A mechanistic study to assess whether isoproterenol can reverse the negative chronotropic effect of fingolimod.

The sphingosine-1-phosphate receptor modulator fingolimod (FTY720) elicits a negative chronotropic effect at treatment initiation that attenuates thereafter. The authors determined whether isoproterenol can counteract this effect. In this randomized, crossover study, 14 healthy subjects received 5 infusions of isoproterenol (titrated to increase heart rate to 100-120 bpm) or intravenous placebo. The first infusion was 2 hours before and the other 4 infusions were between 3 and 6 hours after a 5-mg oral dose of fingolimod. Telemetry and pharmacokinetic data were collected for 24 hours. During isoproterenol infusion 1 (before fingolimod administration), heart rate was increased 80% from preinfusion 68 +/- 9 bpm to a maximum 122 +/- 15 bpm. Administration of fingolimod decreased heart rate from 73 +/- 11 bpm predose to a nadir of 57 +/- 8 bpm. The subsequent isoproterenol infusion 2 in the presence of fingolimod increased mean heart rate by 85% to a maximum 105 +/- 21 bpm. A 41% higher total isoproterenol dose was needed to increase heart rate to the target range with fingolimod (97 +/- 6 mcg) compared with isoproterenol alone (69 +/- 27 mcg). Isoproterenol infusions 3 to 5 had similar effects on heart rate as infusion 2. Fingolimod had no significant influence on blood pressure responses to isoproterenol. Isoproterenol did not alter the pharmacokinetics of fingolimod. The pure beta-agonist isoproterenol can reverse the heart rate reduction that occurs transiently after initiating fingolimod treatment.

The ability of atropine to prevent and reverse the negative chronotropic effect of fingolimod in healthy subjects.

AIMS: The authors determined whether intravenous atropine can prevent or counteract the negative chronotropic effect of the immunomodulator fingolimod. METHODS: In this randomized, placebo-controlled, two-period, crossover study, 12 healthy subjects received 5 mg fingolimod orally concurrently with intravenous atropine (titrated to a heart rate of 110-120 beats min(-1)) or intravenous placebo. A second group of 12 subjects received atropine/placebo 4 h after the fingolimod dose. Continuous telemetry measurements were made for 24 h after each fingolimod dose. RESULTS: Fingolimod administration alone yielded a heart rate nadir of 51 +/- 5 beats min(-1) at a median 4 h postdose with heart rate remaining depressed at 51-64 beats min(-1) over the rest of the day. Concurrent administration of fingolimod and atropine yielded a nadir of 66 +/- 6 beats min(-1) resulting in an atropine: placebo ratio (90% confidence interval) of 1.30 (1.22, 1.36). When atropine was administered at the time of the nadir, it was able to reverse the negative chronotropic effect of fingolimod from a heart rate of 56 +/- 9 beats min(-1) (placebo) to 64 +/- 8 beats min(-1) (atropine) resulting in an atropine: placebo ratio of 1.15 (1.04, 1.26). Atropine had no influence on the pharmacokinetics of fingolimod. CONCLUSIONS: Atropine administered concurrently with fingolimod prevented the heart rate nadir that typically occurs 4 h postdose. Atropine administered at the time of the heart rate nadir was able to reverse the negative chronotropic effect of fingolimod.

Co-medication and interference testing in bioanalysis: a European Bioanalysis Forum recommendation.

Interference testing of co-medication in bioanalytical method validation has become an area of debate in view of the increased specificity offered by current state-of-the-art technology in both LC-MS/MS and ligand-binding assay platforms. In view of this, and considering the extensive experience within the European Bioanalysis Forum member companies, we evaluated the impact of co-medication on the performance of hundreds of bioanalytical methods with the aim of providing a science-based recommendation on how to evaluate and document potential interference from co-medication on the PK parameters in clinical studies in patients and volunteers.

Longitudinal noninvasive magnetic resonance imaging of brain microhemorrhages in BACE inhibitor-treated APP transgenic mice.

Currently, several immunotherapies and BACE (Beta Site APP Cleaving Enzyme) inhibitor approaches are being tested in the clinic for the treatment of Alzheimer's disease. A crucial mechanism-related safety concern is the exacerbation of microhemorrhages, which are already present in the majority of Alzheimer patients. To investigate potential safety liabilities of long-term BACE inhibitor therapy, we used aged amyloid precursor protein (APP) transgenic mice (APP23), which robustly develop cerebral amyloid angiopathy. T2*-weighted magnetic resonance imaging (MRI), a translational method applicable in preclinical and clinical studies, was used for the detection of microhemorrhages throughout the entire brain, with subsequent histological validation. Three-dimensional reconstruction based on in vivo MRI and serial Perls' stained sections demonstrated a one-to-one matching of the lesions thus allowing for their histopathological characterization. MRI detected small Perls' positive areas with a high spatial resolution. Our data demonstrate that volumetric assessment by noninvasive MRI is well suited to monitor cerebral microhemorrhages in vivo. Furthermore, 3 months treatment of aged APP23 with the potent BACE-inhibitor NB-360 did not exacerbate microhemorrhages in contrast to Aß-antibody ß1. These results substantiate the safe use of BACE inhibitors regarding microhemorrhages in long-term clinical studies for the treatment of Alzheimer's disease.

Role of epigenetic mechanisms in the development of chronic complications of diabetes.

There is growing evidence that epigenetic regulation of gene expression including post-translational histone modifications (PTHMs), DNA methylation and microRNA (miRNA)-regulation of mRNA translation could play a crucial role in the development of chronic, diabetic complications. Hyperglycemia can induce an abnormal action of PTHMs and DNA methyltransferases as well as alter the levels of numerous miRNAs in endothelial cells, vascular smooth muscle cells, cardiomyocytes, retina, and renal cells. These epigenetic abnormalities result in changes in the expression of numerous genes contributing to effects such as development of chronic inflammation, impaired clearance of reactive oxygen species (ROS), endothelial cell dysfunction and/or the accumulation of extracellular matrix in the kidney, which causing the development of retinopathy, nephropathy or cardiomyopathy. Some epigenetic modifications, for example PTHMs and DNA methylation, become irreversible over time. Therefore, these processes have gained much attention in explaining the long-lasting detrimental consequences of hyperglycaemia causing the development of chronic complications even after improved glycaemic control is achieved. Our review suggests that the treatment of chronic complications should focus on erasing metabolic memory by targeting chromatin modification enzymes and by restoring miRNA levels.

Dermal PK/PD of a lipophilic topical drug in psoriatic patients by continuous intradermal membrane-free sampling.

BACKGROUND: Methodologies for continuous sampling of lipophilic drugs and high-molecular solutes in the dermis are currently lacking. We investigated the feasibility of sampling a lipophilic topical drug and the locally released biomarker in the dermis of non-lesional and lesional skin of psoriatic patients over 25h by means of membrane-free dermal open-flow microperfusion probes (dOFM) and novel wearable multi-channel pumps. METHODS: Nine psoriatic patients received a topical p-38 inhibitor (BCT194, 0.5% cream) on a lesional and a non-lesional application site once daily for 8 days. Multiple dOFM sampling was performed for 25 h from each site on day 1 and day 8. Patients were mobile as dOFM probes were operated by a novel light-weight push-pull pump. Ultrasound was used to verify intradermal probe placement, cap-LC-MS/MS for BCT194 and ELISA for TNFα analysis. RESULTS: dOFM was well tolerated and demonstrated significant drug concentrations in lesional as well as non-lesional skin after 8 days, but did not show significant differences between tissues. On day 8, TNFα release following probe insertion was significantly reduced compared to day 1. CONCLUSIONS: Novel membrane-free probes and wearable multi-channel pumps allowed prolonged intradermal PK/PD profiling of a lipophilic topical drug in psoriatic patients. This initial study shows that dOFM overcomes limitations of microdialysis sampling methodology, and it demonstrates the potential for PK/PD studies of topical products and formulations in a clinical setting.

Oral-intravenous crossover study of fingolimod pharmacokinetics, lymphocyte responses and cardiac effects.

OBJECTIVE: The pharmacokinetics and lymphocyte responses to the immunomodulator fingolimod (FTY720) were characterized after oral and intravenous administration. METHODS: In this randomized, two-period crossover study 11 evaluable healthy subjects received single doses of fingolimod 1.25 mg orally and 1 mg intravenously infused over 2 h. The pharmacokinetics of fingolimod, blood lymphocyte counts and heart rate were characterized for 28 days after each dose. RESULTS: After oral administration, Cmax was 1.1+/-0.2 ng/ml occurring at 12 h postdose and the AUC was 201+/-31 ng.h/ml. After intravenous infusion, Cmax was 4.9+/-0.8 ng/ml, AUC was 175+/-50 ng. h/ml, clearance was 6.3+/-2.3 l/h and distribution volume was 1199+/-260 l. The oral/intravenous ratio of dose-normalized AUCs was 0.94 (95%CI: 0.78-1.12). The pharmacologically active metabolite fingolimod-phosphate was quantifiable near its peak after oral administration but not after intravenous administration. The mean lymphocyte nadir occurred on day 1 and was 35% lower after oral (0.74x10(9)/l) than after intravenous (1.15x10(9)/l) administration. Lymphocytes recovered to the normal range by day 15 for both treatments. The mean heart rate nadir occurred 3-4 h postdose and was 11% lower after oral administration (47 bpm) versus intravenous administration (53 bpm). CONCLUSIONS: Average systemic exposure to fingolimod was similar after oral and intravenous administration. However, the acute decrease in lymphocyte counts was weaker after intravenous administration, likely because of lower blood levels of the active metabolite fingolimod-phosphate compared with oral administration.