Plasma DYRK1A as a novel risk factor for Alzheimer's disease.

To determine whether apparent involvement of DYRK1A in Alzheimer's disease (AD) pathology makes it a candidate plasma biomarker for diagnosis, we developed a method to quantify plasma DYRK1A by immunoblot in transgenic mouse models having different gene dosages of Dyrk1a, and, consequently, different relative protein expression. Then, we measured plasma DYRK1A levels in 26 patients with biologically confirmed AD and 25 controls (negative amyloid imaging available on 13). DYRK1A was detected in transgenic mouse brain and plasma samples, and relative levels of DYRK1A correlated with the gene copy number. In plasma from AD patients, DYRK1A levels were significantly lower compared with controls (P<0.0001). Results were similar when we compared AD patients with the subgroup of controls confirmed by negative amyloid imaging. In a subgroup of patients with early AD (CDR=0.5), lower DYRK1A expression was confirmed. In contrast, no difference was found in levels of DYRK1B, the closest relative of DYRK1A, between AD patients and controls. Further, AD patients exhibited a positive correlation between plasma DYRK1A levels and cerebrospinal fluid tau and phosphorylated-tau proteins, but no correlation with amyloid-ß42 levels and Pittsburgh compound B cortical binding. DYRK1A levels detected in lymphoblastoid cell lines from AD patients were also lower when compared with cells from age-matched controls. These findings suggest that reduced DYRK1A expression might be a novel plasma risk factor for AD.

Classification of human chromosome 21 gene-expression variations in Down syndrome: impact on disease phenotypes.

Down syndrome caused by chromosome 21 trisomy is the most common genetic cause of mental retardation in humans. Disruption of the phenotype is thought to be the result of gene-dosage imbalance. Variations in chromosome 21 gene expression in Down syndrome were analyzed in lymphoblastoid cells derived from patients and control individuals. Of the 359 genes and predictions displayed on a specifically designed high-content chromosome 21 microarray, one-third were expressed in lymphoblastoid cells. We performed a mixed-model analysis of variance to find genes that are differentially expressed in Down syndrome independent of sex and interindividual variations. In addition, we identified genes with variations between Down syndrome and control samples that were significantly different from the gene-dosage effect (1.5). Microarray data were validated by quantitative polymerase chain reaction. We found that 29% of the expressed chromosome 21 transcripts are overexpressed in Down syndrome and correspond to either genes or open reading frames. Among these, 22% are increased proportional to the gene-dosage effect, and 7% are amplified. The other 71% of expressed sequences are either compensated (56%, with a large proportion of predicted genes and antisense transcripts) or highly variable among individuals (15%). Thus, most of the chromosome 21 transcripts are compensated for the gene-dosage effect. Overexpressed genes are likely to be involved in the Down syndrome phenotype, in contrast to the compensated genes. Highly variable genes could account for phenotypic variations observed in patients. Finally, we show that alternative transcripts belonging to the same gene are similarly regulated in Down syndrome but sense and antisense transcripts are not.

Stiripentol: efficacy and tolerability in children with epilepsy.

PURPOSE: Stiripentol (STP) is a new antiepileptic drug (AED) that inhibits cytochrome P450, resulting in increased plasma concentrations of concomitant AEDs. The efficacy and tolerability of STP as an add-on therapy in children were assessed. METHODS: Two hundred twelve patients with refractory epilepsy, aged from 1 month to 20.5 years, received STP either in a single-blind, placebo-controlled trial (108 patients) or in a further open trial (104 other patients selected by epilepsy syndrome for possible efficacy based on the results of the previous trial). RESULTS: Among the 97 patients who could be analyzed for efficacy in the placebo-controlled study, the median seizure frequency was lower at 3 months with STP than with the placebo (p<0.0001); 49% responded to the drug, including 10% who became seizure free. Patients with partial epilepsy had the highest response rate (57%). Results were confirmed in the open study where 68% of the 91 patients receiving STP responded at 3 months. These patients were mainly those with partial epilepsy (73%) who were receiving carbamazepine (CBZ) (75%) as comedication (p<0.001). Ten of the 20 children with severe myoclonic epilepsy in infancy also responded with clobazam (CLB) as comedication. Efficacy was sustained long term in 74% of the 94 patients still receiving STP at a mean 30-month follow-up. Adverse events were reported in 48% of the 212 patients, mainly anorexia and loss of weight, but these events required STP discontinuation in only nine cases. Side effects were minimized in the open trial by optimizing the dose of comedication. CONCLUSIONS: STP seems to be a promising add-on drug, particularly when combined with CBZ in patients with partial childhood epilepsy refractory to vigabatrin (VGB) and with CLB in patients with severe myoclonic epilepsy in infancy.

Saccharomyces boulardii prevents diarrhea in critically ill tube-fed patients. A multicenter, randomized, double-blind placebo-controlled trial.

OBJECTIVE: To assess the preventive effect of Saccharomyces boulardii on diarrhea in critically ill tube-fed patients and to evaluate risk factors for diarrhea. DESIGN: Prospective, multicenter, randomized, double-blind placebo-controlled study. SETTING: Eleven intensive care units in teaching and general hospitals. PATIENTS: Critically ill patients whose need for enteral nutrition was expected to exceed 6 days. INTERVENTION: S. boulardii 500 mg four times a day versus placebo. MEASUREMENTS AND RESULTS: Diarrhea was defined by a semiquantitative score based on the volume and consistency of stools. A total of 128 patients were studied, 64 in each group. Treatment with S. boulardii reduced the mean percentage of days with diarrhea per feeding days from 18.9 to 14.2% [odds ratio (OR) = 0.67, 95% confidence interval (CI) = 0.50-0.90, P = 0.0069]. In the control group, nine risk factors were significantly associated with diarrhea: nonsterile administration of nutrients in open containers, previous suspension of oral feeding, malnutrition, hypoalbuminemia, sepsis syndrome, multiple organ failure, presence of an infection site, fever or hypothermia, and use of antibiotics. Five independent factors were associated with diarrhea in a multivariate analysis: fever or hypothermia, malnutrition, hypoalbuminemia, previous suspension of oral feeding, and presence of an infection site. After adjustment for these factors, the preventive effect of S. boulardii on diarrhea was even more significant (OR = 0.61, 95% CI = 0.44-0.84, P < 0.0023). CONCLUSIONS: S. boulardii prevents diarrhea in critically ill tube-fed patients, especially in patients with risk factors for diarrhea.

Use of a microbial model for the determination of drug effects on cell metabolism and energetics: study of citrulline-malate.

Euglena gracilis can be used as a microbial model to study the effect of drugs on lactate metabolism and gluconeogenetic synthesis. The cell growth and metabolism have been characterized in a 33 mM lactate medium, non-supplemented or supplemented by dl-malate or by l-citrulline alone or by the compound formed by the stoichiometric combination of the two components: the citrulline-malate (Stimol). The malate of the complex accelerated the ammonium disappearance, while the citrulline facilitated the lactate consumption. A synergistic action of the complex, by comparison with the additive effects of the individual components, on most of the parameters studied was detected. A remarkable resistance to anoxia, and a quicker recovery under aeration of the cells supplemented with CM, were evident: after carbonation for 2 min the total nucleotides in the medium were increased by 44 per cent with an unchanged energy charge; and after a prolonged (20 min) anoxia followed by an aeration, the capacities of the cells to synthesize ATP in the presence of excesses of both ADP and phosphate were two-fold higher in Stimol treated cells than in control.

Comparison of the concentration-effect relationship of a local antiinflammatory agent and oral acetylsalicylic acid: the value of local application.

Using a pharmacological model, the comparison between acetylsalicylic acid (ASA), administered orally, and a solution combining two salicylate derivatives (ethyl 5-methoxy-salicylate and 3-phenyl-propyl-salicylate), applied locally, demonstrated the value of the local application. Indeed, the pharmacological activity was highly significant and directly related to the tissue concentration of salicyl ions, which was higher after local application of the solution than after oral administration of ASA. The local solution also resulted in a lower plasma concentration of salicylate ions, allowing high plasma salicylate concentrations to be avoided.

Stiripentol kinetics in epilepsy: nonlinearity and interactions.

Stiripentol kinetics during oral therapy were assessed in six patients with epilepsy who were receiving other antiepileptic drugs. Steady-state levels at 600, 1200, and 2400 mg/day increased in a nonlinear fashion, indicating Michaelis-Menten kinetics. Oral clearance of stiripentol at 600 mg/day was 41.5 +/- 23.4 l/day/kg (mean +/- SD), greater than that at 1200 mg/day (20.3 +/- 8.8 l/day/kg; P less than 0.05) or 2400 mg/day (8.5 +/- 3.8 l/day/kg; P less than 0.01). The apparent in vivo Michaelis-Menten parameters were determined from three mean steady-state concentrations. The average velocity of conversion of stiripentol to its metabolites (Vm), Michaelis constant (Km), and the ratio Vm/Km were 49.3 +/- 13.1 mg/day/kg, 1.35 +/- 1.08 mg/l, and 50.2 +/- 27.5 l/day/kg. Stiripentol reduced the elimination clearances of concomitant antiepileptic drugs. Phenytoin clearance was reduced in all five subjects who received this drug, from a mean control of 29.5 +/- 13.4 l/day to 18.5 +/- 4.6 l/day at a stiripentol dose of 1200 mg/day (P = 0.05) and to 6.48 +/- 2.59 l/day at 2400 mg/day (P less than 0.01). Stiripentol reduced the clearance of carbamazepine in one subject from a control value of 209 l/day to 128 l/day (1200 mg/day) and 61 l/day (2400 mg/day). Stiripentol reduced phenobarbital clearance in two subjects from 3.8 and 5.1 l/day to 2.3 and 3.4 l/day (2400 mg/day). The Michaelis-Menten kinetics of stiripentol, as well as its interactions with other antiepileptic drugs, have important implications in the designing of controlled clinical trials.

Michaelis-Menten kinetics of stiripentol in normal humans.

Michaelis-Menten kinetic parameters for stiripentol, and anticonvulsant, were assessed in six normal volunteers. Stiripentol was administered orally three times a day in dosage increments of 600, 1,200, and 1,800 mg/day for consecutive periods of 3, 4, and 7 days, respectively. Stiripentol steady-state levels at the three dosing rates increased more than proportionally with dose. The mean +/- SD oral clearance of stiripentol at 600 mg/day (1,090 +/- 624 L/day) was significantly greater (p less than 0.01) than at 1,200 (506 +/- 219 L/day) or 1,800 (405 +/- 151 L/day) mg/day. Average steady-state concentrations predicted from individually determined Vm and Km parameters were in good agreement with experimentally observed levels, indicating that the kinetics of stiripentol are of the Michaelis-Menten type. The mean Vm, Km, and Vm/Km ratio were 2,299 +/- 490 mg/day, 2.20 +/- 1.28 mg/L, and 1,241 +/- 837 L/day, respectively. Neuropsychological tests carried out before and after 14 days of stiripentol treatment showed a significant decline in verbal learning ability (p = 0.038) and a significant improvement in a test of memory and attention (p less than 0.01).

Pharmacokinetics of stiripentol in normal man: evidence of nonlinearity.

The pharmacokinetics and metabolism of stiripentol, a new antiepileptic drug, were investigated in normal male subjects after single-dose and multiple-dose administration. Each of six subjects received single doses of 300, 600, and 1200 mg of stiripentol in powder form and another 600 mg in solution. In the multiple-dose study, each of six subjects received a 300-mg dose on day 1 and multiple doses (1200 mg/day) from day 2 to day 8. Five of these six subjects participated also in the single-dose study. Stiripentol and several of its metabolites, namely, stiripentol conjugate, DiOH, P-OH, and M-OH, were analyzed in plasma and urine. After single doses, the elimination curve of stiripentol appeared multiphasic. The oral clearance was 1.3 to 1.8 liter/hr/kg. The average mean residence time was 4 hours. There were no statistically significant differences in clearance or mean residence time among the three doses. However, dose dependence was found in all the four pathways when formation clearances were compared. Only trace amounts of the drug were excreted unchanged in urine. The active metabolite, P-OH, was not detectable in plasma. Stiripentol was very highly bound to plasma proteins in plasma from dosed subjects as well as spiked human plasma (free fraction of 1 per cent). In the multiple-dose study, there was a decrease (nearly eightfold) in oral clearance of stiripentol between day 1 and day 8. The fractions of dose metabolized through conjugation and methylenedioxy ring opening increased 183 and 49 per cent, respectively, but the formation clearances for all the pathways were decreased. These findings suggest that the steady-state plasma level/dose ratio of stiripentol will increase with the daily dose.