Small molecules, big targets: drug discovery faces the protein-protein interaction challenge.

Protein-protein interactions (PPIs) are of pivotal importance in the regulation of biological systems and are consequently implicated in the development of disease states. Recent work has begun to show that, with the right tools, certain classes of PPI can yield to the efforts of medicinal chemists to develop inhibitors, and the first PPI inhibitors have reached clinical development. In this Review, we describe the research leading to these breakthroughs and highlight the existence of groups of structurally related PPIs within the PPI target class. For each of these groups, we use examples of successful discovery efforts to illustrate the research strategies that have proved most useful.JS, DES and ARB thank the Wellcome Trust for funding.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/nrd.2016.2

Digestion of Starch Granules from Maize, Potato and Wheat by Larvae of the the Yellow Mealworm, Tenebrio molitor and the Mexican Bean Weevil, Zabrotes subfasciatus

Scanning electron microscopy images were taken of starch granules from different sources following exposure in vivo and in vitro to gut α-amylases isolated from Tenebrio molitor L. (Coleoptera: Tenebrionidae) and Zabrotes subfasciatus Boheman (Coleoptera: Bruchidae). One α-amylase was isolated from whole larval midguts of T. molitor using non-denaturing SDS-PAGE, while two other α-amylase fractions were isolated from whole larval midguts of Z. subfasciatus using hydrophobic interaction chromatography., Digested starch granules from larvae fed on maize, potato or wheat were isolated from midgut contents. Combinations of starch granules with isolated α-amylases from both species showed similar patterns of granule degradation. In vitro enzymatic degradation of maize starch granules by the three different α-amylase fractions began by creating small holes and crater-like areas on the surface of the granules. Over time, these holes increased in number and area resulting in extensive degradation of the granule structure. Granules from potato did not show formation of pits and craters on their surface, but presented extensive erosion in their interior. For all types of starch, as soon as the interior of the starch granule was reached, the inner layers of amylose and amylopectin were differentially hydrolyzed, resulting in a striated pattern. These data support the hypothesis that the pattern of starch degradation depends more on the granule type than on the α-amylase involved

Incident risk factors as predictors of HIV seroconversion in the Lisbon cohort of men who have sex with men: first results, 2011-2014

HIV incidence in men who have sex with men (MSM) is increasing in western countries, including Portugal. We aimed to estimate HIV incidence and to assess how individual short-term changes in exposures over time predict seroconversion. We evaluated participants of an open cohort of HIV-negative MSM enrolled after testing at a community-based voluntary HIV counselling and testing centre in Lisbon. At each evaluation a structured questionnaire was completed and HIV status was ascertained using rapid followed by confirmatory testing. Between April 2011 and February 2014, 804 MSM were followed for a total of 893 person-years. Predictors of HIV seroconversion were identified using Poisson generalised linear regression. The overall seroincidence was 2.80/100 person-years (95% confidence interval: 1.89–4.14). Men who seroconverted had a higher mean number of tests per year. Seroconversions were significantly associated with partner disclosure of HIV status during follow-up, newly-adopted unprotected anal intercourse (UAI) with a steady partner and being newly-diagnosed with syphilis during follow-up. Likewise, sexual intercourse with HIV-positive men, having an HIV-positive steady partner at least once during follow-up and persistent UAI with occasional partners were predictors of seroconversion. High HIV incidence in this cohort is likely driven by short-term contextual and behavioural changes during follow-up

The Lisbon Cohort of men who have sex with men

Purpose Newly diagnosed HIV infections among men who have sex with men (MSM) are rising in many European countries. Surveillance tools must be tailored to the current state of the epidemic, and include decentralised prospective monitoring of HIV incidence and behavioural changes in key populations. In this scenario, an open prospective cohort study was assembled—The Lisbon Cohort of MSM—aiming to dynamically monitor the frequency of disease and its predictors. Participants The Lisbon Cohort of MSM is an ongoing observational prospective study conducted at a community-based voluntary HIV counselling and testing centre in Lisbon, Portugal (CheckpointLX). Men testing negative for HIV, aged 18 or over and reporting having had sex with men are invited to follow-up visits every 6 months. At each evaluation, a face-to-face interview using a structured questionnaire is conducted, and HIV and syphilis rapid tests are performed by trained peer counsellors. From April 2011 to February 2014, 3106 MSM were eligible to the cohort of whom 923 (29.7%) did not participate. The remaining 2183 (70.3%) MSM were enrolled and 804 had at least one follow-up evaluation, for a total of 893 person-years of observation. Future plans The study findings will be disseminated in peer-reviewed journals and presented at national and international conferences. The follow-up of this cohort of HIV-negative MSM will be a valuable tool for monitoring HIV incidence in a setting where limited prospective information existed. Moreover, it will allow for a deeper analytical approach to the study of population time trends and individual changes in risk factors that currently shape the HIV epidemic among MSM

Clinical and Cardiometabolic Effects of Reducing Sedentary Behavior in Postmenopausal Women with Rheumatoid Arthritis

Purpose: We investigated the effects of a 4-month intervention targeting sedentary behavior on sedentary time and physical activity level, clinical parameters, cardiometabolic risk factors, inflammatory markers, and health-related quality of life in post-menopausal women with rheumatoid arthritis. Methods: This was a 4-month, parallel-group, randomized controlled trial (ClinicalTrials.gov identifier: NCT03186924). One-hundred-and-three postmenopausal rheumatoid arthritis patients were randomized (1:1) to either a newly developed intervention targeting sedentary behavior (Take a STAND for Health; TS4H) or standard of care (SOC). Sedentary behavior (primary outcome) and physical activity levels, clinical parameters, anthropometric parameters and body composition, blood samples and oral glucose tolerance test, blood pressure, muscle function, and health-related quality of life were assessed at baseline (Pre) and after 4 months (Post). Between- and within-group differences were tested using linear mixed models following the intention-to-treat principle. Results: Total sedentary time, time in prolonged sitting bouts, standing, and stepping did not change in either group (all p≥0.337). No significant between- and within-group differences were detected for any of the clinical parameters, markers of cardiometabolic health and inflammation, and health-related quality of life variables (all p≥0.136). Among responders in TS4H group (those who reduced sedentary time by ≥30 min/d), Pre to Post IL-10 concentrations tended to reduce (group*time: p=0.086; estimated mean difference [EMD]: -12.0 pg/mL [-23.5 to -0.6], p=0.037), and general health (group*time: p=0.047; EMD: 10.9 A.U. [-1.1 to 22.9], p=0.086) and overall physical health tended to improve (group*time: p=0.067; EMD: 7.9 A.U. [-0.9 to 16.6], p=0.089). Conclusions: TS4H did not change sedentary behavior, physical activity levels, clinical, cardiometabolic, inflammatory or health-related quality of life outcomes. However, TS4H tended to reduce IL-10 levels and improve health-related quality of life in responders

Effects of different concentrations of carbamide peroxide and bleaching periods on the roughness of dental ceramics

The wide use of dental bleaching treatment has brought concern about the possible effects of hydrogen peroxide on dental tissue and restorative materials. The objective of this study was to evaluate in vitro the effect of nightguard bleaching on the surface roughness of dental ceramics after different periods of bleaching treatment. Fifteen specimens of 5 × 3 × 1 mm were created with three dental ceramics following the manufacturers' instructions: IPS Classic (Ivoclar-Vivadent); IPS d.Sign (Ivoclar-Vivadent); and VMK-95 (Vita). A profilometer was used to evaluate baseline surface roughness (Ra values) of all ceramics by five parallel measurements with five 0.25 mm cut off (Λc) at 0.1 mm/s. Afterwards, all specimens were submitted to 6-h daily bleaching treatments with 10% or 16% carbamide peroxide (Whiteness- FGM) for 21 days, while control groups from each ceramic system were stored in artificial saliva. The surface roughness of all groups was evaluated after 18 h, 42 h, 84 h, and 126 h of bleaching treatment. The surface roughness of each specimen (n = 5) was based on the mean value of five parallel measurements in each time and all data were submitted to two-way repeated measures ANOVA and Tukey's post-hoc test (α = 0.05). No significant differences in ceramic surface roughness were observed between untreated and bleached ceramic surfaces, regardless of bleaching intervals or bleaching treatments. This study provided evidence that at-home bleaching systems do not cause detrimental effects on surface roughness of dental ceramics

Position and sequence conservation in Amniota of polymorphic enhancer HS1.2 within the palindrome of IgH 3'Regulatory Region

<p>Abstract</p> <p>Background</p> <p>The Immunoglobulin heavy chain (IgH) 3' Regulatory Region (3'RR), located at the 3' of the constant alpha gene, plays a crucial role in immunoglobulin production. In humans, there are 2 copies of the 3'RR, each composed of 4 main elements: 3 enhancers and a 20 bp tandem repeat. The single mouse 3'RR differs from the two human ones for the presence of 4 more regulative elements with the double copy of one enhancer at the border of a palindromic region.</p> <p>Results</p> <p>We compared the 3'RR organization in genomes of vertebrates to depict the evolutionary history of the region and highlight its shared features. We found that in the 8 species in which the whole region was included in a fully assembled contig (mouse, rat, dog, rabbit, panda, orangutan, chimpanzee, and human), the shared elements showed synteny and a highly conserved sequence, thus suggesting a strong evolutionary constraint. In these species, the wide 3'RR (~30 kb in human) bears a large palindromic sequence, consisting in two ~3 kb complementary branches spaced by a ~3 kb sequence always including the HS1.2 enhancer. In mouse and rat, HS3 is involved by the palindrome so that one copy of the enhancer is present on each side. A second relevant feature of our present work concerns human polymorphism of the HS1.2 enhancer, associated to immune diseases in our species. We detected a similar polymorphism in all the studied Catarrhini (a primate parvorder). The polymorphism consists of multiple copies of a 40 bp element up to 12 in chimpanzees, 8 in baboons, 6 in macaque, 5 in gibbons, 4 in humans and orangutan, separated by stretches of Cytosine. We show specific binding of this element to nuclear factors.</p> <p>Conclusions</p> <p>The nucleotide sequence of the palindrome is not conserved among evolutionary distant species, suggesting pressures for the maintenance of two self-matching regions driving a three-dimensional structure despite of the inter-specific divergence at sequence level. The information about the conservation of the palindromic structure and the settling in primates of the polymorphic feature of HS1.2 show the relevance of these structures in the control and modulation of the Ig production through the formation of possible three-dimensional structures.</p

VE-statin/egfl7 Expression in Endothelial Cells Is Regulated by a Distal Enhancer and a Proximal Promoter under the Direct Control of Erg and GATA-2

Angiogenesis is the process by which new blood vessels arise from existing ones by the budding out of endothelial cell capillaries from the luminal side of blood vessels. Blood vessel formation is essential for organ development during embryogenesis and is associated with several physiological and pathological processes, such as wound healing and tumor development. The VE-statin/egfl7 gene is specifically expressed in endothelial cells during embryonic development and in the adult. We studied here the regulatory mechanisms that control this tissue-specific expression. RT-qPCR analyses showed that the specificity of expression of VE-statin/egfl7 in endothelial cells is not shared with its closest neighbor genes notch1 and agpat2 on the mouse chromosome 2. Chromatin-immunoprecipitation analysis of histone modifications at the VE-statin/egfl7 locus showed that the chromatin is specifically opened in endothelial cells, but not in fibroblasts at the transcription start sites. A 13 kb genomic fragment of promoter was cloned and analyzed by gene reporter assays which showed that two conserved regions are important for the specific expression of VE-statin/egfl7 in endothelial cells; a −8409/−7563 enhancer and the −252/+38 region encompassing the exon-1b transcription start site. The latter contains essential GATA and ETS-binding sites, as assessed by linker-scanning analysis and site-directed mutagenesis. An analysis of expression of the ETS and GATA transcription factors showed that Erg, Fli-1 and GATA-2 are the most highly expressed factors in endothelial cells. Erg and GATA-2 directly control the expression of the endogenous VE-statin/egfl7 while Fli-1 probably exerts an indirect control, as assessed by RNA interference and chromatin immunoprecipitation. This first detailed analysis of the mechanisms that govern the expression of the VE-statin/egfl7 gene in endothelial cells pinpoints the specific importance of ETS and GATA factors in the specific regulation of genes in this cell lineage