Targeting ROS production through inhibition of NADPH oxidases.

NADPH oxidases (NOXs) are transmembrane enzymes that are devoted to the production of reactive oxygen species (ROS). In cancers, dysregulation of NOX enzymes affects ROS production, leading to redox unbalance and tumor progression. Consequently, NOXs are a drug target for cancer therapeutics, although current therapies have off-target effects: there is a need for isoenzyme-selective inhibitors. Here, we describe fully validated human NOX inhibitors, obtained from an in silico screen, targeting the active site of Cylindrospermum stagnale NOX5 (csNOX5). The hits are validated by in vitro and in cellulo enzymatic and binding assays, and their binding modes to the dehydrogenase domain of csNOX5 studied via high-resolution crystal structures. A high-throughput screen in a panel of cancer cells shows activity in selected cancer cell lines and synergistic effects with KRAS modulators. Our work lays the foundation for the development of inhibitor-based methods for controlling the tightly regulated and highly localized ROS sources.

Phenotypes favoring fractional exhaled nitric oxide discordance vs guideline-based uncontrolled asthma.

BACKGROUND: Despite potential value of identification of allergic inflammation with fractional exhaled nitric oxide (FeNO) in managing asthma, randomized clinical trials have not consistently shown better outcomes compared with guideline management alone. OBJECTIVE: To assess the effectiveness of FeNO vs non-FeNO-based therapeutic algorithms in managing asthma, and the phenotypic profile associated with FeNO >35 ppb yet well controlled by guidelines, as a potential model to predict better FeNO-based algorithm outcomes. METHODS: This is a randomized controlled study (RCT) in 88 high-risk children with asthma 7 to 18 years of age across 352 visits over a 1-year period. Generalized estimating equations analysis assessed algorithm group differences in outcomes and characteristics associated with higher odds uncontrolled by FeNO alone in the treatment decision algorithm. RESULTS: The FeNO treatment algorithm did not show superiority in reducing exacerbations and morbidity (P > .05). Phenotypes that more than doubled the odds FeNO alone identified uncontrolled asthma included adolescence, non-adherence, high atopy (>6+), and baseline FeNO >35 ppb, whereas obesity, FEF25-75% < 65% predicted, and bronchodilator response >10% decreased the odds. Uncontrolled asthma by FeNO alone (F) vs guidelines alone (G) showed overall F/G > 1.0 in adolescents, but <1.0 in younger patients unless the FeNO threshold was reduced to >20 ppb. CONCLUSION: Our study suggests that age and phenotypes play a key role in FeNO discordance compared with the conventional guideline-based uncontrolled asthma. The FeNO-based therapeutic algorithm, if confirmed further, could provide the clinician with an effective asthma management tool. The clinical implication could improve future FeNO-based RCTs and treatment decision algorithms in managing asthma by considering phenotypes and age-dependent FeNO thresholds.

Initial assessment of medical post-traumatic stress among patients with chronic esophageal diseases.

BACKGROUND: Diagnosis and management of chronic esophageal disease requires the use of potentially traumatic medical procedures, performed with or without sedation. Medical trauma and post-traumatic stress (PTS) are emerging as important considerations in patients with digestive illness. To date, no study assesses medical PTS from procedures in patients with esophageal disease. METHODS: Adult patients with achalasia, eosinophilic esophagitis, gastroesophageal reflux disease, or functional esophageal disease at a university-based gastroenterology clinic completed: Post-Traumatic Stress Disorder Checklist for DSM-5 (PCL-5), Gastroesophageal Disease Questionnaire, Brief Esophageal Dysphagia Questionnaire, Northwestern Esophageal Quality of Life scale (HRQoL), NIH-PROMIS Depression scale, and a study-specific questionnaire about esophageal procedures (endoscopy with sedation; functional lumen imaging probe (FLIP) with sedation; high-resolution manometry (HRM); wireless pH testing; or 24-h pH-impedance testing). KEY RESULTS: Half of 149 participants reported at least one traumatic procedure, with HRM most often cited. Only 2.7% met the cutoff for PTS on PCL-5. This increased to 7.1% for patients with a traumatic procedure combined with experiencing intense fear. Rates of moderate-severe PTS ranged from 7.4%-12% for all patients and 14%-29% for those with a traumatic procedure with fear. Medical PTS was associated with poorer HRQoL, and increased esophageal symptoms, depression, and hypervigilance and symptom anxiety. CONCLUSIONS & INFERENCES: Preliminary evidence suggests medical PTS affects few patients with esophageal disease. However traumatic procedures, most often associated with HRM, significantly increase PTS symptoms. The potential impacts of medical PTS on esophageal patient assessment and outcomes are considerable and warrants further study.

Bioisosteric replacement based on 1,2,4-oxadiazoles in the discovery of 1H-indazole-bearing neuroprotective MAO B inhibitors.

Following a hybridization strategy, a series of 5-substituted-1H-indazoles were designed and evaluated in vitro as inhibitors of human monoamine oxidase (hMAO) A and B. Among structural modifications, the bioisostere-based introduction of 1,2,4-oxadiazole ring returned the most potent and selective human MAO B inhibitor (compound 20, IC50 = 52 nM, SI > 192). The most promising inhibitors were studied in cell-based neuroprotection models of SH-SY5Y and astrocytes line against H2O2. Moreover, preliminary drug-like features (aqueous solubility at pH 7.4; hydrolytic stability at acidic and neutral pH) were assessed for selected 1,2,4-oxadiazoles and compared to amide analogues through RP-HPLC methods. Molecular docking simulations highlighted the crucial role of molecular flexibility in providing a better shape complementarity for compound 20 within MAO B enzymatic cleft than rigid analogue 18. Enzymatic kinetics analysis along with thermal stability curves (Tm shift = +2.9 °C) provided clues of a tight-binding mechanism for hMAO B inhibition by 20.

Magnet-associated intestinal perforation results in a new institutional policy of ferromagnetic screening prior to MRI.

Foreign body ingestions are common and the vast majority pass through the gastrointestinal tract without complication. Some ingestions, however, result in serious morbidity and mortality. We present a case in which the patient's chief complaint of severe posterior neck pain was unrelated to his foreign body ingestion (multiple magnets). The ingestion of magnets was not disclosed by the child to either the providing medical team or to the patient's family. In order to evaluate the patient's complaint of severe focal neck pain, MRI of the neck was performed. The authors believe it to be feasible that the MRI scan resulted in intestinal perforations that might not have occurred during the natural course of the ingestion. This complication might have been prevented if the patient had undergone screening with a ferromagnetic detector prior to entering the MRI suite. Because of the serious complications related to this case, all pediatric patients at our institution are now screened with ferromagnetic detectors prior to entering the MRI suite. We encourage nationwide policy revision to prevent further incidents similar to the one described in this case.

Psychosocial Aspects of Metabolic and Bariatric Surgeries and Endoscopic Therapies.

Obesity is a prevalent progressive and relapsing disease for which there are several levels of intervention, including metabolic and bariatric surgery (MBS) and now endoscopic bariatric and metabolic therapies (EBMTs). Preoperative psychological assessment focused on cognitive status, psychiatric symptoms, eating disorders, social support, and substance use is useful in optimizing patient outcomes and minimizing risks in MBS. Very little is known about the psychosocial needs of patients seeking EBMTs, though these investigations will be forthcoming if these therapies become more widespread. As MBS and EBMT inherently alter the gastrointestinal (GI) tract, considerations for the longer-term GI functioning of the patient are relevant and should be considered and monitored.

Evolutionary and structural analyses of the NADPH oxidase family in eukaryotes reveal an initial calcium dependency.

Reactive oxygen species are unstable molecules generated by the partial reduction of dioxygen. NADPH oxidases are a ubiquitous family of enzymes devoted to ROS production. They fuel an array of physiological roles in different species and are chemically demanding enzymes requiring FAD, NADPH and heme prosthetic groups in addition to either calcium or a various number of cytosolic mediators for activity. These activating partners are exclusive components that partition and distinguish the NOX members from one another. To gain insight into the evolution of these activating mechanisms, and in general in their evolutionary history, we conducted an in-depth phylogenetic analysis of the NADPH oxidase family in eukaryotes. We show that all characterized NOXs share a common ancestor, which comprised a fully formed catalytic unit. Regarding the activation mode, we identified calcium-dependency as the earliest form of NOX regulation. The protein-protein mode of regulation would have evolved more recently by gene-duplication with the concomitant loss of the EF-hands motif region. These more recent events generated the diversely activated NOX systems as observed in extant animals and fungi.

Using factorial mediation analysis to better understand the effects of interventions.

To improve understanding of how interventions work or why they do not work, there is need for methods of testing hypotheses about the causal mechanisms underlying the individual and combined effects of the components that make up interventions. Factorial mediation analysis, i.e., mediation analysis applied to data from a factorial optimization trial, enables testing such hypotheses. In this commentary, we demonstrate how factorial mediation analysis can contribute detailed information about an intervention's causal mechanisms. We briefly review the multiphase optimization strategy (MOST) and the factorial experiment. We use an empirical example from a 25 factorial optimization trial to demonstrate how factorial mediation analysis opens possibilities for better understanding the individual and combined effects of intervention components. Factorial mediation analysis has important potential to advance theory about interventions and to inform intervention improvements.

Obesity Status on associations between cancer-related beliefs and health behaviors in cancer survivors: Implications for patient-clinician communication.

OBJECTIVE: Associations between cancer beliefs and health behavior engagement are largely unexplored in cancer survivors, particularly among those with overweight and obesity. We investigated belief-behavior associations for cancer survivors, and whether obesity altered these associations. METHODS: Cancer survivors were identified from the National Cancer Institute HINTS Survey 5 data and classified as having had an obesity-related cancer or not. Linear and multiple logistic regression analyses examined whether cancer risk beliefs and self-efficacy predicted dining out behaviors and physical activity (PA). Restricted analyses were conducted in those with overweight or obesity. RESULTS: Low self-efficacy to take care of one's health was associated with longer sitting time in the overall sample (p = 0.04). In cancer survivors with overweight or obesity, engagement in healthier behaviors was associated with 1) feeling less overwhelmed by cancer risk recommendations and 2) believing that PA or obesity influences cancer development (both p < 0.05). Among those with overweight and obesity, associations between cancer beliefs and health behaviors were not significantly different by cancer type (obesity-related vs. not). CONCLUSIONS: Obesity altered associations between cancer risk beliefs and health behavior engagement from the overall sample. PRACTICE IMPLICATIONS: Weight status may be a useful tailoring factor when delivering health-promoting interventions for cancer survivors.

Using the Preparation Phase of the Multiphase Optimization Strategy to Develop a Messaging Component for Weight Loss: Formative and Pilot Research.

BACKGROUND: Mobile messaging is often used in behavioral weight loss interventions, yet little is known as to the extent to which they contribute to weight loss when part of a multicomponent treatment package. The multiphase optimization strategy (MOST) is a framework that researchers can use to systematically investigate interventions that achieve desirable outcomes given specified constraints. OBJECTIVE: This study describes the use of MOST to develop a messaging intervention as a component to test as part of a weight loss treatment package in a subsequent optimization trial. METHODS: On the basis of our conceptual model, a text message intervention was created to support self-regulation of weight-related behaviors. We tested the messages in the ENLIGHTEN feasibility pilot study. Adults with overweight and obesity were recruited to participate in an 8-week weight loss program. Participants received a commercially available self-monitoring smartphone app, coaching calls, and text messages. The number and frequency of text messages sent were determined by individual preferences, and weight was assessed at 8 weeks. RESULTS: Participants (n=9) in the feasibility pilot study lost 3.2% of their initial body weight over the 8-week intervention and preferred to receive 1.8 texts per day for 4.3 days per week. Researcher burden in manually sending messages was high, and the cost of receiving text messages was a concern. Therefore, a fully automated push notification system was developed to facilitate sending tailored daily messages to participants to support weight loss. CONCLUSIONS: Following the completion of specifying the conceptual model and the feasibility pilot study, the message intervention went through a final iteration. Theory and feasibility pilot study results during the preparation phase informed critical decisions about automation, frequency, triggers, and content before inclusion as a treatment component in a factorial optimization trial. TRIAL REGISTRATION: ClinicalTrials.gov NCT01814072; https://clinicaltrials.gov/ct2/show/NCT01814072.

A closer look into NADPH oxidase inhibitors: Validation and insight into their mechanism of action.

NADPH-oxidases (NOXs) purposefully produce reactive-oxygen-species (ROS) and are found in most kingdoms of life. The seven human NOXs are each characterized by a specific expression profile and a fine regulation to spatio-temporally tune ROS concentration in cells and tissues. One of the best known roles for NOXs is in host protection against pathogens but ROS themselves are important second messengers involved in tissue regeneration and the modulation of pathways that induce and sustain cell proliferation. As such, NOXs are attractive pharmacological targets in immunomodulation, fibrosis and cancer. We have studied an extensive number of available NOX inhibitors, with the specific aim to identify bona fide ligands versus ROS-scavenging molecules. Accordingly, we have established a comprehensive platform of biochemical and biophysical assays. Most of the investigated small molecules revealed ROS-scavenging and/or assay-interfering properties to various degrees. A few compounds, however, were also demonstrated to directly engage one or more NOX enzymes. Diphenylene iodonium was found to react with the NOXs' flavin and heme prosthetic groups to form stable adducts. We also discovered that two compounds, VAS2870 and VAS3947, inhibit NOXs through the covalent alkylation of a cysteine residue. Importantly, the amino acid involved in covalent binding was found to reside in the dehydrogenase domain, where the nicotinamide ring of NADPH is bound. This work can serve as a springboard to guide further development of bona fide ligands with either agonistic or antagonistic properties toward NOXs.

A Factorial Experiment to Optimize Remotely Delivered Behavioral Treatment for Obesity: Results of the Opt-IN Study.

OBJECTIVE: Intensive behavioral obesity treatments face scalability challenges, but evidence is lacking about which treatment components could be cut back without reducing weight loss. The Optimization of Remotely Delivered Intensive Lifestyle Treatment for Obesity (Opt-IN) study applied the Multiphase Optimization Strategy to develop an entirely remotely delivered, technology-supported weight-loss package to maximize the amount of weight loss attainable for ≤$500. METHODS: Six-month weight loss was examined among adults (N = 562) with BMI ≥ 25 who were randomly assigned to conditions in a factorial experiment crossing five dichotomous treatment components set to either low/high (12 vs. 24 coaching calls) or off/on (primary care provider reports, text messaging, meal replacements, and buddy training). RESULTS: About 84.3% of participants completed the final assessment. The treatment package yielding maximum weight loss for ≤$500 included 12 coaching calls, buddy training, and primary care provider progress reports; produced average weight loss of 6.1 kg, with 57.1% losing ≥5% and 51.8% losing ≥7%; and cost $427 per person. The most expensive candidate-treatment component (24 vs. 12 coaching calls) was screened out of the optimized treatment package because it did not increase weight loss. CONCLUSIONS: Systematically testing each treatment component's effect on weight loss made it possible to eliminate more expensive but less impactful components, yielding an optimized, resource-efficient obesity treatment for evaluation in a randomized controlled trial.

Effective Care Management for Children With Special Health Care Needs in the Era of Value-Based Payment.

Objectives. To evaluate the impact of a pediatric-specific care coordination program for Medicaid children with special health care needs under a fully capitated payment model and assess whether sufficient savings can be achieved to offset the cost of the care coordination program. Methods. 442 children with special health care needs, receiving health care under a Medicaid capitation payment program, were enrolled in a care coordination program. ED and inpatient utilization were measured for 1-year pre and post intervention. Use rates and costs for ED and inpatient services were evaluated using a Poisson random effect regression model. Results. There was a statistically significant reduction in ED utilization (31% reduction, P < .0001), inpatient admissions (38% reduction, P = .0002), and inpatient length of stay (34% reduction, P = .0112) comparing the pre and post intervention periods. Medical cost savings attributed to the reduction in ED and inpatient utilization was approximately 3 times the program costs. Conclusions. Enrolling children with special health care needs in a care management program was associated with a significant reduction in ED utilization, inpatient admissions, and hospital length of stay when compared with baseline expenditures. Under a fully capitated Medicaid model, the cost savings greatly exceeded the costs of the interventions. These results serve to highlight the efficacy of pediatric-specific care management programs for children with special health care needs, both clinically and economically. Such models can inform other interventions and contracting strategies to assure children receive the care they deserve in a sustainable cost model.

Toward a precision behavioral medicine approach to addressing high-risk sun exposure: a qualitative analysis.

OBJECTIVES: Precision behavioral medicine techniques integrating wearable ultraviolet radiation (UVR) sensors may help individuals avoid sun exposure that places them at-risk for skin cancer. As a preliminary step in our patient-centered process of developing a just-in-time adaptive intervention, this study evaluated reactions and preferences to UVR sensors among melanoma survivors. MATERIALS AND METHODS: Early stage adult melanoma survivors were recruited for a focus group (n = 11) or 10-day observational study, which included daily wearing a UVR sensor and sun exposure surveys (n = 39). Both the focus group moderator guide and observational study exit interviews included questions on UVR sensing as a potential intervention strategy. These responses were transcribed and coded using an inductive strategy. RESULTS: Most observational study participants (84.6%) said they would find information provided by UVR sensors to be useful to help them learn about how specific conditions (eg, clouds, location) impact sun exposure and provide in-the-moment alerts. Focus group participants expressed enthusiasm for UVR information and identified preferred qualities of a UVR sensor, such as small size and integration with other devices. Participants in both studies indicated concern that UVR feedback may be difficult to interpret and some expressed that a UVR sensor may not be convenient or desirable to wear in daily life. DISCUSSION: Melanoma survivors believe that personalized UVR exposure information could improve their sun protection and want this information delivered in a method that is meaningful and actionable. CONCLUSION: UVR sensing is a promising component of a precision behavioral medicine strategy to reduce skin cancer risk.

A Tail-Based Mechanism Drives Nucleosome Demethylation by the LSD2/NPAC Multimeric Complex.

LSD1 and LSD2 are homologous histone demethylases with opposite biological outcomes related to chromatin silencing and transcription elongation, respectively. Unlike LSD1, LSD2 nucleosome-demethylase activity relies on a specific linker peptide from the multidomain protein NPAC. We used single-particle cryoelectron microscopy (cryo-EM), in combination with kinetic and mutational analysis, to analyze the mechanisms underlying the function of the human LSD2/NPAC-linker/nucleosome complex. Weak interactions between LSD2 and DNA enable multiple binding modes for the association of the demethylase to the nucleosome. The demethylase thereby captures mono- and dimethyl Lys4 of the H3 tail to afford histone demethylation. Our studies also establish that the dehydrogenase domain of NPAC serves as a catalytically inert oligomerization module. While LSD1/CoREST forms a nucleosome docking platform at silenced gene promoters, LSD2/NPAC is a multifunctional enzyme complex with flexible linkers, tailored for rapid chromatin modification, in conjunction with the advance of the RNA polymerase on actively transcribed genes.

Development of alkyl glycerone phosphate synthase inhibitors: Structure-activity relationship and effects on ether lipids and epithelial-mesenchymal transition in cancer cells.

In aggressive tumors, alkylglyceronephosphate synthase (AGPS) controls cellular ether phospholipid utilization and metabolism to promote cancer cell proliferation and motility. SAR studies on the first-in-class AGPS inhibitor 1, discovered by our group, led to the 2,6-difluoro analog 2i which showed higher binding affinity than 1in vitro. In 231MFP cancer cells, 2i reduced ether lipids levels and cell migration rate. When tested in PC-3 and MDA-MB-231 cancer cells, 2i specifically impaired epithelial to mesenchymal transition (EMT) by modulating E-cadherin, Snail and MMP2 expression levels. Moreover, the combination of siRNAs against AGPS and 2i provided no additive effect, confirming that the modulation of 2i on EMT specifically relies on AGPS inhibition. Finally, this compound also affected cancer cell proliferation especially in MDA-MB-231 cells expressing higher AGPS level, whereas it provided negligible effects on MeT5A, a non-tumorigenic cell line, thus showing cancer specificity.

DNA-immunisation with dengue virus E protein domains I/II, but not domain III, enhances Zika, West Nile and Yellow Fever virus infection.

Dengue virus (DENV), the causative agent of dengue disease, is among the most important mosquito-borne pathogens worldwide. DENV is composed of four closely related serotypes and belongs to the Flaviviridae family alongside other important arthropod-borne viral pathogens such as Zika virus (ZIKV), West Nile virus (WNV) and Yellow Fever virus (YFV). After infection, the antibody response is mostly directed to the viral E glycoprotein which is composed of three structural domains named DI, DII and DIII that share variable degrees of homology among different viruses. Recent evidence supports a close serological interaction between ZIKV and DENV. The possibility of worse clinical outcomes as a consequence of antibody-dependent enhancement of infection (ADE) due to cross-reactive antibodies with poor neutralisation activity is a matter of concern. We tested polyclonal sera from groups of female Balb/C mice vaccinated with DNA constructs expressing DI/DII, DIII or the whole sE from different DENV serotypes and compared their activity in terms of cross-reactivity, neutralisation of virus infection and ADE. Our results indicate that the polyclonal antibody responses against the whole sE protein are highly cross-reactive with strong ADE and poor neutralisation activities due to DI/DII immunodominance. Conversely, anti-DIII polyclonal antibodies are type-specific, with no ADE towards ZIKV, WNV and YFV, and strong neutralisation activity restricted only to DENV.