Genomic effects of once-weekly, intramuscular interferon-beta1a treatment after the first dose and on chronic dosing: Relationships to 5-year clinical outcomes in multiple sclerosis patients.

PURPOSE: To characterize gene expression in multiple sclerosis (MS) patients after the first dose and chronic dosing of 30 microg, once weekly, intramuscular interferon-beta1a (IFN-beta) and to delineate the pharmacogenomic differences between Good Responders and Partial Responders to IFN-beta therapy. METHODS: The treatment responses after the first IFN-beta dose and chronic IFN-beta dosing were assessed in 22 relapsing MS patients (17 females, 5 males; average age: 41.5+/-SD 10.4 years). Gene expression profiles in peripheral blood mononuclear cells were obtained prior to treatment and at 1, 2, 4, 8, 24, 48, 120, 168 h after the first IFN-beta dose and at 1, 6 and 12 months after chronic dosing with once-weekly 30 microg IFN-beta-1a intramuscularly. Repeated measures statistics with false discovery rate control were used. The functional characteristics, biological pathways and transcription factor sites were analyzed. RESULTS: Of the 1000 genes modulated following the first dose and upon chronic dosing of IFN-beta in MS patients, approximately 35% were up-regulated and 65% were down- regulated; the percentage of modulated genes in common was approximately 50%. The expression of the pharmacodynamic mRNA markers of IFN-beta effect showed differences in time profiles for the Good Responder and Partial Responders to IFN-beta therapy and the Jak-STAT, TNFRSF10B, IL6, TGFbeta, retinoic acid and CDC42 pathways were differentially modulated. The patients with side effects to therapy showed differences in the TGFbeta1, IFNG/STAT3 and TNF pathways. CONCLUSIONS: Gene expression is a valuable tool for understanding the molecular mechanisms of IFN-beta action in MS patients.

Immune cell BDNF secretion is associated with white matter volume in multiple sclerosis.

PURPOSE: To investigate the relationship between immune cell secretion of brain-derived neurotrophic factor (BDNF) with clinical and MRI variables in multiple sclerosis (MS) patients. BACKGROUND: BDNF exerts beneficial effects on neuronal growth and repair and is secreted by both neurons and immune cells. Consequently, it may mediate the crosstalk between the immune system and CNS in autoimmune diseases such as MS. METHODS: Fifty-two relapsing MS patients (41 females, age: 48.8+/-6.6 years, disease duration: 12.7+/-8.4 years) were enrolled. Clinical and MRI measurements (including, T1-, T2- and contrast-enhancing (CE) lesion volumes (LVs); normalized measures of whole brain, white matter (WM) and gray matter (GM) volumes; diffusion weighted imaging measure of mean whole brain (WB) parenchyma diffusivity and magnetization transfer ratio (MTR) measures were obtained. RESULTS: Immune cell BDNF secretion after anti-CD3 plus anti-CD28 stimulation was positively associated with increased CE-LV (p=0.026). The MTR of CE-LV and normal-appearing (NA) WM (NAWM) were negatively associated with immune cell BDNF secretion after anti-CD3 plus anti-CD28 stimulation. Immune cell BDNF secretion after anti-CD3 plus anti-CD28 was positively associated with higher WM volume (p=0.027). Immune cell BDNF secretion after anti-CD3 plus anti-CD28 stimulation was decreased with increasing disease duration (p=0.031). The BDNF secretion was independent of the BDNF Val66Met (dBSNP ID: rs6265) SNP genotype. CONCLUSIONS: Immune cell BDNF secretion is associated with the sites of higher inflammatory activity as evidenced by CE lesions and may represent an important factor associated with the WM volume of patients with MS.

Interferon inhibitory activity in patients with multiple sclerosis.

BACKGROUND: Interferon inhibitory activity (IIA) is a logical candidate for explaining neutralizing antibody-negative partial responsiveness to interferon beta in multiple sclerosis (MS), but its role has not been evaluated. OBJECTIVE: To investigate the role of IIA and soluble interferon-alpha/beta receptor (sIFNR) in determining response of patients with MS to interferon beta therapy. DESIGN: Parallel-group, open-label study. SETTING: Baird Multiple Sclerosis Center, Buffalo, NY. Patients Blood was obtained before and 24 hours after injection of interferon beta-1a from 38 anti-interferon beta neutralizing antibody-negative patients with relapsing-remitting MS and 16 untreated healthy controls. On the basis of clinical parameters of response to interferon beta therapy, the patients were divided into stable or good-responder (n = 20) and active or partial-responder (n = 18) groups. MAIN OUTCOME MEASURES: Quantitative analyses of magnetic resonance imaging were obtained; the IIA and sIFNR levels were measured using bioassay and enzyme-linked immunosorbent assay, respectively. RESULTS: The IIA and sIFNR levels were elevated in MS patients compared with controls (P<.001). The IIA levels were higher in active or partial responders compared with stable or good responders (P<.001); the sIFNR levels were not different between groups. The Extended Disability Status Score and T2 lesion volumes were higher in the active or partial-responder group compared with the stable or good-responder group. Interferon beta-1a did not have short-term effects on the IIA and sIFNR levels. In univariate general linear model and stepwise regression analyses, IIA levels were associated with T2 lesion volume. CONCLUSION: The levels of IIA are associated with increased MS disease activity and with responsiveness to interferon beta therapy in anti-interferon beta neutralizing antibody-negative MS patients.

Interferon-beta modulates bone-associated cytokines and osteoclast precursor activity in multiple sclerosis patients.

PURPOSE: Multiple sclerosis (MS) patients have a high risk of low bone density. The purpose of this study was to examine the molecular mechanisms potentially capable of modulating bone homeostasis in response to interferon-beta-1a (IFN-beta-1a) treatment and the focus was the bone-modulating system comprised of receptor activator of nuclear factor-kappaB (RANK), its ligand RANKL and its decoy receptor, osteoprotegerin (OPG). METHODS: In this open-label pharmacodynamic study, peripheral blood was obtained from relapsing-remitting MS patients just prior to and at multiple time points after intramuscular injection of 30 microg IFN-beta-1a. Samples were analysed for RANKL, tumour necrosis factor related apoptosis-inducing ligand (TRAIL), OPG and macrophage inflammatory protein-1 alpha/beta expression. Osteoclast precursor differentiation from peripheral blood cells of MS patients in the presence of exogenously added IFN-beta-1a was also assessed. Additionally, the changes in plasma levels of osteocalcin and the C-telopeptides after 1 year of treatment were measured as surrogate markers of bone formation and degradation, respectively. RESULTS: IFN-beta-1a treatment modulated RANKL and OPG in a selective, time-dependent manner. The levels of OPG protein decreased 25% at the 8-h time point, then increased 43% at the 24-h time point. The levels of free RANKL reached a maximum at the 8-h time point. Increases in the levels of macrophage inflammatory protein-1beta (MIP-1beta), a chemokine that increases osteolysis, were observed. The levels of the bone formation marker, osteocalcin, were lower in MS patients compared to controls and increased after one year of treatment. Ex vivo treatment of peripheral blood lymphocytes with IFN-beta resulted in a marked reduction of osteoclast-like cells in the presence of RANKL and macrophage colony stimulating factor. CONCLUSIONS: IFN-beta treatment induces complex, specific and time-dependent changes in multiple proteins and mRNAs related to bone homeostasis in MS patients.

Study of mitoxantrone for the treatment of recurrent neuromyelitis optica (Devic disease).

BACKGROUND: Neuromyelitis optica is a severe demyelinating disease that selectively involves the optic nerves and the spinal cord but usually spares the brain. It is considered to have a B-cell-induced pathogenesis. Mitoxantrone hydrochloride, a synthetic anthracenedione approved for worsening relapsing-remitting multiple sclerosis and secondary progressive multiple sclerosis, has been shown to primarily suppress the humoral response. OBJECTIVE: To evaluate the benefit of mitoxantrone treatment in patients with relapsing neuromyelitis optica. DESIGN: Prospective 2-year study. SETTING: Academic multiple sclerosis center. PATIENTS: Five patients (3 women and 2 men) with an age range of 20 to 51 years and an Expanded Disability Status Scale score of 2.5 to 6.5 (mean +/- SD, 4.40 +/- 1.88). INTERVENTIONS: Monthly intravenous infusions of mitoxantrone hydrochloride, 12 mg/m2, for 6 months followed by 3 additional treatments every 3 months. MAIN OUTCOME MEASURES: Expanded Disability Status Scale score measured every 3 months and during relapses; findings on orbital, brain, and spinal cord magnetic resonance images performed at baseline and at 3, 6, 12, 18, and 24 months; and visual evoked potentials and results of ophthalmologic evaluations performed at baseline and annually. RESULTS: During the 2 years of treatment, 2 patients each had a relapse once within the initial 5 months of treatment (1 severe and 1 moderate). Improvement was seen clinically and on magnetic resonance images in 4 patients. Patients generally tolerated the treatment well, although 1 patient had a reversible decrease in cardiac ejection fraction. CONCLUSION: Our results suggest a beneficial effect of mitoxantrone treatment for relapsing neuromyelitis optica.

Low fat dietary intervention with omega-3 fatty acid supplementation in multiple sclerosis patients.

OBJECTIVES: To determine whether a low fat diet supplemented with omega-3 positively affects quality of life (QOL) in relapsing-remitting MS (RRMS) patients. In this 1-year long double-blind, randomized trial, patients were randomized to two dietary interventions: the "Fish Oil" (FO) group received a low fat diet (15% fat) with omega-3 FOs and the "Olive Oil" (OO) group received the AHA Step I diet (fat 30%) with OO supplements. The primary outcome measure was the Physical Components Summary Scale (PCS) of the Short Health Status Questionnaire (SF-36). Additional measures using MS specific QOL questionnaires, neurological status and relapse rate were obtained. RESULTS: 31 RRMS patients were enrolled, with mean follow up over 11 +/- SD 2.9 months. Clinical benefits favoring the FO group were observed on PCS/SF-36 (P = 0.050) and MHI (P = 0.050) at 6 months. Reduced fatigue was seen on the OO diet at 6 months (P = 0.035). The relapse rate decreased in both groups relative to the rates during the 1 year preceding the study: mean change in relapse rate in the FO group: -0.79 +/- SD 1.12 relapses/year (P = 0.021) vs. -0.69 +/- SD 1.11 (P = 0.044) in the OO group. This study suggests that a low fat diet supplemented with omega-3 PUFA can have moderate benefits in RRMS patients on concurrent disease modifying therapies.

Risk of bone loss in men with multiple sclerosis.

CONTEXT: Osteoporosis and the increased fracture risk associated with osteoporosis become apparent in men approximately 10 years later than women. However, in recent studies, approximately 20% of healthy men in the age range 55-64 years were found to be osteopenic. Emerging data suggest a significantly increased prevalence of osteoporosis in men and women with multiple sclerosis (MS) compared to age-matched controls, but no specific clinical testing recommendations are available for men. OBJECTIVE: To determine the proportion of male MS patients with osteoporosis and to identify the factors associated with the reduction in bone mass. DESIGN: Consecutive male MS patients seen at our MS clinic were screened with dual-X-ray absorptiometry (DEXA) scan for determining the bone mineral density (BMD). All patients had neurological Expanded Disability Status Scale (EDSS) evaluation. The results were compared to healthy age-matched male reference population using the Z score and to a cohort of women MS patients and women controls. Calcium, total testosterone, sex-hormone binding globulin (SHBG), 25-hydroxy-vitamin-D, and parathyroid hormone (PTH) were evaluated in male patients with decreased BMD. Relevant data on body mass index (BMI), medication, alcohol consumption, smoking, and sexual dysfunction were recorded. SETTING: Academic MS Centre. PATIENTS AND OTHER PARTICIPANTS: Forty consecutive male MS patients, age mean 51.2 +/- 8.7 years, and mean EDSS of 5.8 +/- 1.9 were evaluated with DEXA scan. Of these, 17.5% patients were relapsing-remitting (RR) MS, 57.5% were secondary progressive (SP) MS and 25% were primary progressive (PP) MS. MAIN OUTCOME MEASURE: Proportion of male MS patients with reduced BMD at the lumbar spine and femoral neck. RESULTS: Thirty-two (80%) of our patients had a reduced bone mass of either lumbar spine or the femoral neck; of these 17 patients (42.5%) had osteopenia and 15 patients (37.5%) had osteoporosis. Twenty-one per cent (eight out of 38 patients) had vertebral, rib or extremities fractures. Multivariate linear regression analysis indicated that the EDSS (P < 0.0001) and BMI (P = 0.0004) were the important factors associated with low BMD at the femoral neck and the EDSS was the important factor (P = 0.0017) associated with low BMD at the lumbar spine. The same factors emerged as significantly associated with the corresponding Z scores, which are corrected for age and sex. No clear association between intravenous steroid therapy and BMD was evident in the multivariate analysis. Low levels of 25-hydroxy-vitamin-D were seen in 37.5% of patients. CONCLUSIONS: The proportion of male MS patients with reduced bone mass is high and disproportionate to their age and ambulation, consistent with an association between the MS disease process and pathological bone loss. Increased awareness and bone density screening of male and female MS patients over 40 years of age is warranted.

Genomic effects of IFN-beta in multiple sclerosis patients.

The purpose of this report was to characterize the dynamics of the gene expression cascades induced by an IFN-beta-1a treatment regimen in multiple sclerosis patients and to examine the molecular mechanisms potentially capable of causing heterogeneity in response to therapy. In this open-label pharmacodynamic study design, peripheral blood was obtained from eight relapsing-remitting multiple sclerosis patients just before and at 1, 2, 4, 8, 24, 48, 120, and 168 h after i.m. injection of 30 micro g of IFN-beta-1a. The total RNA was isolated from monocyte-depleted PBL and analyzed using cDNA microarrays containing probes for >4000 known genes. IFN-beta-1a treatment resulted in selective, time-dependent effects on multiple genes. The mRNAs for genes implicated in the anti-viral response, e.g., double-stranded RNA-dependent protein kinase, myxovirus resistance proteins 1 and 2, and guanylate binding proteins 1 and 2 were rapidly induced within 1-4 h of IFN-beta treatment. The mRNAs for several genes involved in IFN-beta signaling, such as IFN-alpha/beta receptor-2 and Stat1, were also increased. The mRNAs for lymphocyte activation markers, such as IFN-induced transmembrane protein 1 (9-27), IFN-induced transmembrane protein 2 (1-8D), beta(2)-microglobulin, and CD69, were also increased in a time-dependent manner. The findings demonstrate that IFN-beta treatment induces specific and time-dependent changes in multiple mRNAs in lymphocytes of multiple sclerosis patients that could provide a framework for rapid monitoring of the response to therapy.

Dynamics of immune cell trafficking in interferon-beta treated multiple sclerosis patients.

PURPOSE: To investigate the effects of interferon-beta-1a (IFN-beta-1a) on the trafficking of cell populations in peripheral blood cells of multiple sclerosis (MS) patients. METHODS: In this open-label pharmacodynamic study, peripheral blood was obtained from 10 relapsing-remitting (RR) MS patients just prior to and at 1, 2, 4, 8, 24, 48, 120, and 168 h after intramuscular injection of 30-microg IFN-beta-1a. Timed samples were also obtained from five controls at 0, 8, 24, 48 and 168 h. The blood cells were analyzed using four-color flow cytometry with antibody conjugates directed against cell surface proteins specific for T cells, B cells, NK cells, and the activation marker, CD69. RESULTS: IFN-beta-1a treatment resulted in selective, time-dependent effects on many cell populations in peripheral blood. The trafficking of T-helper and T-suppressor/cytotoxic subsets of T cells were qualitatively different. The most prominent effects were on the trafficking of natural killer cells, the levels of which decreased to 23.5% of pretreatment values at 8 h after treatment. The levels of CD69-positive NK cells increased to a peak value of 606% of pretreatment levels at the 24-h time point. In untreated controls, these characteristic trafficking effects were not observed. There was inter-patient heterogeneity in the levels of activated NK cells at the 6-month time point that may potentially be relevant for individualizing IFN-beta therapy. CONCLUSIONS: IFN-beta treatment can induce specific, selective, and time-dependent trafficking of cells and its effects on different subsets of a given cell type are not qualitatively similar. The dynamics indicate that the activation of NK cells by IFN-beta is possibly dependent on the trafficking of NK cells. The activated NK cell levels after prolonged therapy may potentially provide a surrogate marker for IFN-beta exposure.