Robust Strategy for Hit-to-Lead Discovery: NMR for SAR.

Establishing robust structure-activity relationships (SARs) is key to successful drug discovery campaigns, yet it often remains elusive due to screening and hit validation artifacts (false positives and false negatives), which frequently result in unproductive downstream expenditures of time and resources. To address this issue, we developed an integrative biophysics-driven strategy that expedites hit-to-lead discovery, mitigates false positives/negatives and common hit validation errors, and provides a robust approach to obtaining accurate binding and affinity measurements. The advantage of this method is that it vastly improves the clarity and reproducibility for affinity-driven SAR by monitoring and eliminating confounding factors. We demonstrate the ease at which high-quality micromolar binders can be generated from the initial millimolar fragment screening hits against an "undruggable" protein target, HRas.

Schedule Thinning Following Functional Communication Training: Effects of Chained and Multiple Schedules.

Functional communication training (FCT) is used to reduce rates of problem behavior by teaching communicative responses that access functionally equivalent reinforcers. During FCT, the communicative response is typically placed on a dense schedule of reinforcement that is unlikely to be maintained in the natural environment. Experiment 1 evaluated the effects of two schedule-thinning procedures (chained schedules and multiple schedules) on problem behavior maintained by escape from demands for three participants following FCT. The chained and multiple-schedule procedures were effective in reducing rates of problem behavior. Compliance increased under both schedules, but the chained schedule resulted in higher levels of compliance with two participants. In Experiment 2, participants' preference for the chained or multiple-schedule procedure was evaluated using a modified concurrent-chain procedure. One participant preferred the chained schedule. One participant preferred the multiple schedule. One participant did not appear to discriminate between conditions.

Management of ponatinib dosing in chronic myeloid leukemia patients.

Ponatinib (Iclusig®, Takeda/Incyte) is a third-generation highly-potent-pan-inhibitor of tyrosine kinases, active in all single resistance ABL kinase mutations including the T315l mutation. It is approved to treat of chronic myeloid leukemia (CML) in every phase of disease-resistant or intolerant to second-generation tyrosine kinase inhibitors (2GTKIs) and for whom imatinib is not clinically appropriate as well as for patients with T315I mutation. The drug is also indicated for Ph+ acute lymphoblastic leukemia (ALL). The approved starting dose for ponatinib is 45 mg once daily. Available data revealed ponatinib dose-dependent increased risk of cardiovascular toxicity. There is still no consensus about the optimal, initial dose of ponatinib and its management during therapy. It is crucial to start treatment with the risk-adjusted dose and assess the benefit-risk profile of ponatinib dosing. Evaluation of dosing modification should be considered during treatment, mainly if toxicity occurs. Our study summarizes current knowledge and recommendations about the choice of starting dose of ponatinib and management of ponatinib dosing during the treatment.

Probing the free-state solution behavior of drugs and their tendencies to self-aggregate into nano-entities.

The free-state solution behaviors of drugs profoundly affect their properties. Therefore, it is critical to properly evaluate a drug's unique multiphase equilibrium when in an aqueous enviroment, which can comprise lone molecules, self-associating aggregate states and solid phases. To date, the full range of nano-entities that drugs can adopt has been a largely unexplored phenomenon. This protocol describes how to monitor the solution behavior of drugs, revealing the nano-entities formed as a result of self-associations. The procedure begins with a simple NMR 1H assay, and depending on the observations, subsequent NMR dilution, NMR T2-CPMG (spin-spin relaxation Carr-Purcell-Meiboom-Gill) and NMR detergent assays are used to distinguish between the existence of fast-tumbling lone drug molecules, small drug aggregates and slow-tumbling colloids. Three orthogonal techniques (dynamic light scattering, transmission electron microscopy and confocal laser scanning microscopy) are also described that can be used to further characterize any large colloids. The protocol can take a non-specialist between minutes to a few hours; thus, libraries of compounds can be evaluated within days.

Interventions for inappropriate sexual behavior in individuals with intellectual and developmental disabilities: A brief review.

Although prevalence rates vary, 6% to 28% of individuals with intellectual or developmental disabilities (IDDs) engage in inappropriate sexual behavior (ISB), ranging from public masturbation to sexually aggressive behavior. Along with increased risk for contacting the criminal justice system, people with IDDs who display ISB may encounter negative social consequences, restricted community access and barriers to independence, and a variety of counter-therapeutic outcomes. The purpose of the present review is to highlight recent, efficacious behavior-analytic treatments for ISB in individuals with IDDs. Ethical considerations and areas for future research will be discussed.

Anesthetic Management During Pediatric Cytoreductive Surgery and Hyperthermic Intraperitoneal Chemotherapy With Cisplatin in a Small Child: A Case Report and Systematic Literature Review.

Cytoreductive surgery (CS) and hyperthermic intraperitoneal chemotherapy (HIPEC) present a challenging task for anesthesia providers. Anesthesia management may be complicated by hyperthermia, fluid shifts, and distinct inflammatory response. Only a few reports dealing with the anesthesia management of pediatric CS and HIPEC have been published. We report a case of a 2-year-old child with a relapse of an alveolar rhabdomyosarcoma of the uterus and peritoneal carcinomatosis treated with CS and HIPEC. For children, careful temperature measurement, intraoperative prevention of hyperthermia, and sufficient volume management are important, as well as postoperative pediatric intensive care with experience CS and HIPEC patients.

Correction: Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Blue Light Covers Increase Stereotypy and Decrease On-Task Behavior for Students with Autism.

Some recommended strategies for individuals with autism spectrum disorder (ASD) are not empirically based. The purpose of the study was to evaluate effects of blue light covers on levels of stereotypy and on-task behavior. Four male children with ASD who engaged in repetitive behavior participated. Placing light covers over the classroom's fluorescent lights relative to normal classroom lighting did not improve on-task behavior or stereotypy.

Exposing Small-Molecule Nanoentities by a Nuclear Magnetic Resonance Relaxation Assay.

Small molecules can self-assemble in aqueous solution into a wide range of nanoentity types and sizes (dimers, n-mers, micelles, colloids, etc.), each having their own unique properties. This has important consequences in the context of drug discovery including issues related to nonspecific binding, off-target effects, and false positives and negatives. Here, we demonstrate the use of the spin-spin relaxation Carr-Purcell-Meiboom-Gill NMR experiment, which is sensitive to molecular tumbling rates and can expose larger aggregate species that have slower rotational correlations. The strategy easily distinguishes lone-tumbling molecules versus nanoentities of various sizes. The technique is highly sensitive to chemical exchange between single-molecule and aggregate states and can therefore be used as a reporter when direct measurement of aggregates is not possible by NMR. Interestingly, we found differences in solution behavior for compounds within structurally related series, demonstrating structure-nanoentity relationships. This practical experiment is a valuable tool to support drug discovery efforts.

LMA Protector™ Airway: first experience with a new second generation laryngeal mask.

BACKGROUND: The LMA Protector™ Airway (The Laryngeal Mask Company Ltd., Teleflex Incorporated, Athlone, Ireland) is a new supraglottic airway promising a better seal, an improved drainage of gastric secretions and the opportunity of a simplified fiberscopy-guided tracheal intubation. The aim of this study was to present a primary evaluation of the LMA Protector in a clinical setting. METHODS: After informed consent 50 patients, scheduled for minor/moderate surgery in supine position, were recruited. Pharyngeal seal pressures were examined in neutral position of the patients' head and in maximum passive extension of the neck. Additionally, the fiberscopic view on the glottis was graduated and the feasibility of fiberscope guided tracheal intubation through the device was evaluated. RESULTS: The median pharyngeal seal pressure of the LMA Protector in neutral position of the head was 34 cmH2O. Passive extension of the neck did not cause a reduction of the pharyngeal seal (median pharyngeal seal pressure: 34.7 cmH2O; P<0.039). The LMA Protector was applicable for fiberscopic tracheal intubation but is not reliable for blind tracheal intubation. CONCLUSIONS: The LMA Protector provides a high pharyngeal seal. Uncommon for laryngeal masks its pharyngeal seal is not affected by the extension of the patient's neck. As a second generation supraglottic airway which is also suitable for simplified fiberscopic guided tracheal intubation, the LMA Protector could be considered as a supraglottic airway of the third generation.

Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph + ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1.

Minimal residual disease (MRD) is a powerful prognostic factor in acute lymphoblastic leukemia (ALL) and is used for patient stratification and treatment decisions, but its precise role in Philadelphia chromosome positive ALL is less clear. This uncertainty results largely from methodological differences relating to the use of real-time quantitative PCR (qRT-PCR) to measure BCR-ABL1 transcript levels for MRD analysis. We here describe the first results by the EURO-MRD consortium on standardization of qRT-PCR for the e1a2 BCR-ABL1 transcript in Ph + ALL, designed to overcome the lack of standardisation of laboratory procedures and data interpretation. Standardised use of EAC primer/probe sets and of centrally prepared plasmid standards had the greatest impact on reducing interlaboratory variability. In QC1 the proportion of analyses with BCR-ABL1/ABL1 ratios within half a log difference were 40/67 (60%) and 52/67 (78%) at 10-3 and 36/67 (53%) and 53/67 (79%) at 10-4BCR-ABL1/ABL1. Standardized RNA extraction, cDNA synthesis and cycler platforms did not improve results further, whereas stringent application of technical criteria for assay quality and uniform criteria for data interpretation and reporting were essential. We provide detailed laboratory recommendations for the standardized MRD analysis in routine diagnostic settings and in multicenter clinical trials for Ph + ALL.

Fecal elastase 1 and vitamin D3 in patients with osteoporotic bone fractures.

BACKGROUND AND AIMS: The aim of the present study was to clarify if patients with osteoporotic bone fractures have exocrine pancreatic insufficiency, especially reduced fecal elastase 1, connected with lowered serum levels of vitamin D3 that could be relevant for predominant osteoporosis. METHODS: Between October 1999 and September 2001, we investigated on 167 patients with an average age of approx. 69 years suffering from typical osteoporotic bone fractures, as well as 20 healthy controls with an average age of 53 years. A standardized osteodensitometry via dual energy X-ray absorptiometry (DEXA) was performed in all participants. Levels of PTH, 1,25(OH)(2) Vitamin D(3), 25(OH) Vitamin D(3), calcium and phosphate in serum, elastase 1 in feces as well as the body mass index were determined in all patients and controls. RESULTS: In patients 25(OH)D3 was more than 60% and 1,25(OH)(2)D(3) was more than 53% decreased compared to controls. Fecal elastase 1 was lower than the lowest reference of 200 microg/g feces in more than 34% of the patients and it was more than 65% reduced in comparison to healthy controls (fecal elastase 1 patients: 240.7 +/- 96.3 microg/g; controls 694.9 +/- 138.6 microg/g). Separation of the patients in accordance with the elastase 1 contend in feces into four groups (below 100 microg/g, between 100 and 200 microg/g, between 201 and 300 microg/g and above 300 microg/g) resulted in significant variations for 25(OH)D(3), 1,25(OH)(2)D(3), calcium and PTH between these groups (p < 0.01). Furthermore 25(OH)D(3), 1,25(OH)(2)D(3), calcium and PTH correlated significantly with elastase 1 in feces (p < 0.01) the way, that lower fecal elastase 1 was connected with lower levels of the other parameters. BMI shows no relevant differences within the patients or between patients and controls. CONCLUSION: Exocrine pancreatic insufficiency, especially lowered fecal elastase 1, may be much more frequent in patients with osteoporotic bone fractures than suggested so far. Lowered exocrine pancreatic function with lowered fecal elastase 1 seems to be relevant as a reason for reduced levels of circulating vitamin D3 metabolites being an appropriate additional cause for predominant osteoporosis.

Further evaluation of leisure items in the attention condition of functional analyses.

Research suggests that including leisure items in the attention condition of a functional analysis may produce engagement that masks sensitivity to attention. In this study, 4 individuals' initial functional analyses indicated that behavior was maintained by nonsocial variables (n = 3) or by attention (n = 1). A preference assessment was used to identify items for subsequent functional analyses. Four conditions were compared, attention with and without leisure items and control with and without leisure items. Following this, either high- or low-preference items were included in the attention condition. Problem behavior was more probable during the attention condition when no leisure items or low-preference items were included, and lower levels of problem behavior were observed during the attention condition when high-preference leisure items were included. These findings suggest how preferred items may hinder detection of behavioral function.

Effects of Particle Size and Surface Chemistry on the Dispersion of Graphite Nanoplates in Polypropylene Composites.

Carbon nanoparticles tend to form agglomerates with considerable cohesive strength, depending on particle morphology and chemistry, thus presenting different dispersion challenges. The present work studies the dispersion of three types of graphite nanoplates (GnP) with different flake sizes and bulk densities in a polypropylene melt, using a prototype extensional mixer under comparable hydrodynamic stresses. The nanoparticles were also chemically functionalized by covalent bonding polymer molecules to their surface, and the dispersion of the functionalized GnP was studied. The effects of stress relaxation on dispersion were also analyzed. Samples were removed along the mixer length, and characterized by microscopy and dielectric spectroscopy. A lower dispersion rate was observed for GnP with larger surface area and higher bulk density. Significant re-agglomeration was observed for all materials when the deformation rate was reduced. The polypropylene-functionalized GnP, characterized by increased compatibility with the polymer matrix, showed similar dispersion effects, albeit presenting slightly higher dispersion levels. All the composites exhibit dielectric behavior, however, the alternate current (AC) conductivity is systematically higher for the composites with larger flake GnP.

Mechanistic insights into allosteric regulation of the A2A adenosine G protein-coupled receptor by physiological cations.

Cations play key roles in regulating G-protein-coupled receptors (GPCRs), although their mechanisms are poorly understood. Here, 19F NMR is used to delineate the effects of cations on functional states of the adenosine A2A GPCR. While Na+ reinforces an inactive ensemble and a partial-agonist stabilized state, Ca2+ and Mg2+ shift the equilibrium toward active states. Positive allosteric effects of divalent cations are more pronounced with agonist and a G-protein-derived peptide. In cell membranes, divalent cations enhance both the affinity and fraction of the high affinity agonist-bound state. Molecular dynamics simulations suggest high concentrations of divalent cations bridge specific extracellular acidic residues, bringing TM5 and TM6 together at the extracellular surface and allosterically driving open the G-protein-binding cleft as shown by rigidity-transmission allostery theory. An understanding of cation allostery should enable the design of allosteric agents and enhance our understanding of GPCR regulation in the cellular milieu.

Treatment and outcome of 2904 CML patients from the EUTOS population-based registry.

The European Treatment and Outcome Study (EUTOS) population-based registry includes data of all adult patients newly diagnosed with Philadelphia chromosome-positive and/or BCR-ABL1+ chronic myeloid leukemia (CML) in 20 predefined countries and regions of Europe. Registration time ranged from 12 to 60 months between January 2008 and December 2013. Median age was 55 years and median observation time was 29 months. Eighty percent of patients were treated first line with imatinib, and 17% with a second-generation tyrosine kinase inhibitor, mostly according to European LeukemiaNet recommendations. After 12 months, complete cytogenetic remission (CCyR) and major molecular response (MMR) were achieved in 57% and 41% of patients, respectively. Patients with high EUTOS risk scores achieved CCyR and MMR significantly later than patients with low EUTOS risk. Probabilities of overall survival (OS) and progression-free survival for all patients at 12, 24 and 30 months was 97%, 94% and 92%, and 95%, 92% and 90%, respectively. The new EUTOS long-term survival score was validated: the OS of patients differed significantly between the three risk groups. The probability of dying in remission was 1% after 24 months. The current management of patients with tyrosine kinase inhibitors resulted in responses and outcomes in the range reported from clinical trials. These data from a large population-based, patient sample provide a solid benchmark for the evaluation of new treatment policies.

Effects of treatment-integrity failures on a response-cost procedure.

Effects of incorrect or partial implementation (poor treatment integrity) on response cost are largely unknown. We evaluated reduced treatment integrity during response cost on rates of 2 concurrently available responses. College students earned points by clicking on either a black circle or a red circle on a computer screen. Experiment 1 compared 2 types of treatment-integrity failures (omission and commission errors) across 2 levels of integrity (20% and 50%). Compared to 100% integrity conditions, omission errors did not suppress responding to the same extent, and commission errors reduced target responding but also decreased rates of alternative behavior. Experiment 2 compared the effects of 20% and 50% omission errors within subjects. Implementation at 50% integrity adequately suppressed responding, but treatment effects were lost at 20% integrity. There may be a critical level at which response cost must be implemented to suppress responding, which has important implications for application.

Assessing the efficacy of pictorial preference assessments for children with developmental disabilities.

Past research has demonstrated that pictorial preference assessments can predict subsequent reinforcement effects for individuals with developmental disabilities only when access to the selected stimulus is provided contingent on a pictorial selection. The purpose of the present investigation was to assess more comprehensively the feasibility of the pictorial format with children with developmental disabilities. In Experiment 1, prerequisite skill assessments were conducted, and the role of a contingent reinforcer was assessed by comparing the results from the pictorial assessment without contingent access to a reinforcer assessment. If contingent access was found to be necessary, the effects of schedule thinning were evaluated to determine whether a pictorial format could be made more practical in Experiment 2. The pictorial format without contingent access was successful with only some participants. However, schedule thinning was found to be an effective method to establish conditioned reinforcement properties for pictorial stimuli to create a more practical assessment for a subset of participants.

Development and evaluation of a secondary reference panel for BCR-ABL1 quantification on the International Scale.

Molecular monitoring of chronic myeloid leukemia patients using robust BCR-ABL1 tests standardized to the International Scale (IS) is key to proper disease management, especially when treatment cessation is considered. Most laboratories currently use a time-consuming sample exchange process with reference laboratories for IS calibration. A World Health Organization (WHO) BCR-ABL1 reference panel was developed (MR(1)-MR(4)), but access to the material is limited. In this study, we describe the development of the first cell-based secondary reference panel that is traceable to and faithfully replicates the WHO panel, with an additional MR(4.5) level. The secondary panel was calibrated to IS using digital PCR with ABL1, BCR and GUSB as reference genes and evaluated by 44 laboratories worldwide. Interestingly, we found that >40% of BCR-ABL1 assays showed signs of inadequate optimization such as poor linearity and suboptimal PCR efficiency. Nonetheless, when optimized sample inputs were used, >60% demonstrated satisfactory IS accuracy, precision and/or MR(4.5) sensitivity, and 58% obtained IS conversion factors from the secondary reference concordant with their current values. Correlation analysis indicated no significant alterations in %BCR-ABL1 results caused by different assay configurations. More assays achieved good precision and/or sensitivity than IS accuracy, indicating the need for better IS calibration mechanisms.

Ethanol differentially affects ATP-gated P2X(3) and P2X(4) receptor subtypes expressed in Xenopus oocytes.

P2X receptors are cation-selective, ligand-gated ion channels activated by synaptically released, extracellular adenosine 5'-triphosphate (ATP). ATP-gated currents are inhibited by ethanol when tested in dorsal root ganglion and CA1 neurons. Recently, we reported differences in sensitivity to ethanol inhibition between homomeric P2X(2) and P2X(4) receptors expressed in Xenopus oocytes, which suggested that subunit composition of native P2X receptors determines their ethanol sensitivity. The present study extended the investigation to P2X(3) receptors. The effects of ethanol and zinc ions (Zn(2+)) were tested on homomeric P2X(3) and P2X(4) receptors expressed in Xenopus oocytes using two-electrode voltage clamp. Ethanol potentiated ATP-gated P2X(3) receptor currents in a concentration dependent manner. In contrast, ethanol inhibited P2X(4) receptor function. Ethanol did not directly alter receptor function, nor did it alter the Hill coefficient or maximal ATP response (E(max)) in either P2X(3) or P2X(4) receptors. Ethanol increased the maximal response to Zn(2+) ATP-gated currents in P2X3 receptors which suggests that ethanol and Zn(2+) act on different sites. The differences in ethanol response of P2X(3) and P2X(4) receptors set the stage for future investigations that will use chimeric P2X receptors or other molecular manipulations of P2X structure to investigate the molecular sites and mechanisms of action of ethanol.