Identification of active main metabolites of anti-infective inhibitors of the macrophage infectivity potentiator protein by liquid chromatography using mass detection.

Due to increasing antibiotic resistance, the development of anti-infectives with new mechanisms of action is crucial. Virulence factors such as the "macrophage infectivity potentiator" (Mip) protein, which catalyzes the folding of proline-containing proteins by means of their cis-trans isomerase (PPIase) activity, have come into focus as a potential new target. Since the inhibition of Mip by small molecules has been shown to lead to reduced virulence and survival in vitro, especially of Gram-negative bacteria such as Burkholderia pseudomallei (Bp), Neisseria meningitidis (Nm), and Neisseria gonorrhoeae (Ng), or Coxiella burnetii (Cb), among many others, a library of Mip inhibitors was developed. As drug metabolism has a significant impact on the overall therapeutic outcome, this report describes the biotransformation of the most potent Mip inhibitors. Therefore, the anti-infectives were treated using human liver microsomes in vitro. Liquid chromatography with tandem mass spectrometry (LC/MS-MS) methods were applied to identify the metabolites and quantify the metabolic degradation of the hit compounds. Active metabolites, N-oxides, were found, leading to new opportunities for further drug development.

Measuring dysfunctional interpersonal beliefs: validation of the Interpersonal Cognitive Distortions Scale among a heterogeneous German-speaking sample.

BACKROUND: Dysfunctional interpersonal beliefs (DIBs) are a key symptom domain in numerous mental disorders. Because DIBs exert a strong influence on social experience and behavior, they play an important role in a mental disorder's development and progression. To date, only the Interpersonal Cognitive Distortions Scale (ICDS) captures DIBs independently of specific disorders, populations, or contexts. The present study's aim was to psychometrically evaluate and validate a German translation of the ICDS. METHODS: The ICDS was administered along with indicators of convergent (rejection sensitivity, depressive expectations, interpersonal trust, interpersonal problems, perceived social support), discriminant (self-efficacy, perseverative negative thinking, optimism), and clinical validity (psychopathology, perceived stress, well-being) to a pooled sample incorporating non-clinical (N = 114) and clinical (N = 94) participants. RESULTS: An exploratory factor analysis (EFA) suggested a five-factor solution (factor loadings: .44 to .85). Correlational analyses demonstrated acceptable convergent (ρ = -.29 to -.35, ρ = .27 to .59), suboptimal discriminant (ρ = -.27 to -.38, ρ = .52), and acceptable clinical validity (ρ = -.21, ρ = .36 to .44) at the total-scale level. However, results at the subscale level were mixed and required nuanced interpretation. Likewise, internal consistency was acceptable at the total-scale level (α = .76), but ranged from good to poor at the subscale level (α = .61 to .80). DIBs mediated the negative relationship between mental disorder onset and psychopathology levels. DISCUSSION: Our results imply DIBs' relevance to mental health and related outcomes. When working with the ICDS's German version, we recommend employing only the "insecurity" subscale, as this was the only scale revealing acceptable psychometric properties. Future studies should improve the construct validity of the ICDS (and its subscales), e.g., by adding more items to the respective subscales and further classes of DIBs.

The solvent- and surface-dependent adsorption of the lipopeptide antibiotic daptomycin: The general necessity of adsorption tests.

The impact of poor or non-reproducible analyte recoveries due to non-specific drug adsorption on various analytical assays is often underestimated. Even internationally approved guidelines for pharmaceutical analysis such as the EMA guideline on bioanalytical method validation, the ICH guideline M10 on bioanalytical method validation and study sample analysis or the FDA bioanalytical method validation guidance do not adequately encourage more detailed investigations. Furthermore, other areas of research in which the concentration of active pharmaceutical compounds plays a crucial role, for example screening for minimal inhibitory concentrations of bacterial isolates, are potentially affected as well. The aim of this study was to demonstrate the general necessity of drug adsorption tests, using the lipopeptide antibiotic daptomycin as an example. A wide range of typical materials used in processing samples in pharmaceutical and biological analysis, as well as various solvents and biological matrices were included in the experiments. A fully validated LC-MS/MS method was applied for the determination of daptomycin concentrations, which were subsequently used to calculate the recovery. Recovery results (n = 3) ranged from 0.00% to 102.12% with a maximum relative standard deviation of 12.78%. These findings demonstrate that recovery can vary greatly depending on the solvent and the contact material, indicating the need to be optimized and, if applicable, validated. Hence, high reproducibility can only be achieved if all materials (and their manufacturers) used in a method are specified, not just those used in steps considered critical.

How to modify expectations of social rejection? An experimental study using a false-feedback paradigm.

BACKGROUND AND OBJECTIVES: Negative expectations (NEs) are fundamental to various mental disorders. Finding ways to modulate NEs would help to improve clinical treatment. The present study investigated how previously formed expectations of social rejection are revised in the context of novel positive social experiences, and whether their revision can be modulated by differentially shifting participants' attentional focus. METHODS: Our sample of 124 healthy participants was randomly assigned to four experimental conditions and received manipulated social feedback in multiple alleged webcam conferences. All groups went through three experimental phases that began with predominantly negative social feedback, then either transitioned to predominantly positive social feedback or continued to predominantly negative social feedback, and ultimately transitioning to a phase with no explicit social feedback. The experimental conditions differed in what they were instructed to focus on when receiving positive social feedback. RESULTS: Receiving novel positive social feedback led to substantial changes in social expectations, but this effect was not modulated by the instructions the participants were given. Descriptive trends revealed that both instructions improved NE modification, although this effect was not robust to extinction in one condition. LIMITATIONS: To prevent our cover story from being compromised, we could not perform an immediate manipulation check of the instructions given. Nevertheless, some of the sample seemed suspicious about the cover story. CONCLUSION: Our results suggest that established expectations of social rejection can be revised when unexpectedly experiencing social acceptance. Nevertheless, more research is needed on potential instructions that could be used to optimize the modification of NEs.

Fluorescent probe for the identification of potent inhibitors of the macrophage infectivity potentiator (Mip) protein of Burkholderia pseudomallei.

The macrophage infectivity potentiator (Mip) protein belongs to the immunophilin superfamily. This class of enzymes catalyzes the interconversion between the cis and trans configuration of proline-containing peptide bonds. Mip has been shown to be important for the virulence of a wide range of pathogenic microorganisms, including the Gram-negative bacterium Burkholderia pseudomallei. Small molecules derived from the natural product rapamycin, lacking its immunosuppression-inducing moiety, inhibit Mip's peptidyl-prolyl cis-trans isomerase (PPIase) activity and lead to a reduction in pathogen load in vitro. Here, a fluorescence polarization assay (FPA) to enable the screening and effective development of BpMip inhibitors was established. A fluorescent probe was prepared, derived from previous pipecolic scaffold Mip inhibitors labeled with fluorescein. This probe showed moderate affinity for BpMip and enabled a highly robust FPA suitable for screening large compound libraries with medium- to high-throughput (Z factor ∼ 0.89) to identify potent new inhibitors. The FPA results are consistent with data from the protease-coupled PPIase assay. Analysis of the temperature dependence of the probe's binding highlighted that BpMip's ligand binding is driven by enthalpic rather than entropic effects. This has considerable consequences for the use of low-temperature kinetic assays.

Development, validation and application of a selective and sensitive LC-MS/MS method for the quantification of daptomycin in a suspension of Mammaliicoccus sciuri in Mueller-Hinton broth.

Reports of therapy failures related to the use of daptomycin (DAP) are steadily increasing. This is mainly due to emerging DAP resistance for which, however, the underlying mechanism is often unknown. To elucidate the mode of action of novel DAP resistance traits it is indispensable to reliably detect and quantify DAP in complex matrices such as bacterial culture media. In this study, we established a selective and sensitive quantification method for DAP upon growth of a DAP resistant Mammaliicoccus sciuri strain in Mueller-Hinton medium. The method combined methanol-induced protein precipitation followed by high performance liquid chromatography with gradient elution coupled to mass spectrometry (LC-MS/MS) using daptomycin-d5 as internal standard. All validation parameters met the acceptance criteria of the European Medicines Agency (EMA) guideline on bioanalytical method validation. We successfully applied this highly selective and sensitive LC-MS/MS method for the quantification of DAP in in vitro studies addressing DAP resistance mechanisms in Gram-positive bacteria.

Design, synthesis and biological evaluations of quinolone amides against African trypanosomiasis with improved solubility.

The human African trypanosomiasis is a devastating parasitic infection, which is caused by the protozoan Trypanosoma brucei and transmitted by the bite of the tsetse fly. An untreated infection usually results in death and only few drugs with significant drawbacks are currently available for treatment. Previous investigations revealed the quinolone amide MB007 as a lead compound with an excellent selectivity for T. b. brucei. Here, new quinolone amides were synthesized for deeper insights into the structure-activity relationship. Furthermore, the aqueous solubility of the compounds was analyzed, as the poor solubility of previous quinolone amides impeded in vivo studies for target identification. The biological evaluation led to the new lead structure 9f, which exhibits a promising in vitro activity against T. b. brucei (IC50 = 22 nM) and showed no cytotoxicity against macrophages. Moreover, compounds 10b and 10c were discovered, which possessed an improved solubility combined with a decent selectivity.

How negative mood hinders belief updating in depression: results from two experimental studies.

BACKGROUND: In two experimental studies, we tested the hypothesis that negative mood would hinder the revision of negative beliefs in response to unexpectedly positive information in depression, whereas positive mood was expected to enhance belief updating. METHODS: In study 1 (N = 101), we used a subclinical sample to compare the film-based induction of sad v. happy mood with a distraction control group. Subsequently, participants underwent a well-established paradigm to examine intra-individual changes in performance-related expectations after unexpectedly positive performance feedback. In study 2, we applied the belief-updating task from study 1 to an inpatient sample (N = 81) and induced sad v. happy mood via film-clips v. recall of autobiographic events. RESULTS: The results of study 1 showed no significant group differences in belief updating; the severity of depressive symptoms was a negative predictor of belief revision, though, and there was a non-significant trend suggesting that the presence of sad mood hindered belief updating in the subgroup of participants with a diagnosed depressive episode. Study 2 revealed that participants updated their expectations significantly less in line with positive feedback when they underwent the induction of negative mood prior to feedback, relative to positive mood. CONCLUSIONS: By indicating that the presence of negative mood can hinder the revision of negative beliefs in clinically depressed people, our findings suggest that learning from new experiences can be hampered if state negative mood is activated. Thus, interventions relying on learning from novel positive experiences should aim at reducing state negative mood in depression.

Using expectation violation models to improve the outcome of psychological treatments.

Expectations are a central maintaining mechanism in mental disorders and most psychological treatments aim to directly or indirectly modify clinically relevant expectations. Therefore, it is crucial to examine why patients with mental disorders maintain dysfunctional expectations, even in light of disconfirming evidence, and how expectation-violating situations should be created in treatment settings to optimize treatment outcome and reduce the risk of treatment failures. The different psychological subdisciplines offer various approaches for understanding the underlying mechanisms of expectation development, persistence, and change. Here, we convey recommendations on how to improve psychological treatments by considering these different perspectives. Based on our expectation violation model, we argue that the outcome of expectation violation depends on several characteristics: features of the expectation-violating situation; the dynamics between the magnitude of expectation violation and cognitive immunization processes; dealing with uncertainties during and after expectation change; controlled and automatic attention processes; and the costs of expectation changes. Personality factors further add to predict outcomes and may offer a basis for personalized treatment planning. We conclude with a list of recommendations derived from basic psychology that could contribute to improved treatment outcome and to reduced risks of treatment failures.

Oversampled and undersolved: Depressive rumination from an active inference perspective.

Rumination is a widely recognized cognitive deviation in depression. Despite the recognition, researchers have struggled to explain why patients cannot disengage from the process, although it depresses their mood and fails to lead to effective problem-solving. We rethink rumination as repetitive but unsuccessful problem-solving attempts. Appealing to an active inference account, we suggest that adaptive problem-solving is based on the generation, evaluation, and performance of candidate policies that increase an organism's knowledge of its environment. We argue that the problem-solving process is distorted during rumination. Specifically, rumination is understood as engaging in excessive yet unsuccessful oversampling of policy candidates that do not resolve uncertainty. Because candidates are sampled from policies that were selected in states resembling one's current state, "bad" starting points (e.g., depressed mood, physical inactivity) make the problem-solving process vulnerable for generating a ruminative "halting problem". This problem leads to high opportunity costs, learned helplessness and diminished overt behavior. Besides reviewing evidence for the conceptual paths of this model, we discuss its neurophysiological correlates and point towards clinical implications.

How social rejection expectations and depressive symptoms bi-directionally predict each other - A cross-lagged panel analysis.

BACKGROUND: Although research suggests that social rejection expectations play a crucial role in the development and maintenance of depressive symptoms, it is not clear whether such expectations are a risk factor for depression or rather a consequence thereof. The present study addressed this issue by investigating the time-lagged bi-directional effects of social rejection expectations and depressive symptoms. METHODS: In an online survey, participants (N = 347) completed measures of social rejection expectations, depressive symptoms, interpersonal competencies, and perceived social support at baseline and 2 months later. The relationships between the variables were examined using path models and cross-lagged path analyses. RESULTS: Cross-lagged path analyses provided evidence for a substantial positive effect of social rejection expectations at baseline on depressive symptoms at follow-up in addition to the reverse effect. A mediator analysis indicated that neither interpersonal competencies nor perceived social support mediated these bi-directional effects. CONCLUSION: The current results demonstrate that social rejection expectations and depressive symptoms bi-directionally predict each other. Thus, social rejection expectations appears to be both a risk factor for - and a symptom of - depression. In order to prevent a vicious circle of social rejection expectations and depressive symptoms, we recommend the early detection and treatment of social rejection expectations. Moreover, social rejection expectations should be specifically addressed in cognitive-behavioural treatment of depression.

Cognitive and emotional variables predicting treatment outcome of cognitive behavior therapies for patients with medically unexplained symptoms: A meta-analysis.

OBJECTIVE: Cognitive behavior therapy (CBT) is the best-evaluated psychological approach to treat patients with medically unexplained symptoms (MUS). We still need a better understanding of what characterizes patients with MUS who benefit more or less from CBT. This systematic review aimed to identify patients' cognitive-emotional characteristics predicting the outcome of CBT for MUS. METHODS: A systematic literature search (PubMed, PsycINFO, Web of Science) revealed 37 eligible studies, 23 of these provided data for meta-analyses. Mean correlation coefficients between predictor variables and the outcomes (symptom intensity, physical or social-emotional functioning) were calculated using a random-effects model. Differences between syndromes of MUS were investigated with moderator analyses. RESULTS: Meta-analyses showed that patients with a comorbid mood disorder (r = 0.32, p < .01) or anxiety disorder (r = 0.18, p < .01), symptom catastrophizing and worries (r = 0.34, p < .01), tendencies of somatosensory amplification (r = 0.46, p = .04), and low symptom acceptance or self-efficacy (r = 0.25, p < .01) have a less favorable CBT outcome. Moderator analyses revealed that these associations between predictors and treatment outcome are pronounced in patients with chronic fatigue syndrome and irritable bowel syndrome. CONCLUSIONS: Our results show that pre-treatment differences in patients' cognitive-emotional characteristics predict patients' outcome in CBT. Patient-tailored CBT could be a promising approach to address MUS patients' widely varying needs more effectively. PROTOCOL REGISTRATION: The protocol of this systematic review and meta-analysis was registered in the PROSPERO registry (CRD 42018098649).

No1LikesU! - A Pilot Study on an Ecologically Valid and Highly Standardised Experimental Paradigm to Investigate Social Rejection Expectations and Their Modification.

Background: Dysfunctional expectations have been suggested as core features in the development and maintenance of mental disorders. Thus, preventing development and promoting modification of dysfunctional expectations through intervention might improve clinical treatment. While there are well-established experimental procedures to investigate the acquisition and modification of dysfunctional performance expectations in major depression, paradigms for investigating other important types of dysfunctional expectations (e.g. social rejection expectations) are currently lacking. We introduce an innovative associative learning paradigm, which can be used to investigate the development, maintenance, and modification of social rejection expectations. Method: A pilot sample of 28 healthy participants experienced manipulated social feedback after answering personal questions in supposed webcam conferences. While participants repeatedly received social rejection feedback in a first phase, differential feedback was given in a second phase (social rejection vs. social appreciation). In a third phase, explicit social feedback was omitted. Results: Participants developed social rejection expectations in the first phase. For the second phase, we found an interaction effect of experimental condition; i.e. participants adjusted their expectations according to the differential social feedback. In the third phase, learned social expectations remained stable in accordance to the social feedback in the second phase. Conclusion: Results indicate that the paradigm can be used to investigate the development, maintenance, and modification of social rejection expectations in healthy participants. This offers broad applications to explore the differential acquisition and modification of social rejection expectations in healthy vs. clinical samples. Further, the paradigm might be used to investigate therapeutic strategies to facilitate expectation change.

Belief updating in depression is not related to increased sensitivity to unexpectedly negative information.

BACKGROUND: People with depression differ from healthy people in the extent to which they use novel positive information to adjust negative expectations. In this study, we examined whether the two groups also differ in updating positive expectations after receiving unexpectedly negative information. METHODS: Examining 76 people with depressive symptoms and 81 healthy controls, we used an adapted version of a previously validated paradigm. After the initial establishment of positive performance expectations, participants worked on the TEMINT performance test, whereupon half of the participants received standardized feedback that confirmed previous positive expectations while the other half received disconfirming negative feedback. Subsequently, participants' performance expectations were assessed again. Additionally, we assessed participants' appraisal of the feedback, particularly whether they tended to disregard it. RESULTS: Results indicated that healthy subjects had overall more positive expectations than people with depressive symptoms, but the two samples did not differ in updating their expectations: both groups changed their expectations in a negative direction after receiving negative feedback; similarly, there were no differences between the two samples after receiving confirmatory positive feedback. Both people with and without depressive symptoms were more likely to disregard the feedback received if the feedback was negative, and such a negative appraisal of the feedback was associated with smaller expectation update. CONCLUSIONS: In combination with prior work, the current findings suggest that people with depressive symptoms do not over-sensitively react to unexpectedly negative information; rather, the main problem of depression seems to be the integration of novel positive information, as shown previously.

De novo design of heat-repressible RNA thermosensors in E. coli.

RNA-based temperature sensing is common in bacteria that live in fluctuating environments. Most naturally-occurring RNA thermosensors are heat-inducible, have long sequences, and function by sequestering the ribosome binding site in a hairpin structure at lower temperatures. Here, we demonstrate the de novo design of short, heat-repressible RNA thermosensors. These thermosensors contain a cleavage site for RNase E, an enzyme native to Escherichia coli and many other organisms, in the 5' untranslated region of the target gene. At low temperatures, the cleavage site is sequestered in a stem-loop, and gene expression is unobstructed. At high temperatures, the stem-loop unfolds, allowing for mRNA degradation and turning off expression. We demonstrated that these thermosensors respond specifically to temperature and provided experimental support for the central role of RNase E in the mechanism. We also demonstrated the modularity of these RNA thermosensors by constructing a three-input composite circuit that utilizes transcriptional, post-transcriptional, and post-translational regulation. A thorough analysis of the 24 thermosensors allowed for the development of design guidelines for systematic construction of similar thermosensors in future applications. These short, modular RNA thermosensors can be applied to the construction of complex genetic circuits, facilitating rational reprogramming of cellular processes for synthetic biology applications.