upregulated Th2/regulatory cytokines and inhibited Th1 cytokines in sera or PBMC supernatants 3 and 6months into treatment. We found two prognostic models with clinical utility. A model based on IL-18 at baseline, the change in TNFa from baseline to 3months, the change in IL-4 from baseline to 6months, and the change in the log of the ratio of TNFa/IL-4 from baseline to 6months had an area under the curve (AUC) of 0.80. A high IL-18 level at baseline and a reduction of TNF-alpha over time are associated with a response to GA. Although the study identified predictive biomarkers of clinical response to GA, the results will need to be validated in other data sets.

Overlapping and distinct mechanisms of action of multiple sclerosis therapies.

In the last two decades MS has changed from an idiopathic condition with only symptomatic treatments to a disease with better characterized pathophysiologic underpinnings and several treatments that modify its long-term course based on specific mechanisms of action. There are now several FDA approved options for therapy at the onset of disease, and discussions have begun on choosing the best treatment in individual patients and what option to choose next in patients who are failing their current treatment. Numerous studies have begun to highlight that the underlying pathology of CNS damage may be different in subsets of patients, raising the possibility that some may respond to a treatment with a mechanism of action that is targeted to 'their' MS. Trials are ongoing of numerous new agents with different mechanisms of action and some combination therapies. A better understanding of how each therapy works may guide decisions on initiating, combining or changing therapy in a more rational way to improve patient outcomes. Further knowledge of underlying mechanisms of disease in different patients with 'the same' disease may lead to more targeted therapies, as will biomarkers that predict clinical response to therapy. Studies of the effects of various agents used in MS reveal both overlapping and distinct mechanisms of actions that may be relevant to their efficacy and side effects in individual patients. However, it is important to remember that most agents are approved based on their reduction of MRI lesions and relapse rates over a short time frame. These measures only partially correlate with long-term disability, which may be the most relevant clinical outcome for people with MS. Fixed disability requires years to become apparent, and there is a lack of large studies of biomarkers for chronic outcomes. In addition, few large studies correlate response to therapy with cognitive outcomes, which are a major cause of chronic disability. This review will attempt to summarize clinically relevant knowledge of the mechanisms of action of current FDA approved therapies for MS in the context of ongoing clinical trials of combination therapy and address rational approaches to changing therapy in a patient suspected to be 'unresponsive' to their current treatment.

Clinical response to glatiramer acetate correlates with modulation of IFN-gamma and IL-4 expression in multiple sclerosis.

OBJECTIVE: To determine whether glatiramer acetate (GA)-induced lymphoproliferation and IFN-gamma and IL-4 modulation correlate with the clinical response in multiple sclerosis (MS). BACKGROUND: GA therapy involves the induction of anti-inflammatory cytokine shifts. However, it is not known whether this response correlates with the clinical outcome. METHODS: Thirty-six relapsing-remitting (RR) MS patients were treated with GA for at least two years, and classified clinically as GA-responders (GA-R=22) or hypo/non-responders (GA-HR/NR = 14). Proliferation of peripheral blood mononuclear cells (PBMC) to GA and Tetanus toxoid (TT), as well as IL-4 and IFN-gamma ELISPOT, were performed. FINDINGS: There was no difference in PBMC proliferation to GA or TT between GA-R and GA-HR/NR before and during treatment (P>0.05). The mean number of IFN-gamma ELISPOTS in unstimulated, TT and anti-CD3/CD28-stimulated PBMC was lower among GA-R (unstimulated: GA-R =10.1+/-6.21 (n=22) versus GA-HR/NR=17.8+/-12.7 (n=14), P=0.04; TT-GA-R =12.2+/-4.06 (n=12) versus GA-HR/NR=26.8+/-21.0 (n=8), P=0.028; anti-CD-3/CD28 GA-R=217.3+/-140.4 (n=22) versus GA-HR/NR=368.5+/-170.1 (n=14), P=0.006). In contrast, the number of IL-4 ELISPOTS remained unchanged in the GA-R group, but was progressively reduced in the GA-HR/NR group during GA therapy (GA-HR/NR IL-4: pre-Rx: 59+/-34 versus 22+/-11 at 12 months (n =6), P=0.0429). The IL-4/ IFN-gamma ratio in anti-CD3/CD28-stimulated PBMC was significantly higher among GA-R compared to GA-HR/NR (P=0.0474). INTERPRETATION: Lymphoproliferation to GA did not differentiate GA-R from GA-HR/NR. However, reduced IFN-gamma expression and stable IL-4 expression in anti-CD3/CD28-stimulated PBMC, and an increased IL-4/IFN-gamma ratio was associated with favorable clinical response. More data are needed to validate the prospective use of IL-4/IFN-gamma expression in PBMC as a biomarker of clinical response to GA for individual patients.

Interferon-beta-1a induces increases in vascular cell adhesion molecule: implications for its mode of action in multiple sclerosis.

We investigated soluble vascular cell adhesion molecule-1 (sVCAM) levels and MRI lesions over 24 weeks in 15 Relapsing Remitting MS (RRMS) patients randomized prospectively to receive once-weekly (qw) IFN-beta-1a 30 mug intramuscularly (IM) (Group I, 8 patients) or three-times-weekly (tiw) IFN-beta-1a 44 mug subcutaneously (SC) (Group II, 7 patients). Both groups demonstrated a significant increase in sVCAM during treatment when compared to pre-treatment levels. Patients on IFN-beta-1a 44 mug SC tiw had a significant (p<0.0001) mean increase in sVCAM of 321.9 ng/ml which was significantly greater (p<0.0001) than with IFN-beta-1a 30 mug IM qw (68.6 ng/ml). There was a negative correlation between combined unique (CU) MRI lesions and sVCAM levels within the IFN-beta-1a 44 mug SC tiw group (slope=-0.00106, p=0.009). We postulate that the mode of action of IFN-beta therapy in MS may involve the induction of an increase in sVCAM. sVCAM could bind VLA-4 on T-cells and intercept their adhesion to the blood brain barrier (BBB). This mechanism is consistent with the observed clinical effect of IFN-beta in reducing MRI contrast enhancing lesions.

Brain-derived neurotrophic factor (BDNF) gene delivery into the CNS using bone marrow cells as vehicles in mice.

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, is protective in animal models of neurodegenerative diseases. However, BDNF has a short half-life and its efficacy in the CNS when delivered peripherally is limited due to the blood-brain barrier. In the present study, bone marrow cells were used as vehicles to deliver the BDNF gene into the CNS. Marrow cells obtained from 6 to 8 week-old SJL/J mice were transduced with BDNF expressing pro-virus. RT-PCR analysis revealed that BDNF mRNA was expressed in transduced but not in non-transduced marrow cells. Additionally, virus transduced marrow cells expressed the BDNF protein (296+/-1.2 unit/ml). BDNF-transduced marrow cells were then transplanted into irradiated mice through the tail vein. Three months post-transplantation, significant increases in BDNF as well as glutamic acid decarboxylase (GAD(67)) mRNA were detected in the brains of BDNF transplanted mice compared to untransplanted animals, indicating biological activity of the BDNF transgene. Thus, bone marrow cells can be used as vehicles to deliver the BDNF gene into the brain with implications for the treatment of neurological diseases.

Effect of combined IFNbeta-1a and glatiramer acetate therapy on GA-specific T-cell responses in multiple sclerosis.

The combined treatment with interferon beta (IFNbeta) and glatiramer acetate (GA) is of current interest in multiple sclerosis (MS). The therapeutic effect of GA in MS is believed to be mediated by GA-specific Th2 cells. IFNbeta has a significant anti-proliferative effect on GA-induced lymphoproliferation in vitro. Therefore, we examined the possibility that IFNbeta may interfere with the generation and phenotype of GA T-cell responses in MS patients receiving combined therapy. Sixty-six GA-specific T-cell lines (TCL) were generated ex vivo from five MS patients enrolled in an open-label dinical trial of combined IFNbeta/GA treatment. Controls included 83 pretreatment and 131 on-treatment GA-TCL from 11 MS patients treated with GA only, and five GA-TCL generated from four patients receiving IFNbeta-1a monotherapy. IFNgamma and IL-5 (markers of Th1 and Th2 responses, respectively) were assayed by ELISA in GA-TCL supematants. Th1/Th2 bias was defined by the IFNgamma/IL-5 level ratio ( >2 = Th1 bias, <0.5 = Th2 bias, and 0.5-2 = Th0 bias). The frequency with which GA-reactive TCL were generated was 37.0% for the patients in the combination trial compared to 33.3% in the patients receiving GA alone. The mean stimulation index of the GA-TCL was 8.41 (range 2-42) for the combination compared to a mean of 6.29 (range 2-37) for the GA-treated group--a nonsignificant difference. Mean GA-TCL IFNgamma production was significantly lower in all treatment groups compared to pretreatment IL-5 levels were enhanced in all treatment groups compared to pretreatment levels, but the change was not statistically significant. The Th1/Th0/Th2 distribution of GA-TCL was 7%/30%/63% for the GA+IFNbeta group, 8%/9%/83% for the GA group, compared to 48%/21%/31% pre-GA treatment. All five GA-TCL from the IFNbeta-1a monotherapy patients were Th2-biased. We conclude that IFNbeta-1a does not affect the generation of GA-reactive T cells in vivo. Although more Th0 G4-TCL occurred with combination therapy than with G4 treatment alone, both groups shared an overall Th2 bias. Therefore, we speculate that combined therapy is unlikely to reduce the efficacy of GA treatment in MS.

Sustained immunological effects of Glatiramer acetate in patients with multiple sclerosis treated for over 6 years.

The availability of a group of multiple sclerosis (MS) patients at the University of Maryland, who had participated in the pivotal Copaxone trial in the early 1990s, provided an opportunity to examine the long-term immunologic effects of Glatiramer acetate (GA) treatment in MS. Forty-eight GA-reactive T-cell lines (TCL) were generated from 10 MS patients who have been receiving GA treatment for 6-9 years. Proliferative responses, cytokine production, and cross-reactivity with myelin basic protein (MBP) and the MBP immunodominant peptide 83-99 were compared to responses obtained from 10 MS patients who were tested pretreatment and after a shorter period of treatment ranging from 1 to 10 months. The results indicate that while long-term treatment with GA results in a 2.9-fold decrease in the estimated precursor frequency of GA-reactive T-cells, the sustained response to GA remains Th2-biased and in part cross-reactive with MBP and MBP (83-99) as measured by proliferation and cytokine release assays. The results indicate that despite a drop in the precursor frequency of GA-reactive T-cells with long-term treatment, the sustained response remains predominantly Th2-biased and cross-reactive with MBP, which is consistent with the anti-inflammatory effects of the drug and bystander suppression.

Glatiramer acetate induces a Th2-biased response and crossreactivity with myelin basic protein in patients with MS.

Glatiromer acetate (GA) is an approved treatment for multiple sclerosis (MS). The proposed mechanism of action is the induction of GA-specific T cells characterized by protective anti-inflammatory Th2 response. We tested this hypothesis in 11 MS patients treated with GA from 1-19 months. Interferon-gamma and IL-5 (markers of Th1 and Th2 responses respectively) were assayed by ELISA in GA-specific T-cell lines (TCL) supernatants. Th1/Th2 bias was defined based on the ratio of IFN-gamma/IL-5 secretion. Fifty-eight pre-treatment and 75 on-treatment GA-specific TCL were generated. On-treatment mean IL-5 levels in GA-TCL increased significantly, whereas those for IFN-gamma were markedly reduced. Consequently, the ratio of IFN-gamma IL-5 also shifted in favor of a Th2 response. The percentage of GA-TCL classified as Th1 was decreased, whereas those classified as Th2 increased on-treatment as compared to pre-treatment. Some GA-specific TCL, (approximately 25%) generated during treatment secreted predominantly IL-5 in response to MBP and the immunodominant MBP peptide 83-99, indicating that these crossreactive antigens can act as partial agonists for GA-reactive TCL. These results strongly suggest that the mechanism of action of GA in MS involves the induction of crossreactive GA-specific T cells with a predominant Th2 cytokine profile.

Mechanism of measles virus failure to activate NF-kappaB in neuronal cells.

Lack of IFN-beta and MHC class I expression in measles virus (MV) infected neurons could impair the host antiviral defense mechanism and result in virus escape from recognition by cytotoxic T-cells. Induction of IFN-beta and MHC class I gene expression requires NF-kappaB activation which depends on degradation of IkappaBalpha, an inhibitory protein of NF-kappaB. In earlier studies we demonstrated that in contrast to glial cells, MV was unable to induce IkappaBalpha degradation in neuronal cells. It is unclear whether this failure is due to the presence of a neuron-specific IkappaBalpha isoform or a defect in the MV signaling cascade that leads to IkappaBalpha phosphorylation and degradation. In this study, an IkappaBalpha-wild type (WT) expression vector was transfected into neuronal and glial cells and subsequently exposed to MV. In contrast to glial cells, IkappaBalpha-WT was degraded in neuronal cells in response to TNFalpha but not MV. The findings eliminate the existence of an IkappaBalpha isoform in neuronal cells that is resistant to phosphorylation by MV. Blocking de novo protein synthesis with cyclohexamide had no effect on neuronal IkappaBalpha, indicating that lack of degradation rather than increased synthesis is responsible for IkappaBalpha accumulation in MV-stimulated neuronal cells. To determine if malfunction in the MV receptor CD46 is responsible for failure of IkappaBalpha phosphorylation and degradation, neuronal cells were transfected with a wild type CD46 (CD46-WT) expression vector. MV stimulation of CD46-WT transfected cells failed to induce IkappaBalpha degradation. Collectively these findings indicate that failure of MV to phosphorylate neuronal IkappaBalpha is not due to a presence of an IkappaBalpha isoform or malfunction of the MV receptor, and is more likely to be due to a defect in the signaling pathway that normally leads to IkappaBalpha phosphorylation and degradation.

Mechanisms of immunomodulation by glatiramer acetate.

OBJECTIVE: To define the mechanism of action of glatiramer acetate (GA; formerly known as copolymer-1) as an immunomodulatory treatment for MS. BACKGROUND: The proposed mechanisms of action of GA include 1) functional inhibition of myelin-reactive T cells by human leukocyte antigen (HLA) blocking, 2) T-cell receptor (TCR) antagonism, and 3) induction of T helper 2 (Th2) immunomodulatory cells. In this report, the authors examined the effects of GA on the functional activation of human T-cell clones (TCC) specific for myelin basic protein (MBP) and for foreign antigens. Several questions were addressed: Is the inhibitory effect of GA specific for autoantigens? Is it mediated by blocking the interaction between peptide and HLA molecule? Is GA a partial agonist or TCR antagonist, or does it induce anergy? Does it induce Th2 modulatory T cells? METHODS: The effects of GA on antigen-induced activation of human TCC specific for MBP, influenza virus hemagglutinin, and Borrelia burgdorferi were studied by proliferation and cytokine measurements, TCR downmodulation, and anergy assays. GA-specific TCC were generated in vitro from the peripheral blood of patients and healthy controls by limiting dilution. RESULTS: GA more strongly inhibited the proliferation of MBP, as compared with foreign antigen-specific TCC; in some MBP-specific TCC, the production of Th1-type cytokines was preferentially inhibited. In addition to HLA competition, the induction of anergy, but not direct TCR antagonism, was observed. Numerous GA-specific TCC were generated from the peripheral blood of both MS patients and normal controls, and a fraction of these showed a Th2 phenotype. CONCLUSIONS: This study confirms a preferential inhibitory effect of GA on autoreactive TCC. With respect to cellular mechanisms, although HLA competition appears to play the most important role in functional inhibition in vitro, a direct effect on the TCR may be involved at least in some autoreactive T cells as shown by anergy induction. Although not confirmed at the clonal level, it is demonstrated further that GA induces T cells that crossreact with myelin proteins. GA-specific, Th2-modulatory cells may play an important role in mediating the effect of the drug in vivo.

IFN-beta-1b inhibits IL-12 production in peripheral blood mononuclear cells in an IL-10-dependent mechanism: relevance to IFN-beta-1b therapeutic effects in multiple sclerosis.

IL-12 is a proinflammatory cytokine secreted by dendritic cells in response to microbial Ags and mitogens. IL-12 is thought to contribute to the pathogenesis of autoimmune diseases such as multiple sclerosis (MS). This is based on studies in experimental allergic encephalomyelitis and the demonstration that PBMC IL-12 production correlates with disease progression in MS. IFN-beta-1b is an effective treatment for MS, which is thought to involve in part inhibition of proinflammatory cytokines. In this study we examined the effect of in vitro treatment with IFN-beta-1b, on mitogen-induced IL-12 production in human PBMC and myelin basic protein-specific T cell lines obtained from healthy donors and MS patients. We demonstrate that IFN-beta-1b significantly inhibits inducible IL-12 p40 up to 80% and biologically active IL-12 p70 up to 70% beginning at a dose of 10 IU/ml. This inhibition is IL-10 dependent, as it could be blocked by anti-IL-10 but not anti-IL-4 or control Abs. Thus, endogenously produced IL-10 is a required cofactor for the IFN-beta-1b inhibitory effect on IL-12 to occur. We conclude that IFN-beta-1b has a profound inhibitory effect on PBMC IL-12 production in vitro, and that this effect is IL-10 dependent. These findings are potentially relevant to the therapeutic mechanism of IFN-beta-1b in MS.

The effect of vesnarinone on TNF alpha production in human peripheral blood mononuclear cells and microglia: a preclinical study for the treatment of multiple sclerosis.

Vesnarinone (OPC-8212) is a synthetic quinolinone derivative with inotropic and immunomodulatory effects. Vesnarinone has been shown to inhibit tumor necrosis factor-alpha (TNF alpha) produced by mitogen stimulated macrophages, and to inhibit phosphodiesterase (PDE) type III in cardiac muscle. TNF alpha and interferon-gamma (IFNgamma) have been implicated in the pathogenesis of autoimmune diseases, and both cytokines are targets for therapeutic intervention. IFNgamma can enhance autoimmune disease through direct effects, and indirectly by priming macrophages to produce TNF alpha. In this study, we demonstrate that while vesnarinone enhances basal TNF alpha levels, it inhibits TNF alpha production in peripheral blood mononuclear cells from multiple sclerosis (MS) patients and healthy donors stimulated with lipopolysaccharide (LPS) or primed with IFNgamma and stimulated with suboptimal doses of LPS. In addition, vesnarinone inhibited TNF alpha production in primary adult human microglial cultures. However, in contrast to rolipram, another TNF alpha inhibiting agent, vesnarinone failed to inhibit TNF alpha production by myelin basic protein specific T-cell lines. As oral TNF inhibitors are currently being considered in the USA for clinical application in MS, the implications of our findings on the development of vesnarinone for treatment of MS are discussed.

Failure of measles virus to activate nuclear factor-kappa B in neuronal cells: implications on the immune response to viral infections in the central nervous system.

Neurons are postmitotic cells that foster virus persistence. These cells lack the HLA class I molecules required for clearance of infected cells. Previously, we showed that HLA class I is induced by measles virus (MV) on glial cells, which is primarily mediated by IFN-beta. In contrast, MV was unable to induce HLA class I or IFN-beta in neuronal cells. This failure was associated with lack of NF-kappa B binding to the positive regulatory domain II element of the IFN-beta promoter, which is essential for virus-induced IFN-beta gene activity. In this study, we demonstrate that the failure to activate NF-kappa B in neuronal cells is due to the inability of MV to induce phosphorylation and degradation of I kappa B, the inhibitor of NF-kappa B. In contrast, TNF-alpha induced degradation of I kappa B alpha in the neuronal cells, suggesting that failure to induce I kappa B alpha degradation is likely due to a defect in virus-mediated signaling rather than to a defect involving neuronal I kappa B alpha. Like MV, mumps virus and dsRNA failed to induce I kappa B alpha degradation in the neuronal cells, suggesting that this defect may be specific to viruses. Autophosphorylation of the dsRNA-dependent protein kinase, a kinase possibly involved in virus-mediated I kappa B alpha phosphorylation, was intact in both cell types. The failure of virus to induce I kappa B alpha phosphorylation and consequently to activate NF-kappa B in neuronal cells could explain the repression of IFN-beta and class I gene expression in virus-infected cells. These findings provide a potential mechanism for the ability of virus to persist in neurons and to escape immune surveillance.

Neutralizing antibodies to interferon beta-1a and interferon beta-1b in MS patients are cross-reactive.

OBJECTIVE: To determine whether neutralizing antibodies (NABs) to interferon beta (IFNbeta)-1a (Avonex) and IFNbeta-1b (Betaseron) cross-react. BACKGROUND: A total of 38% of MS patients treated with IFNbeta-1b and 22% of those treated with IFNbeta-1a were reported to develop NABs, which could reduce the clinical efficacy of the drug. METHODS: Blood from 10 MS patients was collected before and at 3 and 6 months after initiating treatment with IFNbeta-1a. ELISA was performed to detect binding antibodies to IFNbeta-1a. Sera from patients who tested positive for binding antibodies to IFNbeta-1a were then screened for NABs to IFNbeta-1a in a biologic assay based on neutralization of antiviral activity. These serum samples were subsequently tested for cross-reactivity with IFNbeta-1b both in the ELISA and the biologic assay. In the second part of the study, sera from patients who participated in the phase III IFNbeta-1b trial at the University of Maryland were examined for cross-reactivity with IFNbeta-1a in the ELISA and the biologic assay. RESULTS: Of the 10 patients treated with IFNbeta-1a, three developed binding as well as NABs to IFNbeta-1a 6 months after treatment, and these antibodies cross-reacted with IFNbeta-1b both in the binding and the biologic assay. Similarly, sera from six patients with NABs to IFNbeta-1b showed cross-reactivity with IFNbeta-1a in the binding assay. Three of these six serum samples tested for neutralizing activity against IFNbeta-1a demonstrated the presence of NABs to IFNbeta-1a. CONCLUSIONS: NABs to IFNbeta-1a (Avonex) and IFNbeta-1b (Betaseron) cross-react, both in the binding and the biologic assays. This suggests that switching to alternate IFNbeta preparation in patients who develop NABs may not be clinically beneficial. Studies examining cross-reactivity between NABs to IFNbeta-1a and IFNbeta-1b in a large number of patients are indicated.

Differential induction of IL-12 by IFN-beta and IFN-gamma in human macrophages.

Interleukin-12 (IL-12) is a proinflammatory cytokine secreted by antigen-presenting cells (APC) in response to microbial antigens and mitogens. IL-12 induces interferon-gamma (IFN-gamma) production and enhances cellular immune responses. Conversely, IFN-gamma does not induce IL-12 but can prime its production by phagocytic cells in response to antigenic stimuli. In this study, we examined the effect of IFN-beta on IL-12 production in human macrophages, as IFN-beta is a natural protein produced by virus-infected cells. We demonstrate that, unlike IFN-gamma, IFN-beta is able to induce IL-12 production in macrophages. However, IFN-gamma can enhance IFN-beta-induced IL-12 in these cells. These findings suggest that IFN-beta could influence the immune response to virus infection indirectly through IL-12.

Serum interferon beta-1a (Avonex) levels following intramuscular injection in relapsing-remitting MS patients.

BACKGROUND: The pharmacokinetics of IFNbeta-1a in MS patients are poorly understood. We have previously reported an ELISA sensitive and specific for measuring serum IFNbeta-1b levels in patients with MS. OBJECTIVE: We describe an ELISA to measure interferon beta-1a (Avonex) in the serum of MS patients following IM administration. METHODS: We have developed an ELISA for detecting serum IFNbeta-1a in MS patients receiving 6 million units (MU) of IFNbeta-1a, IM once weekly. The specificity of this ELISA was confirmed by the lack of cross-reactivity with other cytokines except for IFNbeta-1b. RESULTS: Serum IFNbeta-1a levels were measured at 3 and 6 months after initiating treatment with IFNbeta-1a in 10 MS patients. At 3 months, all 10 patients had detectable levels ranging from 68 to 86 IU/mL. At 6 months, IFNbeta-1a could be detected in the serum of all but three patients, with levels ranging from 64 to 81 IU/mL. A kinetic study of IFNbeta-1a serum levels in a separate group of six MS patients who had been receiving IFNbeta-1a for several months was carried out. Blood was drawn before and 2, 4, 6, 8, and 24 hours after IFNbeta-1a injection. Peak serum IFNbeta-1a levels were observed at 8 hours and became undetectable at 24 hours after injection. CONCLUSION: The described ELISA may have useful clinical applications in examining the correlation between serum IFNbeta-1a levels and clinical efficacy.

Phase 1 trial of transforming growth factor beta 2 in chronic progressive MS.

Transforming growth factor (TGF)-beta2 is a pleiotropic cytokine associated with remissions in multiple sclerosis (MS) and amelioration of allergic encephalomyelitis. We assessed the safety of TGF-beta2 in an open-label trial of 11 patients with secondary progressive (SP) MS. Five patients had a reversible decline in the glomerular filtration rate. There was no change in expanded disability status scale or MRI lesions during treatment. Systemic TGF-beta2 may be associated with reversible nephrotoxicity, and further investigation of its therapeutic potential in MS should be performed with caution.

Immunomodulating functions of recombinant ovine interferon tau: potential for therapy in multiple sclerosis and autoimmune disorders.

The interferons (IFN) are a family of complex proteins possessing antiviral, antiproliferative, and immunomodulatory activities. Two type I recombinant human IFN have been recently approved for the treatment of multiple sclerosis (MS). However, use of high dose type I IFN treatment in MS patients has been limited by dose-related toxicity. Ovine IFN tau is a unique type I interferon discovered for its role in the animal reproductive cycle. It differs from other type I IFNs in that it is remarkably less toxic even at high concentrations, is able to cross species barriers, and is not inducible by viral infection. Ovine IFN tau has been shown to be very effective in the treatment of animal models of MS. In this study, we examined the toxicity of OvIFN tau on human T-cells at high doses and its immunregulatory properties at equivalent doses. Our experiments confirmed the remarkably non-toxic nature of OvIFN tau on human cells at high concentrations as well as immunomodulating properties consistent with other type I IFNs including an antilymphoproliferative effect and inhibition of IFN gamma-induced HLA class II expression. These results suggest that OvIFN tau could be developed into a potentially less toxic therapeutic option for immune-mediated disorders including MS.

Induction of IP-10 chemokine promoter by measles virus: comparison with interferon-gamma shows the use of the same response element but with differential DNA-protein binding profiles.

Measles virus (MV) and interferon (IFN)-gamma induced IP-10 chemokine mRNA in U373 glioblastoma cells. The minimal response element for both MV and IFN-gamma was localized between nucleotide -231 and -153 of muIP-10 promoter, which contains an IFN-stimulated response element (ISRE) and the distal NF-kappa Bd site. Mutation of individual elements showed that ISRE and NF-kappa Bd were required to function together. DNA-protein binding profiles with the minimal response element showed that IFN-gamma induced a complex consisting of STAT1 while MV induced a complex consisting of p50 and p65 in the absence of new protein synthesis. IFN-gamma and MV also induced IRF-1 DNA binding activity which persisted for longer time periods with IFN-gamma stimulation. Despite the functional requirement of both ISRE and NF-kappa Bd elements, different combinations of DNA binding factors are used in the induction of IP-10 by MV or IFN-gamma.

The effect of interferon beta-1b on cytokine-induced adhesion molecule expression.

Interferon beta-1b (IFN beta-1b) (Betaseron) has been recently shown to alter the course of multiple sclerosis (MS), an inflammatory demyelinating disease of the central nervous system (CNS) where migration of activated lymphocytes across the blood-brain barrier (BBB) is a critical step in the pathogenesis of this disease. Magnetic resonance imaging (MRI) studies performed on patients treated with IFN beta-1b showed a remarkable effect on the BBB as determined by a reduction in the number of gadolinium enhancing lesions, a measure of BBB leakiness. Since adhesion molecules (AM) induced on endothelial cells (EC) play an important role in T-cell migration into the CNS, the objective of this study was to examine the effect of IFN beta-1b on the expression of the AM, ICAM-1, V-CAM and E-selection induced on EC by IFN-gamma, TNF-alpha, or IL-1 beta. Primary cultures of human umbilical vein EC (HUVEC) were used, which under basal conditions expressed low levels of AM. IFN beta-1b (1-1000IU/ml) had minimal effect on basal expression of AM on HUVEC, but AM could be substantially upregulated by IFN-gamma, IL-1 beta or TNF-alpha. The effect of IFN beta-1b on AM expression induced by IFN-gamma, IL-1 beta or TNF-alpha was slightly additive. It is concluded that IFN beta-1b does not downregulate the inducible expression of ICAM-1, V-CAM or E-selectin on HUVEC. These findings suggest alternate mechanisms for the effect of IFN beta-1b on the BBB in MS.