Recent improvements in the development of A<Subscript>2B</Subscript> adenosine receptor agonists

Adenosine is known to exert most of its physiological functions by acting as local modulator at four receptor subtypes named A₁, A_2A, A_2B and A₃ (ARs). Principally as a result of the difficulty in identifying potent and selective agonists, the A_2B AR is the least extensively characterised of the adenosine receptors family. Despite these limitations, growing understanding of the physiological meaning of this target indicates promising therapeutic perspectives for specific ligands. As A_2B AR signalling seems to be associated with pre/postconditioning cardioprotective and anti-inflammatory mechanisms, selective agonists may represent a new therapeutic group for patients suffering from coronary artery disease. Herein we present an overview of the recent advancements in identifying potent and selective A_2B AR agonists reported in scientific and patent literature. These compounds can be classified into adenosine-like and nonadenosine ligands. Nucleoside-based agonists are the result of modifying adenosine by substitution at the N ⁶-, C²-positions of the purine heterocycle and/or at the 5′-position of the ribose moiety or combinations of these substitutions. Compounds 1-deoxy-1-{6-[N′-(furan-2-carbonyl)-hydrazino]-9H-purin-9-yl}-N-ethyl-β-D-ribofuranuronamide (19, hA₁ K _i = 1050 nM, hA_2A K _i = 1550 nM, hA_2B EC₅₀ = 82 nM, hA₃ K _i > 5 μM) and its 2-chloro analogue 23 (hA₁ K _i = 3500 nM, hA_2A K _i = 4950 nM, hA_2B EC₅₀ = 210 nM, hA₃ K _i > 5 μM) were confirmed to be potent and selective full agonists in a cyclic adenosine monophosphate (cAMP) functional assay in Chinese hamster ovary (CHO) cells expressing hA_2B AR. Nonribose ligands are represented by conveniently substituted dicarbonitrilepyridines, among which 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide (BAY-60–6583, hA₁, hA_2A, hA₃ EC₅₀ > 10 μM; hA_2B EC₅₀ = 3 nM) is currently under preclinical-phase investigation for treating coronary artery disorders and atherosclerosis.     

Time-dependent evolution of tissue markers by MALDI-MS imaging

We have used MALDI-MS imaging (MALDI-MSI) to monitor the time dependent appearance and loss of signals when tissue slices are brought rapidly to room temperature for short to medium periods of time. Sections from mouse brain were cut in a cryostat microtome, placed on a MALDI target and allowed to warm to room temperature for 30 s to 3 h. Sections were then refrozen, fixed by ethanol treatment and analysed by MALDI-MSI. The intensity of a range of markers were seen to vary across the time course, both increasing and decreasing, with the intensity of some markers changing significantly within 30 s and markers also showed tissue location specific evolution. The markers resulting from this autolysis were compared directly to those that evolved in a comparable 16 h on-tissue trypsin digest, and the markers that evolved in the two studies were seen to be substantially different. These changes offer an important additional level of location-dependent information for mapping changes and seeking disease-dependent biomarkers in the tissue. They also indicate that considerable care is required to allow comparison of biomarkers between MALDI-MSI experiments and also has implications for the standard practice of thaw-mounting multiple tissue sections onto MALDI-MS targets.     

Quantitative trait locus analysis of circulating adhesion molecules in hyperlipidemic apolipoprotein E-deficient mice

Circulating soluble adhesion molecules have been suggested as useful markers to predict several clinical conditions such as atherosclerosis, type 2 diabetes, obesity, and hypertension. To determine genetic factors influencing plasma levels of soluble vascular cell adhesion molecule-1 (VCAM-1) and P-selectin, quantitative trait locus (QTL) analysis was performed on an intercross between C57BL/6J (B6) and C3H/HeJ (C3H) mouse strains deficient in apolipoprotein E-deficient (apoE^−/−). Female F₂ mice were fed a western diet for 12 weeks. One significant QTL, named sVcam1 (71 cM, LOD 3.9), on chromosome 9 and three suggestive QTLs on chromosomes 5, 13 and 15 were identified to affect soluble VCAM-1 levels. Soluble P-selectin levels were controlled by one significant QTL, named sSelp1 (8.5 cM, LOD 3.4), on chromosome 16 and two suggestive QTLs on chromosomes 10 and 13. Both adhesion molecules showed significant or an apparent trend of correlations with body weight, total cholesterol, and LDL/VLDL cholesterol levels in the F₂ population. These results indicate that plasma VCAM-1 and P-selectin levels are complex traits regulated by multiple genes, and this regulation is conferred, at least partially, by acting on body weight and lipid metabolism in hyperlipidemic apoE^−/− mice. Zuobiao Yuan and Zhiguang Su contributed equally.     

Ethnicity and weight status affect the accuracy of proxy indices of insulin sensitivity

This study tested the hypotheses that correlations between direct measures of insulin sensitivity and proxy indices of insulin sensitivity derived from fasting values, (i) would not be affected by ethnicity, and (ii) would be stronger in overweight vs. weight-reduced states. We further hypothesized that associations between proxy indices and fat distribution would be similar to those between directly measured insulin sensitivity and fat distribution. Testing was performed in weight-stable conditions in 59 African-American (AA) and 62 white-American (WA) overweight, premenopausal women before and after a weight loss intervention. Subjects were retested 1 year following weight loss. Proxy indices were correlated against the insulin sensitivity index S(I) determined via minimal modeling. Fat distribution was assessed using computed tomography. Correlations between Si and proxy indices were consistently stronger among overweight women (r = 0.44-0.52) vs. weight-reduced women (r = 0.18-0.32), and among AA (r = 0.49-0.56, baseline; 0.24-0.36, weight-reduced) vs. WA (r = 0.38-0.46, baseline; 0.19-0.31, weight-reduced). Among subjects who regained >3 kg after 1 year, correlations between S(I) and proxy indices were similar to those observed at baseline, whereas correlations were weak among women who maintained their reduced body weight. S(I) and all proxy indices were similarly correlated with intra-abdominal adipose tissue (IAAT) at baseline, but not after weight loss. In conclusion, correlations between S(I) and proxy indices were affected by both ethnicity and weight status. If proxy indices are used in multiethnic populations, or in populations including both lean and overweight/obese subjects, data should be interpreted with caution.     

Parental recognition of overweight in school-age children

Objective: Examine the accuracy of parental weight perceptions of overweight children before and after the implementation of childhood obesity legislation that included BMI screening and feedback. Methods and Procedures: Statewide telephone surveys of parents of overweight (BMI ≥ 85th percentile) Arkansas public school children before (n = 1,551; 15% African American) and after (n = 2,508; 15% African American) policy implementation were examined for correspondence between parental perception of child's weight and objective classification. Results: Most (60%) parents of overweight children underestimated weight at baseline. Parents of younger children were significantly more likely to underestimate (65%) than parents of adolescents (51%). Overweight parents were not more likely to underestimate, nor was inaccuracy associated with parental education or socioeconomic status. African‐American parents were twice as likely to underestimate as whites. One year after BMI screening and feedback was implemented, the accuracy of classification of overweight children improved (53% underestimation). African‐American parents had significantly greater improvements than white parents (P < 0.0001). Discussion: Parental recognition of childhood overweight may be improved with BMI screening and feedback, and African‐American parents may specifically benefit. Nonetheless, underestimation of overweight is common and may have implications for public health interventions.     

fMRI evidence for a cortical hierarchy of pitch pattern processing

Pitch patterns, such as melodies, consist of two levels of structure: a global level, comprising the pattern of ups and downs, or contour; and a local level, comprising the precise intervals that make up this contour. An influential neuropsychological model suggests that these two levels of processing are hierarchically linked, with processing of the global structure occurring within the right hemisphere in advance of local processing within the left. However, the predictions of this model and its anatomical basis have not been tested in neurologically normal individuals. The present study used fMRI and required participants to listen to consecutive pitch sequences while performing a same/different one-back task. Sequences, when different, either preserved (local) or violated (global) the contour of the sequence preceding them. When the activations for the local and global conditions were contrasted directly, additional activation was seen for local processing in right planum temporale and posterior superior temporal sulcus (pSTS). The presence of additional activation for local over global processing supports the hierarchical view that the global structure of a pitch sequence acts as a "framework" on which the local detail is subsequently hung. However, the lateralisation of activation seen in the present study, with global processing occurring in left pSTS and local processing occurring bilaterally, differed from that predicted by the neuroanatomical model. A re-examination of the individual lesion data on which the neuroanatomical model is based revealed that the lesion data equally well support the laterality scheme suggested by our data. While the present study supports the hierarchical view of local and global processing, there is an evident need for further research, both in patients and neurologically normal individuals, before an understanding of the functional lateralisation of local and global processing can be considered established.     

Steady State Bioequivalence of Generic and Innovator Formulations of Stavudine,Lamivudine, and Nevirapine in HIV-Infected Ugandan Adults

Background

Generic antiretroviral therapy is the mainstay of HIV treatment in resource-limited settings, yet there is little evidence confirming the bioequivalence of generic and brand name formulations. We compared the steady-state pharmacokinetics of lamivudine, stavudine and nevirapine in HIV-infected subjects who were receiving a generic formulation (Triomune®) or the corresponding brand formulations (Epivir®, Zerit®, and Viramune®).

Methodology/Principal Findings

An open-label, randomized, crossover study was carried out in 18 HIV-infected Ugandan subjects stabilized on Triomune-40. Subjects received lamivudine (150 mg), stavudine (40 mg), and nevirapine (200 mg) in either the generic or brand formulation twice a day for 30 days, before switching to the other formulation. At the end of each treatment period, blood samples were collected over 12 h for pharmacokinetic analysis. The main outcome measures were the mean AUC_0–12h and C_max. Bioequivalence was defined as a geometric mean ratio between the generic and brand name within the 90% confidence interval of 0.8–1.25. The geometric mean ratios and the 90% confidence intervals were: stavudine C_max, 1.3 (0.99–1.71) and AUC_0–12h, 1.1 (0.87–1.38); lamivudine C_max, 0.8 (0.63–0.98) and AUC_0–12h, 0.8 (0.65–0.99); and nevirapine C_max, 1.1 (0.95–1.23) and AUC_0–12h, 1.1 (0.95–1.31). The generic formulation was not statistically bioequivalent to the brand formulations during steady state, although exposures were comparable. A mixed random effects model identified about 50% intersubject variability in the pharmacokinetic parameters.

Conclusions/Significant Findings

These findings provide support for the use of Triomune in resource-limited settings, although identification of the sources of intersubject variability in these populations is critical.