A trial-based cost-effectiveness analysis of antibiotic prescription strategies for non-complicated respiratory tract infections in children.

BACKGROUND: Antibiotic prescription for respiratory tract infections (RTIs) in children attending primary care centres is almost double that predicted according to bacterial prevalence. Delayed antibiotic prescription (DAP) is designed to deploy a more rational use of antibiotics. While studies have evaluated DAP efficacy and safety for children with RTIs, little research has been conducted on the economic implications. METHODS: Our trial compared cost-effectiveness for DAP, immediate antibiotic prescription (IAP), and no antibiotic prescription (NAP) for children aged 2-14 years with acute uncomplicated RTIs attended to in 39 primary care centres in Spain. The main outcome was the incremental cost-effectiveness ratio (ICER), measured in euros per gained quality-adjusted life days (QALDs). Net monetary benefit (NMB) was also calculated as a tool for decision making. The analysis was performed from a societal perspective for a time horizon of 30 days, and included healthcare direct costs, non-healthcare direct and indirect costs, and the antimicrobial resistance (AMR) cost. RESULTS: DAP was the most cost-effective strategy, even when the cost of AMR was included. QALD values for the three strategies were very similar. IAP compared to DAP was more costly (109.68 vs 100.90 euros) and similarly effective (27.88 vs 27.94 QALDs). DAP compared to NAP was more costly (100.90 vs 97.48 euros) and more effective (27.94 vs. 27.82 QALDs). The ICER for DAP compared to NAP was 28.84 euros per QALD. The deterministic sensitivity analysis indicated that non-healthcare indirect costs had the greatest impact on the ICER. The cost-effectiveness acceptability curve showed that DAP was the preferred option in approximately 81.75% of Monte Carlo iterations, assuming a willingness-to-pay value of 82.2 euros per gained QALD. CONCLUSIONS: When clinicians are in doubt about whether an antibiotic is needed for children with RTIs attending PC centres, those treated with the DAP strategy will have slightly better efficiency outcomes than those treated with IAP because its costs are lower than those of IAP. DAP is also the most cost-effective strategy over a time horizon of 30 days if AMR is considered, despite higher short-term costs than NAP. However, if in the long term the costs of AMR are larger than estimated, NAP could also be an alternative strategy. TRIAL REGISTRATION: This trial has been registered at www. CLINICALTRIALS: gov (identifier NCT01800747; Date: 28/02/2013 (retrospectively registered).

Interaction of ammonium nutrition with essential mineral cations.

Plant growth and development depend on sufficient nutrient availability in soils. Agricultural soils are generally nitrogen (N) deficient, and thus soils need to be supplemented with fertilizers. Ammonium (NH4+) is a major inorganic N source. However, at high concentrations, NH4+ becomes a stressor that inhibits plant growth. The cause of NH4+ stress or toxicity is multifactorial, but the interaction of NH4+ with other nutrients is among the main determinants of plants' sensitivity towards high NH4+ supply. In addition, NH4+ uptake and assimilation provoke the acidification of the cell external medium (apoplast/rhizosphere), which has a clear impact on nutrient availability. This review summarizes current knowledge, at both the physiological and the molecular level, of the interaction of NH4+ nutrition with essential mineral elements that are absorbed as cations, both macronutrients (K+, Ca2+, Mg2+) and micronutrients (Fe2+/3+, Mn2+, Cu+/2+, Zn2+, Ni2+). We hypothesize that considering these nutritional interactions, and soil pH, when formulating fertilizers may be key in order to boost the use of NH4+-based fertilizers, which have less environmental impact compared with nitrate-based ones. In addition, we are convinced that better understanding of these interactions will help to identify novel targets with the potential to improve crop productivity.

Exploiting Tomato Genotypes to Understand Heat Stress Tolerance.

Increased temperatures caused by climate change constitute a significant threat to agriculture and food security. The selection of improved crop varieties with greater tolerance to heat stress is crucial for the future of agriculture. To overcome this challenge, four traditional tomato varieties from the Mediterranean basin and two commercial genotypes were selected to characterize their responses at high temperatures. The screening of phenotypes under heat shock conditions allowed to classify the tomato genotypes as: heat-sensitive: TH-30, ADX2; intermediate: ISR-10 and Ailsa Craig; heat-tolerant: MM and MO-10. These results reveal the intra-genetical variation of heat stress responses, which can be exploited as promising sources of tolerance to climate change conditions. Two different thermotolerance strategies were observed. The MO-10 plants tolerance was based on the control of the leaf cooling mechanism and the rapid RBOHB activation and ABA signaling pathways. The variety MM displayed a different strategy based on the activation of HSP70 and 90, as well as accumulation of phenolic compounds correlated with early induction of PAL expression. The importance of secondary metabolism in the recovery phase has been also revealed. Understanding the molecular events allowing plants to overcome heat stress constitutes a promising approach for selecting climate resilient tomato varieties.

Putrescine: A Key Metabolite Involved in Plant Development, Tolerance and Resistance Responses to Stress.

Putrescine (Put) is the starting point of the polyamines (PAs) pathway and the most common PA in higher plants. It is synthesized by two main pathways (from ornithine and arginine), but recently a third pathway from citrulline was reported in sesame plants. There is strong evidence that Put may play a crucial role not only in plant growth and development but also in the tolerance responses to the major stresses affecting crop production. The main strategies to investigate the involvement of PA in plant systems are based on the application of competitive inhibitors, exogenous PAs treatments, and the most efficient approaches based on mutant and transgenic plants. Thus, in this article, the recent advances in understanding the role of this metabolite in plant growth promotion and protection against abiotic and biotic stresses will be discussed to provide an overview for future research.

Putrescine biosynthetic pathways modulate root growth differently in tomato seedlings grown under different N sources.

The biosynthesis of putrescine is mainly driven by arginine decarboxylase (ADC) and ornithine decarboxylase (ODC). Hence, in this study, we generated independent ADC and ODC transgenic silenced tomato lines (SilADC and SilODC, respectively) to test the effect of defective ADC and ODC gene expression on root development under nitrate (NN) or ammonium (NA) conditions. The results showed that SilODC seedlings displayed an increase in ADC expression that led to polyamine accumulation, suggesting a compensatory effect of ADC. However, this effect was not observed in SilADC seedlings. These pathways are involved in different growth processes. The SilADC seedlings showed an increase in fresh weight, shoot length, lateral root number and shoot:root ratio under the NN source and an enhancement in fresh weight, and shoot and root length under NA conditions. However, SilODC seedlings displayed greater weight and shoot length under the NN source, whereas a decrease in lateral root density was found under NA conditions. Moreover, two overexpressed ODC lines were generated to check the relevance of the compensatory effect of the ADC pathway when ODC was silenced. These overexpressed lines showed not only an enhancement of almost all the studied growth parameters under both N sources but also an amelioration of ammonium syndrome under NA conditions. Together, these results reflect the importance of both pathways in plant growth, particularly ODC silencing, which requires compensation by ADC induction.

Delayed Antibiotic Prescription for Children With Respiratory Infections: A Randomized Trial.

OBJECTIVES: To assess the effectiveness and safety of delayed antibiotic prescription (DAP) compared to immediate antibiotic prescription (IAP) and no antibiotic prescription (NAP) in children with uncomplicated respiratory infections. METHODS: Randomized clinical trial comparing 3 antibiotic prescription strategies. The participants were children with acute uncomplicated respiratory infections attended to in 39 primary care centers. Children were randomly assigned into prescription arms as follows: (1) DAP, (2) IAP, or (3) NAP. Primary outcomes were symptom duration and severity. Secondary outcomes were antibiotic use, parental satisfaction, parental beliefs, additional primary care visits, and complications at 30 days. RESULTS: In total, 436 children were included in the analysis. The mean (SD) duration of severe symptoms was 10.1 (6.3) for IAP, 10.9 (8.5) for NAP, and 12.4 (8.4) for DAP (P = .539), although the differences were not statistically significant. The median (interquartile range) of the greatest severity for any symptom was similar for the 3 arms (median [interquartile range] score of 3 [2-4]; P = .619). Antibiotic use was significantly higher for IAP (n = 142 [96%]) compared to DAP (n = 37 [25.3%]) and NAP (n = 17 [12.0%]) (P < .001). Complications, additional visits to primary care, and satisfaction were similar for all strategies. Gastrointestinal adverse effects were higher for IAP. CONCLUSIONS: There was no statistically significant difference in symptom duration or severity in children with uncomplicated respiratory infections who received DAP compared to NAP or IAP strategies; however, DAP reduced antibiotic use and gastrointestinal adverse effects.

Temporal stability and clinical validation of the Spanish version of the female sexual function inventory (FSFI).

The Female Sexual Function Index is one of the most common instruments used to evaluate the female sexual function. The present study aimed to analyze the test-retest reliability, internal consistency, and discriminant validity of the index in clinical samples and to determine a specific and sensitive cutoff point for the Spanish version of the Female Sexual Function Index. For that purpose, a sample consisting of 117 Colombian women was recruited to evaluate test-retest reliability, and a second sample, consisting of 185 women, was divided into diagnosis and no-diagnosis groups based on DSM-5 criteria. Results showed adequate test-retest reliability after four weeks, and satisfactory evidence of internal consistency was obtained for subscale and overall scores. The inventory was found to have an adequate criterion validity, and it confirmed the differences between diagnosis and no-diagnosis groups. The instrument's cutoff point was determined to be 26 points, with a specificity of 73.9%, a sensitivity of 87.7%, and an area under the curve of 85.9 (CI = 80.0-91.7). These results confirm that the Spanish version of the FSFI is an adequate tool for evaluating female sexual dysfunction based on DSM-5 criteria.

Novel murine mAbs define specific and cross-reactive epitopes on the latex profilin panallergen Hev b 8.

The production of specific antibodies able to recognize allergens from different sources or block interactions between allergens and antibodies mediating allergic reactions is crucial for developing successful tools for diagnostics and therapeutics. Panallergens are highly conserved proteins present in widely different species, implicated in relevant cross-reactions. The panallergen latex profilin (Hev b 8) has been associated with the latex-food-pollen syndrome. We generated five monoclonal IgGs and one IgE from murine hybridomas against recombinant Hev b 8 and evaluated their interaction with this allergen using ELISA and biolayer interferometry (BLI). Affinity purified mAbs exhibited high binding affinities towards rHev b 8, with KD1 values ranging from 10-10 M to 10-11 M. Some of these antibodies also recognized the recombinant profilins from maize and tomato (Zea m 12 and Sola l 1), and the ash tree pollen (Fra e 2). Competition ELISA demonstrated that some mAb pairs could bind simultaneously to rHev b 8. Using BLI, we detected competitive, non-competitive, and partial-competition interactions between pairs of mAbs with rHev b 8, suggesting the existence of at least two non-overlapping epitopes on the surface of this allergen. Three-dimensional models of the Fv of 1B4 and 2D10 IgGs and docking simulations of these Fvs with rHev b 8 revealed these epitopes. Furthermore, these two mAbs inhibited the interaction of polyclonal IgE and IgG4 antibodies from profilin-allergic patients with rHev b 8, indicating that the mAbs and the antibodies present in sera from allergic patients bind to overlapping epitopes on the allergen. These mAbs can be useful tools for immune-localization studies, immunoassay development, or standardization of allergenic products.

The Apoplast: A Key Player in Plant Survival.

The apoplast comprises the intercellular space, the cell walls, and the xylem. Important functions for the plant, such as nutrient and water transport, cellulose synthesis, and the synthesis of molecules involved in plant defense against both biotic and abiotic stresses, take place in it. The most important molecules are ROS, antioxidants, proteins, and hormones. Even though only a small quantity of ROS is localized within the apoplast, apoplastic ROS have an important role in plant development and plant responses to various stress conditions. In the apoplast, like in the intracellular cell compartments, a specific set of antioxidants can be found that can detoxify the different types of ROS produced in it. These scavenging ROS components confer stress tolerance and avoid cellular damage. Moreover, the production and accumulation of proteins and peptides in the apoplast take place in response to various stresses. Hormones are also present in the apoplast where they perform important functions. In addition, the apoplast is also the space where microbe-associated molecular Patterns (MAMPs) are secreted by pathogens. In summary, the diversity of molecules found in the apoplast highlights its importance in the survival of plant cells.

Exogenous Carbon Compounds Modulate Tomato Root Development.

NO3- is not only a nutrient, but also a signaling compound that plays an important role in several plant processes, like root development. The present study aimed to investigate the effect of three different exogenous C compounds (sucrose, glucose, 2-oxoglutarate) added to NO3- nutrition on C/N, auxin and antioxidant metabolisms in 10-day-old tomato seedlings. Sucrose and glucose supplementation enhanced primary root (PR) length, lateral root number and root density, while 2-oxoglutarate negatively affected them. This phenomenon was accompanied by a slight increase in NRT2.1 and GS1 gene expression, together with an increase in LAX2 and LAX3 and a decrease in LAX4 in the roots growing under sucrose and glucose sources. The addition of 2-oxoglutarate enhanced the expression of NiR, GDH, PEPC1, LAX1, LAX3 and the antioxidant gene SOD Cl. Taken together, these findings contribute to a better understanding of how these C sources can modulate N uptake and C/N, auxin and antioxidant gene expression, which could be useful for improving nitrogen use efficiency.

Tomato root development and N assimilation depend on C and ABA content under different N sources.

Root plasticity is controlled by hormonal homeostasis and nutrient availability. In this work, we have determined the influence of different N regimens on growth parameters and on the expression of genes involved in auxin transport and N-assimilation in tomato seedlings. NH4+ nutrition led to an inhibitory effect on root fresh weight (FW), lateral root (LR) number and root density, while an increase in the primary root (PR) length was observed. The expression of N assimilation genes GS2 and ASN1, is affected by NH4+ nutrition. Moreover, in order to relieve the toxic effect of NH4+ on root development, glucose or 2-oxoglutarate was supplied as a C source during NH4+ treatment. The addition of 2-oxoglutarate improved root parameters compared to the NH4+ regimen. N-assimilation gene analysis showed that NH4+-fed tomato plants try to alleviate the toxic effect by concurrently upregulating ASN1 and anaplerotic PEPC2 expression, whereas when 2-oxoglutarate is supplied, ASN1 induction was not observed. The addition of both C skeletons induced the expression of the ROS-scavenging genes GSH and SOD. In addition, since ABA plays a role in root development, the ABA-synthesis-defective mutant flacca was studied under NO3- and NH4+ regimens. It displayed a decrease in LR number under NO3- conditions, whereas, the NH4+-fed seedlings showed a decrease solely in PR length that was reverted when ABA was exogenously supplied. Moreover, flacca seedlings displayed a reprogramming of the N/C assimilation genes. Altogether, these results reflect the importance of N and C sources and ABA homeostasis in root development of tomato seedlings.

Ammonium mediated changes in carbon and nitrogen metabolisms induce resistance against Pseudomonas syringae in tomato plants.

Predominant NH4+ nutrition causes an "ammonium syndrome" that induces metabolic changes and thereby provides resistance against Pseudomonas syringae infection through the activation of systemic acquired acclimation (SAA). Hence, to elucidate the mechanisms underlying NH4+-mediated SAA, the changes in nutrient balance and C and N skeletons were studied in NH4+-treated plants upon infection by P. syringae. A general decrease in cation and an increase in anion levels was observed in roots and leaves of NH4+-treated plants. Upon NH4+-based nutrition and infection, tomato leaves showed an accumulation of S, P, Zn, and of Mn. Mn accumulation might be required for ROS detoxification since it acts as a cofactor of superoxide dismutase (SOD). Primary metabolism was modified in both tissues of NH4+-fed plants to counteract NH4+ toxicity by decreasing TCA intermediates. A significant increase in Arg, Gln, Asn, Lys, Tyr, His and Leu was observed in leaves of NH4+-treated plants. The high level of the putrescine precursor Arg hints towards the importance of the Glu pathway as a key metabolic check-point in NH4+-treated and infected plants. Taken together, NH4+-fed plants displayed a high level of basal responses allowing them to activate SAA and to trigger defense responses against P. syringae through nutrient imbalances and changes in primary metabolism.

Fra-2-expressing macrophages promote lung fibrosis in mice.

Idiopathic Pulmonary Fibrosis (IPF) is a deadly disease with limited therapies. Tissue fibrosis is associated with Type 2 immune response, although the causal contribution of immune cells is not defined. The AP-1 transcription factor Fra-2 is upregulated in IPF lung sections and Fra-2 transgenic mice (Fra-2tg) exhibit spontaneous lung fibrosis. Here we show that Bleomycin-induced lung fibrosis is attenuated upon myeloid-inactivation of Fra-2 and aggravated in Fra-2tg bone marrow chimeras. Type VI collagen (ColVI), a Fra-2 transcriptional target, is up-regulated in three lung fibrosis models, and macrophages promote myofibroblast activation in vitro in a ColVI- and Fra-2-dependent manner. Fra-2 or ColVI inactivation does not affect macrophage recruitment and alternative activation, suggesting that Fra-2/ColVI specifically controls the paracrine pro-fibrotic activity of macrophages. Importantly, ColVI knock-out mice (KO) and ColVI-KO bone marrow chimeras are protected from Bleomycin-induced lung fibrosis. Therapeutic administration of a Fra-2/AP-1 inhibitor reduces ColVI expression and ameliorates fibrosis in Fra-2tg mice and in the Bleomycin model. Finally, Fra-2 and ColVI positively correlate in IPF patient samples and co-localize in lung macrophages. Therefore, the Fra-2/ColVI pro-fibrotic axis is a promising biomarker and therapeutic target for lung fibrosis, and possibly other fibrotic diseases.

Discovery of novel triazolo[4,3-b]pyridazin-3-yl-quinoline derivatives as PIM inhibitors.

PIM kinase family (PIM-1, PIM-2 and PIM-3) is an appealing target for the discovery and development of selective inhibitors, useful in various disease conditions in which these proteins are highly expressed, such as cancer. The significant effort put, in the recent years, towards the development of small molecules exhibiting inhibitory activity against this protein family has ended up with several molecules entering clinical trials. As part of our ongoing exploration for potential drug candidates that exhibit affinity towards this protein family, we have generated a novel chemical series of triazolo[4,3-b]pyridazine based tricycles by applying a scaffold hopping strategy over our previously reported potent pan-PIM inhibitor ETP-47453 (compound 2). The structure-activity relationship studies presented herein demonstrate a rather selective PIM-1/PIM-3 biochemical profile for this novel series of tricycles, although pan-PIM and PIM-1 inhibitors have also been identified. Selected examples show significant inhibition of the phosphorylation of BAD protein in a cell-based assay. Moreover, optimized and highly selective compounds, such as 42, did not show significant hERG inhibition at 20 µM concentration, and proved its antiproliferative activity and utility in combination with particular antitumoral agents in several tumor cell lines.

Identification of novel PI3K inhibitors through a scaffold hopping strategy.

A scaffold hopping strategy, including intellectual property availability assessment, was successfully applied for the discovery of novel PI3K inhibitors. Compounds were designed based on the chemical structure of the lead compound ETP-46321, a potent PI3K inhibitor, previously reported by our group. The new generated compounds showed good in vitro potency and selectivity, proved to inhibit potently the phosphorylation of AKTSer473 in cells and demonstrated to be orally bioavailable, thus becoming potential back-up candidates for ETP-46321.

Generation of tricyclic imidazo[1,2-a]pyrazines as novel PI3K inhibitors by application of a conformational restriction strategy.

The involvement of the phosphoinositide 3-kinases (PI3Ks) in several diseases, especially in the oncology area, has singled it as one of the most explored therapeutic targets in the last two decades. Many different inhibitor classes have been developed by the industry and academia with a diverse selectivity profile within the PI3K family. In the present manuscript we report a further exploration of our lead PI3K inhibitor ETP-46321 (Martínez González et al., 2012)1 by the application of a conformational restriction strategy. For that purpose we have successfully synthesized novel tricyclic imidazo[1,2-a]pyrazine derivatives as PI3K inhibitors. This new class of compounds had enable the exploration of the solvent-accessible region within PI3K and resulted in the identification of molecule 8q with the best selectivity PI3Kα/δ isoform profile in vitro, and promising in vivo PK data.

Rapid identification of ETP-46992, orally bioavailable PI3K inhibitor, selective versus mTOR.

Phosphoinositide-3-kinases (PI3K) are a family of lipid kinases mediating numerous cell processes such as proliferation, migration and differentiation. PI3K is an important target for cancer therapeutics due to the deregulation of this signaling pathway in a wide variety of human cancers. Herein, we describe the rapid identification of ETP-46992, within 2-aminocarbonyl imidazo [1,2-a] pyrazine series, with suitable pharmacokinetic (PK) properties that allows the establishment of mechanism of action and efficacy in vivo studies. ETP-46992 showed tumor growth inhibition in a GEMM mouse tumor model driven by a K-Ras(G12V) oncogenic mutation and in tumor xenograft models with PI3K pathway deregulated (BT474).

Identification of ETP-46321, a potent and orally bioavailable PI3K α, δ inhibitor.

Phosphoinositide-3-kinase (PI3K) is an important target for cancer therapeutics due to the deregulation of this signaling pathway in a wide variety of human cancers. Herein, we describe the optimization of imidazo [1,2-a] pyrazines, which allow us to identify compound 14 (ETP-46321), with potent biochemical and cellular activity and good pharmacokinetic properties (PK) after oral dosing. ETP-46321 PK/PD studies showed time dependent downregulation of AKT(Ser473) phosphorylation, which correlates with compound levels in tumor tissue and demonstrating to be efficacious in a GEMM mouse tumor model driven by a K-Ras(G12V) oncogenic mutation. Treatment with ETP-46321 resulted in significant tumor growth inhibition.

Imidazo[1,2-a]pyrazines as novel PI3K inhibitors.

Phosphoinositide-3-kinase (PI3K) is an important target for cancer therapeutics due to the deregulation of its signaling pathway in a wide variety of human cancers. We describe herein a novel series of imidazo[1,2-a]pyrazines as PI3K inhibitors.

Phosphorylated insulin-like growth factor binding protein-1 and cervical measurement in women with threatening preterm birth.

OBJECTIVE: To assess the efficacy of the cervical phosphorylated insulin-like growth factor binding protein-1 (pIGFBP-1) in combination with cervical length measurement for the prediction of preterm delivery. DESIGN: Observational prospective study. SETTING: University Hospital, Spain. SAMPLE: A total of 276 women between 24 and 34 weeks gestational age complaining of uterine contractions and intact membranes. METHODS: Transvaginal scan to assess cervical length and cervical swabs for pIGFBP-1 detection. MAIN OUTCOME MEASURES: Prevalence of preterm delivery within 48 hours and 7 days, delivery at < 32, < 34 weeks, admission-to-delivery interval by cervical length and the presence of pIGFBP-1. Results. The prevalence of preterm delivery at < 34 weeks was 14.1% (39/276) and 7.6% (21/276) at < 32 weeks. Cervical pIGFBP-1 was positive in 38.7% (107/276), whereas cervical length < 15 mm was present in 7.2% (20/276) and was > 30 mm for 64.8% (179/276) of the women. The presence of a positive pIGFBP-1 appeared useful for prediction of delivery before 32 (likelihood ratio (LR) = 2.21, confidence interval (CI): 1.63-3.00) and 34 weeks (LR = 1.76, CI: 1.25-2.41), respectively. Receiver operating characteristic curves were significant for cervical length and pIGFBP-1 for prediction of delivery at < 34 weeks, < 32 weeks, < 48 hours and < 7 days. Combining pIGFBP-1 and cervical length had a median interval that was shorter when both were positive as compared to only one positive or both negative, but the CIs overlapped. CONCLUSIONS: Both cervical length measurement and pIGFBP-1 are useful for prediction of spontaneous preterm delivery. A pIGFBP-1 positive result associated with cervical length does not improve preterm delivery prediction rates.