Electrocardiographic assessments and cardiac events after fingolimod first dose - a comprehensive monitoring study.

BACKGROUND: First dose observation for cardiac effects is required for fingolimod, but recommendations on the extent vary. This study aims to assess cardiac safety of fingolimod first dose. Individual bradyarrhythmic episodes were evaluated to assess the relevance of continuous electrocardiogram (ECG) monitoring. METHODS: START is an ongoing open-label, multi-center study. At the time of analysis 3951 patients were enrolled. The primary endpoints are the incidence of bradycardia (heart rate < 45 bpm) and second-/third-degree AV blocks during treatment initiation. The relevance of Holter was assessed by matching ECG findings with the occurrence of clinical symptoms as well as by rigorous analysis of AV blocks with regard to the duration of pauses and the minimal heart rate recorded during AV block. RESULTS: Thirty-one patients (0.8%) developed bradycardia (<45 bpm), 62 patients (1.6%) had second-degree Mobitz I and/or 2:1 AV blocks with a lowest reading (i.e. mean of ten consecutive beats) of 35 bpm and the longest pause lasting for 2.6 s. No Mobitz II or third-degree AV blocks were observed. Only one patient complained about mild chest discomfort and fatigue. After 1 week, there was no second-/third-degree AV block. CONCLUSIONS: Continuous Holter ECG monitoring in this large real-life cohort revealed that bradycardia and AV conduction abnormalities were rare, transient and benign. No further unexpected abnormalities were detected. The data presented here give an indication that continuous Holter ECG monitoring does not add clinically relevant value to patients' safety. TRIAL REGISTRATION: NCT01585298 ; registered April 23, 2012.

Beta-amyloid peptide variants in brains and cerebrospinal fluid from amyloid precursor protein (APP) transgenic mice: comparison with human Alzheimer amyloid.

In this study, we report a detailed analysis of the different variants of amyloid-ß (Aß) peptides in the brains and the cerebrospinal fluid from APP23 transgenic mice, expressing amyloid precursor protein with the Swedish familial Alzheimer disease mutation, at different ages. Using one- and two-dimensional gel electrophoresis, immunoblotting, and mass spectrometry, we identified the Aß peptides Aß(1-40), -(1-42), -(1-39), -(1-38), -(1-37), -(2-40), and -(3-40) as well as minor amounts of pyroglutamate-modified Aß (Aß(N3pE)) and endogenous murine Aß in brains from 24-month-old mice. Chemical modifications of the N-terminal amino group of Aß were identified that had clearly been introduced during standard experimental procedures. To address this issue, we additionally applied amyloid extraction in ultrapure water. Clear differences between APP23 mice and Alzheimer disease (AD) brain samples were observed in terms of the relative abundance of specific variants of Aß peptides, such as Aß(N3pE), Aß(1-42), and N-terminally truncated Aß(2/3-42). These differences to human AD amyloid were also noticed in a related mouse line transgenic for human wild type amyloid precursor protein. Taken together, our findings suggest different underlying molecular mechanisms driving the amyloid deposition in transgenic mice and AD patients.

Optimisation of BACE1 inhibition of tripartite structures by modification of membrane anchors, spacers and pharmacophores - development of potential agents for the treatment of Alzheimer's disease.

Systematic variation of membrane anchor, spacer and pharmacophore building blocks leads to an optimisation of the inhibitory effect of tripartite structures towards BACE1-induced cleavage of the amyloid precursor protein (APP).

A method to prevent cross contamination during 2-DE by ß-amyloid peptides.

A method for the efficient decontamination of aluminium oxide ceramic 2-DE focusing trays from ß-amyloid peptides (Aß) is reported. As these contaminations were resistant to the standard cleaning procedures, additional harsh cleaning steps were necessary for their efficient removal. Our observations suggest that specific surface properties affect the degree of adsorption of the Aß-peptides. "Surface catalysed amyloid aggregation" in the aluminium oxide ceramic trays is proposed as a possible underlying mechanism for the occurrence of proteinase K-resistant forms of Aß.

Structural design, solid-phase synthesis and activity of membrane-anchored ß-secretase inhibitors on Aß generation from wild-type and Swedish-mutant APP.

Covalent coupling of ß-secretase inhibitors to a raftophilic lipid anchor via a suitable spacer by using solid-phase peptide synthesis leads to tripartite structures displaying substantially improved inhibition of cellular secretion of the ß-amyloid peptide (Aß). Herein, we describe a series of novel tripartite structures, their full characterization by NMR spectroscopy and mass spectrometry, and the analysis of their biological activity in cell-based assays. The tripartite structure concept is applicable to different pharmacophores, and the potency in terms of ß-secretase inhibition can be optimized by adjusting the spacer length to achieve an optimal distance of the inhibitor from the lipid bilayer. A tripartite structure containing a transition-state mimic inhibitor was found to be less potent on Aß generation from Swedish-mutant amyloid precursor protein (APP) than from the wild-type protein. Moreover, our observations suggest that specific variants of Aß are generated from wild-type APP but not from Swedish-mutant APP and are resistant to ß-secretase inhibition. Efficient inhibition of Aß secretion by tripartite structures in the absence of appreciable neurotoxicity was confirmed in a primary neuronal cell culture, thus further supporting the concept.

Patient satisfaction and healthcare services in specialized multiple sclerosis centres in Germany.

BACKGROUND: As patients with multiple sclerosis (MS) require lifelong treatment, optimization of therapy with respect to efficacy and safety is needed to limit long-term disease progression. Patients with MS also need a range of health-related services. Satisfaction with these as well as treatment is clinically relevant because satisfied patients are more likely to adhere to therapy. The aim of this study was to determine the status of patient satisfaction and of healthcare services in 70 specialized MS centres in Germany. METHODS: In 2011, patients with MS responded to a questionnaire, which solicited clinical and demographic information, as well as patients' perceptions of their overall situation and their satisfaction with treatment. RESULTS: Of 2791 patients surveyed, 81.9% had relapsing-remitting MS with mild disability [mean (standard deviation) Expanded Disability Status Scale score: 2.6 (1.8)]. Disease activity data were collected from 2205 patients, of whom 57.6% had remained relapse-free during the preceding 12 months. However, 38.9% had experienced one or more relapses, most of whom (67.3%) while receiving immunomodulatory treatment. About one-third of the patients indicated that they were more dissatisfied with their overall situation compared with the time before diagnosis. However, many patients (58.3%) were satisfied with their existing medication. Overall, 72.8% of patients would prefer oral to injectable treatments, assuming there was no difference in their efficacy. CONCLUSIONS: A substantial proportion of patients experienced breakthrough disease on treatment and may potentially benefit from a change of therapy. Although largely satisfied with treatment, most patients with MS would choose oral over injectable treatments.

Evidence for Elevated Cerebrospinal Fluid ERK1/2 Levels in Alzheimer Dementia.

Cerebrospinal fluid (CSF) samples from 33 patients with Alzheimer dementia (AD), 21 patients with mild cognitive impairment who converted to AD during followup (MCI-AD), 25 patients with stable mild cognitive impairment (MCI-stable), and 16 nondemented subjects (ND) were analyzed with a chemiluminescence immunoassay to assess the levels of the mitogen-activated protein kinase ERK1/2 (extracellular signal-regulated kinase 1/2). The results were evaluated in relation to total Tau (tTau), phosphorylated Tau (pTau), and beta-amyloid 42 peptide (Aß42). CSF-ERK1/2 was significantly increased in the AD group as compared to stable MCI patients and the ND group. Western blot analysis of a pooled cerebrospinal fluid sample revealed that both isoforms, ERK1 and ERK2, and low amounts of doubly phosphorylated ERK2 were detectable. As a predictive diagnostic AD biomarker, CSF-ERK1/2 was inferior to tTau, pTau, and Aß42.

Spred-2 steady-state levels are regulated by phosphorylation and Cbl-mediated ubiquitination.

Spred proteins modulate growth factor receptor signaling by inhibiting the Ras-MAPK cascade. Here, we show that Spred-1, Spred-2, and Spred-3 are ubiquitinated in HEK293T cells stimulated with epidermal growth factor (EGF) or pervanadate. Spred-2 tyrosines Y228 and/or Y231 in the Kit binding domain were identified as putative phosphorylation site(s) critical for Spred-2 ubiquitination. Depletion of Cbl and Cbl-b E3 ubiquitin ligases by RNA interference, or overexpression of a Cbl dominant inhibitory mutant (Cbl-N), inhibited Spred-2 ubiquitination, while conversely, wild type Cbl enhanced Spred-2 ubiquitination. Interaction of Spred-2 with Cbl-N was detectable by co-immunoprecipitation and required the Cbl SH2 domain and Spred-2 Y228 and Y231 residues. Studies on endogenous Spred-2 in ME4405 melanoma cells showed that pervanadate induced Spred-2 ubiquitination and a marked reduction in Spred-2 steady-state levels that was partially blocked by the proteasomal inhibitor, MG-132. These results suggest a role for Spred-2 tyrosine phosphorylation and ubiquitination in controlling Spred-2 expression levels.