High Resolution Discovery Proteomics Reveals Candidate Disease Progression Markers of Alzheimer's Disease in Human Cerebrospinal Fluid.

Disease modifying treatments for Alzheimer's disease (AD) constitute a major goal in medicine. Current trends suggest that biomarkers reflective of AD neuropathology and modifiable by treatment would provide supportive evidence for disease modification. Nevertheless, a lack of quantitative tools to assess disease modifying treatment effects remains a major hurdle. Cerebrospinal fluid (CSF) biochemical markers such as total tau, p-tau and Ab42 are well established markers of AD; however, global quantitative biochemical changes in CSF in AD disease progression remain largely uncharacterized. Here we applied a high resolution open discovery platform, dMS, to profile a cross-sectional cohort of lumbar CSF from post-mortem diagnosed AD patients versus those from non-AD/non-demented (control) patients. Multiple markers were identified to be statistically significant in the cohort tested. We selected two markers SME-1 (p<0.0001) and SME-2 (p = 0.0004) for evaluation in a second independent longitudinal cohort of human CSF from post-mortem diagnosed AD patients and age-matched and case-matched control patients. In cohort-2, SME-1, identified as neuronal secretory protein VGF, and SME-2, identified as neuronal pentraxin receptor-1 (NPTXR), in AD were 21% (p = 0.039) and 17% (p = 0.026) lower, at baseline, respectively, than in controls. Linear mixed model analysis in the longitudinal cohort estimate a decrease in the levels of VGF and NPTXR at the rate of 10.9% and 6.9% per year in the AD patients, whereas both markers increased in controls. Because these markers are detected by mass spectrometry without the need for antibody reagents, targeted MS based assays provide a clear translation path for evaluating selected AD disease-progression markers with high analytical precision in the clinic.

Cerebrospinal fluid biomarkers distinguish postmortem-confirmed Alzheimer's disease from other dementias and healthy controls in the OPTIMA cohort.

Cerebrospinal fluid (CSF) amyloid-ß (Aß) and tau have been studied as markers of Alzheimer's disease (AD). Combined Aß42 and t-tau distinguishes AD from healthy controls with a sensitivity and specificity (sens/spec) near 89% across studies. This study examined these markers in the homogeneous OPTIMA cohort, using extensive longitudinal follow up and postmortem evaluation to confirm clinicopathological status. Baseline CSF was analyzed from 227 participants with AD (97% autopsy-confirmed), mild cognitive impairment (MCI; 73% confirmed), other dementia syndrome (ODS; 100% confirmed), and controls (CTL; 27% confirmed, follow up approximately 9-13 years). Biomarker concentrations were analyzed using validated ELISAs. AD patients had lower CSF Aß42 and higher t-tau, p-tau, t-tau/Aß42, and t-tau/Aß40 compared to CTLs, with MCI intermediate. CTL and MCI participants who progressed to AD demonstrated more AD-like profiles. Aß40, sAßPPα, and sAßPPß were lower in AD compared to CTL. High-level discriminators of AD from CTL were t-tau/Aß40 (AUROC 0.986, sens/spec of 92%/94%), p-tau/Aß42 (AUROC 0.972, sens/spec of 94%/90%), and Aß42 (AUROC 0.941, sens/spec of 88%). For discriminating AD from ODS, p-tau/Aß42 demonstrated sens/spec of 88%/100% (95%/86% at the AD versus CTL cutoff) and Aß42 demonstrated sens/spec of 84%/100% (88%/100% at the AD versus CTL cutoff). In a well-characterized, homogeneous population, a single cutoff for baseline CSF Aß and tau markers can distinguish AD with a high level of sens/spec compared to other studies. It may be important to characterize sources of demographic and biological variability to support the effective use of CSF diagnostic assays in the broader AD population.

The effects of laropiprant, a selective prostaglandin D2 receptor 1 antagonist, on the antiplatelet activity of clopidogrel or aspirin.

Laropiprant (LRPT) is being developed in combination with Merck's extended-release niacin (ERN) formulation for the treatment of dyslipidemia. LRPT, an antagonist of the prostaglandin PGD2 receptor DP1, reduces flushing symptoms associated with ERN. LRPT also has affinity for the thromboxane A2 receptor TP (approximately 190-fold less potent at TP compared with DP1). Aspirin and clopidogrel are two frequently used anti-clotting agents with different mechanisms of action. Since LRPT may potentially be co-administered with either one of these agents, these studies were conducted to assess the effects of steady-state LRPT on the antiplatelet activity of steady-state clopidogrel or aspirin. Bleeding time at 24 h post-dose (trough) was pre-specified as the primary pharmacodynamic endpoint in both studies. Two separate, double-blind, randomized, placebo-controlled, crossover studies evaluated the effects of multiple-dose LRPT on the pharmacodynamics of multiple-dose clopidogrel or aspirin. Healthy subjects were randomized to once-daily oral doses of LRPT 40 mg or placebo to LRTP co-administered with clopidogrel 75 mg or aspirin 81 mg for 7 days with at least a 21-day washout between treatments. In both studies, bleeding time and platelet aggregation were assessed 4 and 24 hours post-dose on Day 7. Comparability was declared if the 90% confidence interval for the estimated geometric mean ratio ([LRPT+clopidogrel]/clopidogrel alone or [LRPT+aspirin]/aspirin alone) for bleeding time at 24 hours post-dose on Day 7 was contained within (0.66, 1.50). Concomitant daily administration of LRPT 40 mg with clopidogrel 75 mg or aspirin 81 mg resulted in an approximate 4-5% increase in bleeding time at 24 hours after the last dose vs. bleeding time after treatment with clopidogrel or aspirin alone, demonstrating that the treatments had comparable effects on bleeding time. Percent inhibition of platelet aggregation was not significantly different between LRPT co-administered with clopidogrel or aspirin vs. clopidogrel or aspirin alone at 24 hours post-dose at steady state. At 4 hours after the last dose, co-administration of LRPT 40 mg resulted in 3% and 41% increase in bleeding time vs. bleeding time after treatment with aspirin or clopidogrel alone, respectively. Co-administration of LPRT with clopidogrel or aspirin was generally well tolerated in healthy subjects. Co-administration of multiple doses of LRPT 40 mg and clopidogrel 75 mg or aspirin 81 mg had no clinically important effects on bleeding time or platelet aggregation.

Non-linear relationships of cerebrospinal fluid biomarker levels with cognitive function: an observational study.

INTRODUCTION: Levels of cerebrospinal fluid (CSF) ß-amyloid (Aß) and Tau proteins change in Alzheimer's disease (AD). We tested if the relationships of these biomarkers with cognitive impairment are linear or non-linear. METHODS: We assessed cognitive function and assayed CSF Aß and Tau biomarkers in 95 non-demented volunteers and 97 AD patients. We then tested non-linearities in their inter-relations. RESULTS: CSF biomarkers related to cognitive function in the non-demented range of cognition, but these relations were weak or absent in the patient range; Aß1-40's relationship was biphasic. CONCLUSIONS: Major biomarker changes precede clinical AD and index cognitive impairment in AD poorly, if at all.

Effects of extended release niacin/laropiprant, laropiprant, extended release niacin and placebo on platelet aggregation and bleeding time in healthy subjects.

Laropiprant (LRPT) has been shown to reduce flushing symptoms induced by niacin and has been combined with niacin for treatment of dyslipidemia. LRPT, a potent PGD(2) receptor (DP1) antagonist that also has modest activity at the thromboxane receptor (TP), may have the potential to alter platelet function either by enhancing platelet reactivity through DP1 antagonism or by inhibiting platelet aggregation through TP antagonism. Studies of platelet aggregation ex vivo and bleeding time have shown that LRPT, at therapeutic doses, does not produce clinically meaningful alterations in platelet function. The present study was conducted to assess platelet reactivity to LRPT using methods that increase the sensitivity to detect changes in platelet responsiveness to collagen and ADP. The responsiveness of platelets was quantified by determining the EC(50) of collagen to induce platelet aggregation ex vivo. At 24 hours post-dose on Day 7, the responsiveness of platelets to collagen-induced aggregation was similar following daily treatment with extended-release niacin (ERN) 2 g/LRPT 40 mg or ERN 2 g. At 2 hours post-dose on Day 7, the EC(50) for collagen-induced platelet aggregation was approximately two-fold higher in the presence of LRPT, consistent with a small, transient inhibition of platelet responsiveness to collagen. There was no clinical difference between treatments for bleeding time, suggesting that this small effect on collagen EC(50) does not result in a clinically meaningful alteration of platelet function in vivo. The results of this highly sensitive method demonstrate that LRPT does not enhance platelet reactivity when given alone or with ERN.

Comparative inhibitory activity of etoricoxib, celecoxib, and diclofenac on COX-2 versus COX-1 in healthy subjects.

We determined cyclo-oxygenase-1 and cyclo-oxygenase-2 inhibition in healthy middle-aged subjects (41-65 years) randomly assigned to four 7-day treatment sequences of etoricoxib 90 mg every day, celecoxib 200 mg twice a day, diclofenac 75 mg twice a day, or placebo in a double-blind, randomized, 4-period crossover study. Maximum inhibition of thromboxane B(2) (cyclo-oxygenase-1 activity) in clotting whole blood on day 7 (0-24 hours postdose) was the primary endpoint. Inhibition of lipopolysaccharide-induced prostaglandin E(2) in whole blood (cyclo-oxygenase-2 activity) was assessed on day 7 (0-24 hours postdose) as a secondary endpoint. Diclofenac had significantly greater maximum inhibition of thromboxane B(2) versus each comparator (P < .001); placebo 2.4% (95% confidence interval: -8.7% to 12.3%), diclofenac 92.2% (91.4% to 92.9%), etoricoxib 15.5% (6.6% to 23.5%), and celecoxib 20.2% (11.5% to 28.1%). Prostaglandin E(2) synthesis was inhibited with a rank order of potency of diclofenac > etoricoxib > celecoxib. In summary, at doses commonly used in rheumatoid arthritis, diclofenac significantly inhibits both cyclo-oxygenase-1 and cyclo-oxygenase-2, whereas etoricoxib and celecoxib significantly inhibit cyclo-oxygenase-2 and do not substantially inhibit cyclo-oxygenase-1.

Characterization of etoricoxib, a novel, selective COX-2 inhibitor.

Etoricoxib is a potent selective COX-2 inhibitor in man. Ex vivo whole-blood assays assessed COX-2 inhibition after oral administration of etoricoxib in single (5-500 mg) and multiple (25-150 mg) once-daily doses to healthy human subjects. A separate study examined ex vivo gastric mucosal PGE2 synthesis after etoricoxib (120 mg qd), naproxen (500 mg bid), or placebo for 5 days. The effect of etoricoxib 120 mg qd on the COX-1-mediated antiplatelet effects of low-dose aspirin (ASA) was also assessed. The mean (time)-weighted average inhibition (WAI) of lipopolysaccharide (LPS)-stimulated PGE2 (COX-2 assay) vcrsus placebo was dose related after single (range: 3.1%-99.1%) and multiple doses (range: 52.5%-96.7%). PGE2 remained significantly inhibited 24 hours postdose at steady state. Inhibition of LPS-stimulated PGE2 showed a strong relationship with etoricoxib plasma concentrations; ex vivo, IC50 was almost identical to in vitro. Multiple dosing of etoricoxib (up to 150 mg qd) showed no important effects on serum TXB2, bleeding time, or platelet aggregation (COX-1-mediated effects). The nonselective nonsteroidal anti-inflammatory (NSAID) naproxen significantly inhibited (approximately 78%) ex vivo prostaglandin synthesis in gastric mucosa; etoricoxib had no effect. Etoricoxib did not interfere with the antiplatelet effects of low-dose ASA, as assessed by serum TXB2 and platelet aggregation. Etoricoxib was generally well tolerated, even at doses above the clinical dose range. Based on these results, etoricoxib is a potent selective inhibitor of COX-2 after single and multiple dosing regimens and does not inhibit prostaglandin synthesis in the gastric mucosa, even at doses above the clinical dose range of 60 to 120 mg.