Time course of clinical and neuroradiological effects of delayed-release dimethyl fumarate in multiple sclerosis.

BACKGROUND AND PURPOSE: Delayed-release dimethyl fumarate (DMF, also known as gastro-resistant DMF), demonstrated efficacy and safety in relapsing-remitting multiple sclerosis in the 2-year, randomized, placebo-controlled, phase 3 DEFINE and CONFIRM trials. A post hoc analysis of integrated data from DEFINE and CONFIRM was conducted to determine the temporal profile of the clinical and neuroradiological effects of DMF. METHODS: Eligible patients were randomized to receive placebo, DMF 240 mg twice (BID) or three times (TID) daily or glatiramer acetate (GA; reference comparator; CONFIRM only) for up to 96 weeks. Patients in the GA group were excluded from this analysis. RESULTS: A total of 2301 patients were randomized and received treatment with placebo (n = 771) or DMF BID (n = 769) or TID (n = 761). DMF significantly reduced the annualized relapse rate beginning in weeks 0-12 (BID, P = 0.0159; TID, P = 0.0314); the proportion of patients relapsed beginning at week 10 (BID, P = 0.0427) and week 12 (TID, P = 0.0451); and the proportion of patients with 12-week confirmed disability progression beginning at week 62 (BID, P = 0.0454) and week 72 (TID, P = 0.0399), compared with placebo. These effects were sustained throughout the 2-year study period. DMF significantly reduced the odds of having a higher number of gadolinium-enhancing lesions by 88% (BID) and 75% (TID) and the mean number of new or enlarging T2 lesions by 72% (BID) and 67% (TID), from the first post-baseline magnetic resonance imaging assessment at 24 weeks (all P < 0.0001 versus placebo). CONCLUSIONS: In phase 3 clinical trials, DMF demonstrated rapid and sustained clinical and neuroradiological efficacy in relapsing-remitting multiple sclerosis.

Assessment of hepatic inflammation after spinal cord injury using intravital microscopy.

OBJECTIVES: The liver has been shown to play a particularly important role in the initiation and progression of the early systemic inflammatory response (SIR) to spinal cord injury (SCI). The purpose of this study was to determine the time course of leucocyte recruitment to the liver, and to determine the effect of injury severity on the magnitude of leucocyte recruitment and hepatic injury. METHODS: Rats were randomly assigned to one of the following groups: uninjured, sham-injured (laminectomy and no cord injury), cord compressed or cord transected. At 30 min and 90 min after SCI rats had the left lobe of their livers externalised and visualised using intravital video microscopy. RESULTS: Thirty minutes after injury the total number of leucocytes per post-sinusoidal venule was significantly increased after cord transection compared to that in uninjured and sham-injured rats (P<0.05). Of these leucocytes, significantly more were adherent to venule walls (P<0.05). At 90 min the total number of leucocytes per post-sinusoidal venule and the number of adherent and rolling leucocytes was significantly increased after cord transection and cord compression (P<0.05). DISCUSSION: This is the first study to use intravital microscopy to visualise systemic inflammation in the liver following SCI. We have demonstrated immediate leucocyte recruitment to the liver within 30 min after injury and have shown that systemic inflammation increases with time after injury and with severity of injury.

Gender effects on intramuscular interferon beta-1a in relapsing-remitting multiple sclerosis: analysis of 1406 patients.

BACKGROUND: We aimed to evaluate effects of gender on efficacy and safety of intramuscular (IM) interferon beta (IFNß)-1a in patients with relapsing-remitting MS (RRMS) or clinically isolated syndromes (CIS) characteristic of early MS. METHODS: Pooled data from 1406 (1027 women; 379 men) patients enrolled in five clinical studies of IM IFNß-1a were analyzed. One analysis examined data for all patients treated with IM IFNß-1a from all studies. Separate analyses were conducted of pooled IM IFNß-1a-treated groups from all studies and pooled IFNß-1a-treated and placebo-treated patients from the placebo-controlled studies. Outcome measures included time to first relapse, annualized relapse rate, time to disability progression, number of gadolinium-enhanced lesions, adverse events, laboratory evaluations, and neutralizing antibodies. RESULTS: All efficacy assessments indicated similar treatment effects of IM IFNß-1a in men and women with no significant treatment-by-gender interactions. Women reported more headaches, urinary tract infections, and depression in the analysis; however, these were also common in women who received placebo. Men reported more frequent flu-like symptoms in the placebo-controlled studies only. There were no other differences in the safety profile of IM IFNß-1a between men and women. CONCLUSIONS: We conclude that no significant gender-related differences were found in the efficacy and safety of IM IFNß-1a in patients with RRMS or CIS.

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.

The incidence and significance of anti-natalizumab antibodies: results from AFFIRM and SENTINEL.

OBJECTIVE: To determine the incidence and clinical effects of antibodies that develop during treatment with natalizumab. METHODS: In two randomized, double-blind, placebo-controlled studies (natalizumab safety and efficacy in relapsing remitting multiple sclerosis [MS, AFFIRM] and safety and efficacy of natalizumab in combination with interferon beta-1a [INF beta]1a] in patients with relapsing remitting MS [SENTINEL]) of patients with relapsing multiple sclerosis, blood samples were obtained at baseline and every 12 weeks to determine the presence of antibodies against natalizumab. Antibodies to natalizumab were measured using an ELISA. Patients were categorized as "transiently positive" if they had detectable antibodies (>or=0.5 microg/mL) at a single time point or "persistently positive" if they had antibodies at two or more time points >or=6 weeks apart. RESULTS: In the AFFIRM study, antibodies were detected in 57 of 625 (9%) of natalizumab-treated patients: Twenty (3%) were transiently positive and 37 (6%) were persistently positive. Persistently positive patients showed a loss of clinical efficacy as measured by disability progression (p

MRI outcomes in a placebo-controlled trial of natalizumab in relapsing MS.

BACKGROUND: In a 2-year, placebo-controlled trial (the Natalizumab Safety and Efficacy in Relapsing Remitting Multiple Sclerosis [AFFIRM] study), involving 942 patients with relapsing multiple sclerosis (MS), natalizumab significantly reduced the relapse rate by 68% and progression of sustained disability by 42% vs placebo. We report the effect of natalizumab on MRI measures from the AFFIRM study. METHODS: The number and volume of gadolinium (Gd)-enhancing, new or enlarging T2-hyperintense, and new T1-hypointense lesions and brain parenchymal fraction were measured from annual scans obtained at baseline, 1 year, and 2 years. RESULTS: Compared with placebo, natalizumab produced a 92% decrease in Gd-enhancing lesions (means 2.4 vs 0.2; p < 0.001), an 83% decrease in new or enlarging T2-hyperintense lesions (means 11.0 vs 1.9; p < 0.001), and a 76% decrease in new T1-hypointense lesions (means 4.6 vs 1.1; p < 0.001) over 2 years. Median T2-hyperintense lesion volume increased by 8.8% in the placebo group and decreased by 9.4% in the natalizumab group (p < 0.001); median T1-hypointense lesion volume decreased by 1.5% in the placebo group and decreased by 23.5% in the natalizumab group (p < 0.001). Brain atrophy was greater in year 1 and less in year 2 in natalizumab-treated patients. CONCLUSION: Natalizumab has a sustained effect in preventing the formation of new lesions in patients with relapsing multiple sclerosis.

Natalizumab reduces visual loss in patients with relapsing multiple sclerosis.

OBJECTIVE: To examine the effects of natalizumab on low-contrast letter acuity as a prespecified tertiary endpoint in two randomized clinical trials and to evaluate the usefulness of low-contrast letter acuity testing as a candidate test of visual function in multiple sclerosis (MS). METHODS: AFFIRM and SENTINEL were randomized, double-blind, placebo-controlled, multicenter, phase 3 clinical trials of natalizumab in relapsing MS. Natalizumab was evaluated as monotherapy in AFFIRM and as add-on to interferon beta-1a in SENTINEL. Vision testing was performed at 100% contrast (visual acuity) and low-contrast (2.5% and 1.25%). RESULTS: The risk of clinically significant visual loss (predefined as a two-line worsening of acuity sustained over 12 weeks) at the lowest contrast level (1.25%) was reduced in the natalizumab treatment arms by 35% in AFFIRM (hazard ratio = 0.65; 95% CI: 0.47 to 0.90; p = 0.008) and by 28% in SENTINEL (hazard ratio = 0.72; 95% CI: 0.54 to 0.98; p = 0.038, Cox proportional hazards models). Mean changes in vision scores from baseline were also significantly different, reflecting worsening in non-natalizumab groups. CONCLUSIONS: Natalizumab reduces visual loss in patients with relapsing multiple sclerosis. Low-contrast acuity testing has the capacity to demonstrate treatment effects and is a strong candidate for assessment of visual outcomes in future multiple sclerosis trials.

Lactate dehydrogenase-B is silenced by promoter hypermethylation in human prostate cancer.

In order to identify novel candidates associated with prostate cancer metastasis, we compared the proteomic profile of the poorly metastatic human prostate cancer cell line LNCaP, with its highly metastatic variant LNCaP-LN3, by two-dimensional gel electrophoresis. A major protein spot (pI of 5.9 and molecular weight of 37 kDa) was seen in LNCaP cells, but not in LNCaP-LN3 cells and was identified as lactate dehydrogenase-B (LDHB), by tandem mass spectrometry. Furthermore, enzyme kinetic assays and zymography showed a higher LDH enzyme activity in LNCaP cells compared with LNCaP-LN3. Bisulphite-modified DNA sequencing showed promoter hypermethylation in LNCaP-LN3 cells but not in LNCaP, Du145, PC3, CWR22 or BPH45 cells. Treatment of LNCaP-LN3 cells with 5'-azacytidine caused re-expression of LDHB transcripts. In tissues, LDHB promoter hypermethylation occurred at a higher frequency in prostate cancer, 14/ 31 (45%), compared to adjacent nonmalignant or benign tissue, 2/19 (11%) (P < 0.025). Immunohistochemistry showed a higher frequency of LDHB expression in benign or non-malignant tissues, 59/ 73 (81%), compared to cancer cases, 3/53 (6%) (P < 0.001). Absent LDHB expression was also seen in 7/7 (100%) cases of metastatic cancer in bone. Our data are the first to show loss of LDHB expression in prostate cancer, the mechanism of which appears to involve promoter hypermethylation.

Characterizing the mechanisms of progression in multiple sclerosis: evidence and new hypotheses for future directions.

Major advancements have been achieved in our ability to diagnose multiple sclerosis (MS) and to commence treatment intervention with agents that can favorably affect the disease course. Although MS exacerbations and the emergence of disability constitute the more conspicuous aspects of the disease process, evidence has confirmed that most of the disease occurs on a constitutive and occult basis. Disease-modifying therapies appear to be modest in the magnitude of their treatment effects, particularly in the progressive stage of the disease. Therapeutic strategies currently used for MS primarily target the inflammatory cascade. Several potential mechanisms appear to be involved in the progression of MS. Characterizing these mechanisms will result in a better understanding of the various forms of the disorder and how to effectively treat its clinical manifestations. It is our objective within this 2-part series on progression in MS to offer both evidence-based observations and hypothesis-driven expert perspectives on what constitutes the cause of progression in MS. We have chosen areas of inquiry that appear to have been most productive in helping us to better conceptualize the landscape of what MS looks like pathologically, immunologically, neuroscientifically, radiographically, and genetically. We have attempted to advance hypotheses focused on a deeper understanding of what contributes to the progression of this illness and to illustrate new technical capabilities that are catalyzing novel research initiatives targeted at achieving a more complete understanding of progression in MS.

An open-label safety and drug interaction study of natalizumab (Antegren) in combination with interferon-beta (Avonex) in patients with multiple sclerosis.

In this open-label drug-interaction trial, we studied 38 patients with relapsing-remitting multiple sclerosis (MS) who received 3.0 or 6.0 mg/kg of natalizumab as a single intravenous (i.v.) infusion during stable treatment with intramuscular (i.m.) interferon beta-1a 30 microg (IFNbeta-1a; Avonex). To assess the pharmacokinetic (PK) interaction of natalizumab and IFNbeta-1a, serum concentration-time data for both agents were collected and analysed. Biologic response markers of IFNbeta-1a activity, beta2-microglobulin and neopterin, were also assessed to determine effects of natalizumab on IFNbeta-1a pharmacodynamics (PD). Further, safety and immunogenicity were evaluated. The combination of drug therapies was well tolerated. Although natalizumab serum concentrations (and corresponding PK exposure measures) appeared to be somewhat elevated in the presence of IFNbeta-1a, when compared to the same dose (6.0 mg/kg) administered alone in a concurrent comparator study, the differences were generally small and unlikely to be clinically relevant. In general, natalizumab had no apparent clinically relevant effects on the PK or PD properties of IFNbeta-1a. The presence of antibodies to IFNbeta-1a and natalizumab was relatively low. Overall, the study provided safety, immunogenicity, PK and PD data to support a combination strategy for the use of natalizumab and IFNbeta-1a in the treatment of patients with relapsing-remitting MS. A large clinical study is currently in progress to evaluate the efficacy and long-term safety of this combination drug therapy.

Disease modifying agent related skin reactions in multiple sclerosis: prevention, assessment, and management.

BACKGROUND: The objective for this article is to highlight some of the adverse skin manifestations associated with injectable disease modifying therapy for multiple sclerosis (MS). Early identification and intervention can often lead to minimal consequences and prolonged patient tolerance and compliance with these agents. At the University of Texas Southwestern Medical Center at Dallas and Texas Neurology in Dallas we actively follow approximately 5000 MS patients. The majority of our patients with relapsing-remitting MS (RRMS) or secondary progressive MS (SPMS) are treated with one of the currently available disease modifying agents (DMAs). Our experience with these patients, and the challenges they face in continuing long-term treatment, constitutes the basis of our proposed treatment strategies. CONCLUSION: Skin reactions in response to injectable DMA therapy in MS are generally mild. However, some reactions can evolve into potentially serious lesions culminating in infection, necrosis, and in some circumstances requiring surgical repair.

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.

Quantitative oculographic characterisation of internuclear ophthalmoparesis in multiple sclerosis: the versional dysconjugacy index Z score.

BACKGROUND: There is a poor correlation between multiple sclerosis disease activity, as measured by magnetic resonance imaging, and clinical disability. OBJECTIVE: To establish oculographic criteria for the diagnosis and severity of internuclear ophthalmoparesis (INO), so that future studies can link the severity of ocular dysconjugacy with neuroradiological abnormalities within the dorsomedial brain stem tegmentum. METHODS: The study involved 58 patients with multiple sclerosis and chronic INO and 40 normal subjects. Two dimensional infrared oculography was used to derive the versional dysconjugacy index (VDI)-the ratio of abducting to adducting eye movements for peak velocity and acceleration. Diagnostic criteria for the diagnosis and severity of INO were derived using a Z score and histogram analysis, which allowed comparisons of the VDI from multiple sclerosis patients and from a control population. RESULTS: For a given saccade, the VDI was typically higher for acceleration v velocity, whereas the Z scores for velocity measures were always higher than values derived from comparable acceleration VDI measures; this was related to the greater variability of acceleration measures. Thus velocity was a more reliable measure from which to determine Z scores and thereby the criteria for INO and its level of severity. The mean (SD) value of the VDI velocity derived from 40 control subjects was 0.922 (0.072). The highest VDI for velocity from a normal control subject was 1.09, which was 2.33 SD above the normal control mean VDI. We therefore chose 2 SD beyond this value (that is, a Z score of 4.33) as the minimum criterion for the oculographic confirmation of INO. Of patients thought to have unilateral INO on clinical grounds, 70% (16/23) were found to have bilateral INO on oculographic assessment. CONCLUSIONS: INO can be confirmed and characterised by level of severity using Z score analysis of quantitative oculography. Such assessments may be useful for linking the level of severity of a specific clinical disability with neuroradiological measures of brain tissue pathology in multiple sclerosis.

Polypharmacy in multiple sclerosis.

The goal of this article is to describe therapeutic approaches to multiple sclerosis (MS) which may be applicable to other human conditions such as epilepsy. Polypharmacy is very commonly employed in MS and it is likely to increase in the future. This paper is written with the premise that therapeutic principles and practices underlying the care of patients with MS may be useful for the treatment of people with epilepsy. We will point out some of the analogies between MS and epilepsy and we will describe the polypharmacy approach employed to treat patients with MS. We will review current information as to the etiology and pathogenesis of MS and describe potential therapeutic targets derived from this knowledge. We will identify or suggest certain therapeutic principles underlying polypharmacy in MS which may be relevant to epilepsy or other conditions. Finally, we will describe some of the obstacles to expanding rational polypharmacy and how we might overcome these problems.

Modification of acute experimental autoimmune encephalomyelitis in the Lewis rat by oral administration of type 1 interferons.

The effect of orally administered type 1 interferons on the severity of acute experimental autoimmune encephalomyelitis (EAE), a T cell-mediated autoimmune disease, was examined by inoculation of Lewis rats with guinea pig myelin basic protein (GPMBP) and complete Freund's adjuvant. Rats were fed either rat species-specific or human recombinant type 1 interferon (IFN) or mock IFN daily for 7 days preceding immunization and for 21 days thereafter. There was a significant decrease in the clinical score and inflammatory foci in animals fed 5000 units IFN compared with mock-treated animals. There was a significant decrease in clinical score and number of inflammatory foci in spinal cord in animals fed orally 5000 units human recombinant IFN-alpha PO compared with SC 5000 units recombinant human IFN-alpha. Oral administration of type 1 interferon, as opposed to subcutaneous administration, inhibited the secretion of IFN-gamma from ConA-activated draining popliteal lymph node cells compared with mock-fed animals. These experiments demonstrate that acute EAE is more effectively inhibited by equivalent amounts of orally in contrast to parenterally administered IFN-alpha. These results suggest that type 1 IFNs are active by the oral route and have significant clinical and histologic effects in acute autoimmune disease.

Prolactin stimulates proliferation of cultured human keratinocytes.

The effect of the pituitary hormone prolactin on in vitro proliferation of human keratinocytes has been studied. Cell proliferation was determined by [3H] thymidine incorporation and cell enumeration in culture. Physiologic concentrations of prolactin markedly stimulated proliferation of newborn foreskin keratinocytes in serum-free medium. In addition, it was able to replace almost completely the growth-promoting effects of bovine pituitary extract, a commonly added supplement for keratinocyte culture. This activity was also evident in the absence of epidermal growth factor, but required the presence of insulin. Radioligand-binding studies confirmed the expression of specific prolactin binding sites (Kd 8.9 nM; 1350 sites per cell) on freshly procured keratinocyte membranes. These results extend its hormonal influences to include regulation of in vitro proliferation of human keratinocytes, and suggest the possibility of a completely defined growth medium for keratinocytes.

A clinical therapeutic trial of cyclosporine in myasthenia gravis.

We randomly assigned 39 patients with steroid-dependent generalized myasthenia gravis to treatment with cyclosporine (5 mg/kg per body weight in divided doses) or placebo. Duration of treatment was 6 months. Patients were evaluated monthly. Primary measures of efficacy were quantified strength testing, antihuman acetylcholine receptor antibody titer, and dosage of corticosteroid medication. At the end of the study, patients in the cyclosporine group had significantly greater improvement in strength (p = 0.004) and a reduction in antireceptor antibody titer (p = 0.01). Percentage reduction of steroid medication was greater in the cyclosporine group, although the difference was not statistically significant (p = 0.12). There were no treatment failures, and there was one drug failure in the cyclosporine group. In the placebo group, there were three treatment failures. No significant nephrotoxicity was noted at this dosage during the first 6 months. During the subsequent 18 months of open-label therapy, continued reduction in steroid dosage occurred. Cumulative side effects, however, caused 35% of patients to discontinue the medication; 10% did so secondary to slowly progressive nephrotoxicity.

Prolactin receptor expression by lymphoid tissues in normal and immunized rats.

Prolactin receptor (PRLr) expression and distribution in thymus, spleen, bone marrow, lymph nodes, and peripheral blood lymphocytes from young adult Lewis rats are analyzed using single-color flow cytometry and a well-characterized monoclonal antibody directed against the rat liver PRLr. The in vivo effects of regional immunization on PRLr expression are also examined. PRLr is found to be widely distributed among cells of the immune system and demonstrates lymphoid tissue-specific patterns of expression. Footpad immunization caused the rapid, but transient, induction of PRLr expression in the draining lymph node, with only modest effects on PRLr expression in other distant lymphoid tissues. These studies indicate that PRL may be capable of direct interaction with the immune system through differential expression of the PRL cell surface receptor on select lymphoid target cell populations.

Adoptive transfer of experimental allergic encephalomyelitis using serum-free, chemically defined in vitro culture conditions.

Enhanced adoptive transfer of experimental allergic encephalomyelitis (EAE) in rats requires in vitro culture of encephalitogen-sensitized donor spleen cells with either myelin basic protein or T-cell mitogen for 18-72 h prior to transfer to unimmunized recipients. The required in vitro culture period offers an opportunity to address in detail cellular and molecular immunoregulatory processes involved in the development of EAE. Conventional culture conditions using fetal bovine serum may impose analytical limitations due to the chemical complexity of the media. To permit better definition of the chemical events associated with the development of EAE, we report the successful adoptive transfer of EAE in Lewis rats using completely chemically defined, serum-free culture conditions.