Sputum RNA signature in allergic asthmatics following allergen bronchoprovocation test.

BACKGROUND: Inhaled allergen challenge is a validated disease model of allergic asthma offering useful pharmacodynamic assessment of pharmacotherapeutic effects in a limited number of subjects. OBJECTIVES: To evaluate whether an RNA signature can be identified from induced sputum following an inhaled allergen challenge, whether a RNA signature could be modulated by limited doses of inhaled fluticasone, and whether these gene expression profiles would correlate with the clinical endpoints measured in this study. METHODS: Thirteen non-smoking, allergic subjects with mild-to-moderate asthma participated in a randomised, placebo-controlled, 2-period cross-over study following a single-blind placebo run-in period. Each period consisted of three consecutive days, separated by a wash-out period of at least 3 weeks. Subjects randomly received inhaled fluticasone ((FP) MDI; 500 mcg BID×5 doses in total) or placebo. On day 2, house dust mite extract was inhaled and airway response was measured by FEV1 at predefined time points until 7 h post-allergen. Sputum was induced by NaCl 4.5%, processed and analysed at 24 h pre-allergen and 7 and 24 h post-allergen. RNA was isolated from eligible sputum cell pellets (<80% squamous of 500 cells), amplified according to NuGEN technology, and profiled on Affymetrix arrays. Gene expression changes from baseline and fluticasone treatment effects were evaluated using a mixed effects ANCOVA model at 7 and at 24 h post-allergen challenge. RESULTS: Inhaled allergen-induced statistically significant gene expression changes in sputum, which were effectively blunted by fluticasone (adjusted p<0.025). Forty-seven RNA signatures were selected from these responses for correlation analyses and further validation. This included Th2 mRNA levels for cytokines, chemokines, high-affinity IgE receptor FCER1A, histamine receptor HRH4, and enzymes and receptors in the arachidonic pathway. Individual messengers from the 47 RNA signatures correlated significantly with lung function and sputum eosinophil counts. CONCLUSION: Our RNA extraction and profiling protocols allowed reproducible assessments of inflammatory signatures in sputum including quantification of drug effects on this response in allergic asthmatics. This approach offers novel possibilities for the development of pharmacodynamic (PD) biomarkers in asthma.

The effects of a novel histamine-3 receptor inverse agonist on essential tremor in comparison to stable levels of alcohol.

Essential tremor (ET) is a common movement disorder. Animal studies show that histaminergic modulation may affect the pathological processes involved in the generation of ET. Histamine-3 receptor inverse agonists (H3RIA) have demonstrated attenuating effects on ET in the harmaline rat model. In this double-blind, three-way cross-over, single-dose, double-dummy study the effects of 25 mg of a novel H3RIA (MK-0249) and a stable alcohol level (0.6 g L(-1)) were compared with placebo, in 18 patients with ET. Tremor was evaluated using laboratory tremorography, portable tremorography and a clinical rating scale. The Leeds Sleep Evaluation Questionnaire (LSEQ) and a choice reaction time (CRT) test were performed to evaluate potential effects on sleep and attention, respectively. A steady state of alcohol significantly diminished tremor as assessed by laboratory tremorography, portable tremorography and clinical ratings compared with placebo. A high single MK-0249 dose was not effective in reducing tremor, but caused significant effects on the LSEQ and the CRT test. These results suggest that treatment with a single dose of MK-0249 does not improve tremor in alcohol-responsive patients with ET, whereas stable levels of alcohol as a positive control reproduced the commonly reported tremor-diminishing effects of alcohol.

The effects of TPA023, a GABAAα2,3 subtype-selective partial agonist, on essential tremor in comparison to alcohol.

Essential tremor (ET) is a relatively frequent neurological disorder that responds in some patients to gamma-aminobutyric acid A (GABA(A)) agonists such as the benzodiazepines. Partial subtype-selective GABA(A) agonists may have an improved side effect profile compared to non-selective GABA(A) agonists. However, it is unknown which GABA(A) subtypes are involved in the therapeutic effects of benzodiazepines in ET. The effects of 2 mg TPA023, a GABA(A) α2,3 subtype-selective partial agonist, on ET were compared to the effects of a stable alcohol level (0.6 g/L) and placebo in nine patients with ET. Tremor evaluation included laboratory accelerometry and a performance-based scale. Additional measurements were performed to evaluate other effects on the central nervous system (CNS). Alcohol significantly diminished tremor symptoms in the postural and kinetic condition, as assessed by laboratory accelerometry, but the performance-based rating scale was unaffected. Tremor was also reduced after TPA023 treatment in the kinetic condition, albeit not significantly. Additionally, TPA023 decreased saccadic peak velocity, while alcohol decreased subjective feelings of alertness. This study showed that alcohol reduced maximum tremor power, as assessed by laboratory accelerometry, unlike TPA023, which decreased tremor symptoms to some extent but not significantly. This study showed that treatment with an α2,3 subunit-selective GABA(A) partial agonist was less effective than a stable level of alcohol in reducing ET symptoms. These results provide no support for a therapeutic role of TPA023 in the suppression of ET symptoms.

Effect of the cathepsin K inhibitor odanacatib on bone resorption biomarkers in healthy postmenopausal women: two double-blind, randomized, placebo-controlled phase I studies.

Inhibition of cathepsin K (CatK) is a potential new treatment for osteoporosis. In two double-blind, randomized, placebo-controlled phase I studies, postmenopausal female subjects received odanacatib (ODN), an orally active, potent, and selective CatK inhibitor, once weekly for 3 weeks or once daily for 21 days. Bone turnover biomarkers, safety monitoring, and plasma ODN concentrations were assessed. These studies showed ODN to be well tolerated. Pharmacokinetic (PK) analysis revealed a long half-life (t(1/2); 66-93 h) consistent with once-weekly dosing. Pronounced reductions in C-terminal telopeptide of type I collagen (approximately 62%) and N-terminal telopeptide of type I collagen normalized to creatinine (NTx/Cr) (approximately 62%) at trough (C(168 h)) were seen following weekly administration. Robust reductions in CTx (up to 81%) and NTx/Cr (up to 81%) were seen following daily administration. ODN exhibits robust and sustained suppression of bone resorption biomarkers (CTx and NTx/Cr) at weekly doses > or = 25 mg and daily doses > or = 2.5 mg.

Pharmacodynamic and pharmacokinetic effects of MK-0343, a GABA(A) alpha2,3 subtype selective agonist, compared to lorazepam and placebo in healthy male volunteers.

The use of non-selective gamma-aminobutyric acid (GABA) enhancers, such as benzodiazepines in the treatment of anxiety disorders is still widespread but hampered by unfavourable side effects. some of these may be associated with binding properties to certain subtypes of the GABA(A) receptor that are unnecessary for therapeutic effects. MK-0343 was designed to be a less sedating anxiolytic, based on reduced efficacy at the alpha1 subtype and significant efficacy at alpha2 and alpha3 subtypes of the GABA(A) receptor. This paper is a double-blind, four-way cross-over (n = 12) study to investigate the effects of MK-0343 (0.25 and 0.75 mg) in comparison to placebo and an anxiolytic dose (2 mg) of the non-selective agonist lorazepam. Effects were measured by eye movements, body sway, Visual Analogue scales (VAS) and memory tests. Lorazepam impaired saccadic peak velocity (SPV), VAs alertness scores, postural stability and memory and increased saccadic latency and inaccuracy. MK-0343 0.75 mg was equipotent with lorazepam as indicated by SPV (-42.4 deg/s), saccadic latency (0.02 s) and VAS alertness scores (1.50 ln mm), while effects on memory and postural stability were smaller. MK-0343 0.25 mg only affected postural stability to a similar extent as MK-0343 0.75 mg. The effect profile of MK-0343 0.75 mg is different from the full agonist lorazepam, which could reflect the selective actions of this compound. Although less effect on VAS alertness was expected, diminished effects on memory and postural stability were present. Clinical studies in anxiety patients should show whether this dose of MK-0343 is therapeutically effective with a different side-effect profile.

In vitro availability of calcium, iron, and zinc from first-age infant formulae and human milk.

BACKGROUND: Variation in the bioavailability of calcium (Ca), iron (Fe), and zinc (Zn) occurs because of interactions of food components in the gastrointestinal microenvironment. Bioavailability is preferably determined by in vivo tests, but these are expensive, labor-intensive, time consuming, and often unethical. As an alternative, in vitro methods can be used to predict bioavailability of nutrients from foodstuffs. METHODS: A continuous-flow dialysis model with preliminary intraluminal digestive phase, adapted to the gastrointestinal conditions of infants younger than 6 months, was used. Human milk was the reference standard. Ca, Fe, and Zn content of samples and dialysates after digestion were analyzed by atomic absorption spectrometry. RESULTS: Ca availability is similar in human milk (13.1%+/-0.8%), whey (13.3%+/-1.2%), and soy-based formulae (13.0%+/-1.2%; P > 0.05), and higher in casein-predominant formula (21.2%+/-0.6%; P < 0.05). Availability of Fe is highest in human milk (8.12%+/-0.27%: P < 0.05). Fe availability in whey (1.28%+/-0.28%) and soy formulae (1.48%+/-0.28%) is similar (P > 0.05), but availability is lower in casein-predominant formula (0.48%+/-0.22%; P < 0.05). Zn availability is also highest in human milk (13.1%+/-0.7%; P < 0.05). However, Zn availability is similar in whey (6.7%+/-0.6%) and casein formulae (8.5%+/-1.6%; P > 0.05), but lower in soy formula (2.3%+/-0.4%; P < 0.05). CONCLUSIONS: Our observations are in agreement with previous data from in vivo studies in term infants. This in vitro procedure is an inexpensive, simple, rapid, and reliable method that predicts the bioavailability of Ca, Fe, and Zn in foods.

Indication of silicon essentiality in humans: serum concentrations in Belgian children and adults, including pregnant women.

Serum silicon concentrations were determined in Belgian healthy children and adults, including pregnant women, by electrothermal atomic absorption spectrometry. Serum levels appeared to be significantly higher in healthy children (1-18 yr) than in healthy adults (19-60 yr). Especially, levels in infants (< 1 yr) were higher than in any other group. Compared to age-matched nonpregnants, the serum silicon content was very low in pregnant women. In addition to the fact that this study for the first time provides serum silicon values in adults and children in Belgium, the most important observation is that these serum profiles might be an indication of silicon essentiality in man.

Thickening infant formula with digestible and indigestible carbohydrate: availability of calcium, iron, and zinc in vitro.

BACKGROUND: Regurgitation is common in infants and is usually due to gastroesophageal reflux. Often parental reassurance and dietary management by feeding thickened formulas are the only therapeutic steps necessary. Adding fibers may interfere with the absorption of micronutrients. METHODS: A continuous-flow dialysis in vitro method with a preliminary intraluminal digestive phase, modified to simulate the conditions of infants less than 6 months of age and children from 6 months of age on, was used to study the availability of calcium, iron, and zinc from thickened and nonthickened first-and second-age infant formulas. Pooled mature human milk was used as the reference standard. The elemental content of the samples and dialysate fractions of calcium, iron, and zinc after digestion was determined by atomic absorption spectrometry. RESULTS: In human milk, calcium, iron, and zinc were highly available for absorptive processes. Availability of calcium, iron, and zinc from nonthickened first- and second-age infant formulas tends to be significantly better than in the corresponding products thickened with locust bean gum. Thickening infant formulas with pregelatinized rice starch, however, does not affect the availability of calcium, iron, and zinc. CONCLUSIONS: It appears that human milk provides optimal conditions for the availability of calcium, iron, and zinc. Availability of calcium, iron, and zinc seems to lower when infant formulas are thickened with indigestible carbohydrates, whereas it does not by adding digestible carbohydrates.

Adherence to guidelines for antiretroviral therapy and for preventing opportunistic infections in HIV-infected adults and adolescents in Ryan White-funded facilities in the United States.

To determine adherence by health care providers to guidelines for antiretroviral therapy and for prevention of opportunistic infections (OIs) in adults with HIV infection in federally funded facilities in the United States, we reviewed records of HIV-infected adults (>13 years) in 11 Ryan White Title III facilities in four states for information on eight standard-of-care recommendations during November 1996 through September 1997. Eligibility required a visit to the facility within 6 months before record abstraction and a lowest CD4+ lymphocyte count <500 cells/microl. Reviews were completed for 148 patients in Maryland, 355 in New York, 370 in Georgia, and 538 in Illinois. Adherence to prevention measures by health care providers was >85% for HIV plasma RNA testing, prescription of antiretroviral therapy, Pneumocystis carinii pneumonia (PCP) prophylaxis, anti-Toxoplasma antibody testing, and obtaining Papanicolaou (Pap) smears but lower (69%-80%) for Mycobacterium avium complex (MAC) prophylaxis, tuberculin skin testing (TST), and pneumococcal vaccination. Adherence was similar by patient age, gender, racial/ethnic group, urban versus rural, and hospital versus clinic setting but was generally lower for injecting drug users (IDUs) than for patients with other HIV exposures (p < .05 by multivariate analysis for TST, anti-Toxoplasma antibody testing, Pap smear, and measurement of HIV plasma RNA). Adherence by health care providers to guidelines for preventing OIs in these federally funded facilities is generally high but could be improved for some prevention measures, for instance, MAC prophylaxis, TST, and pneumococcal vaccination, especially for IDUs.

Diffusibility of selenate, selenite, seleno-methionine, and seleno-cystine during simulated gastrointestinal digestion.

The present study was undertaken to evaluate the in vitro availability of chemically varying forms of selenium (Se), supplemented in cow's milk. Two inorganic (selenite and selenate) and two organic (seleno-methionine [Se-Met] and seleno-cystine [Se-Cys]) Se sources were evaluated. The in vitro availability was estimated by the diffusibility of Se during simulated gastrointestinal digestion. First, the diffusibility was compared after adding a constant amount of Se as either selenate, selenite, seleno-methionine, or Se-Cys in milk samples. Se-Met and selenate were found to be significantly more diffusible than seleno-cystine and selenite under the simulated gastrointestinal conditions. The tendency for superior in vitro availability of selenate and Se-Met compared to selenite and Se-Cys was confirmed for a supplementation range of 5-40 ng/g of Se. This study suggests that the high diffusibility of selenate and Se-Met in a simulated gastrointestinal environment may contribute to their high absorption in vivo.

The influence of different food components on the in vitro availability of iron, zinc and calcium from a composed meal.

The availability of iron, zinc and calcium from a composed meal was studied by an in vitro method using equilibrium dialysis after simulated gastric digestion. Four different concentrations of four influencing factors (coffee, vitamin C, wheat bran and pectin) were added to the mixed meal and their effect on the relative index of availability was studied after elemental analysis by atomic absorption spectrometry. Apart from ascorbic acid, all other factors had a negative effect on availability of minerals and trace elements. Most pronounced effect, and for all three elements, was observed for the addition of wheat bran. Zinc was the trace element, which was most sensitive to increased spiking of food constituents.