Action Profiling Research Articles

S102 - NMRA-140, a Novel and Selective Kappa Opioid Receptor Antagonist: Mode of Action Profiling Studies Demonstrate No Opioid Agonist Properties Implicated in Opioid-Related Abuse

S102 - NMRA-140, a Novel and Selective Kappa Opioid Receptor Antagonist: Mode of Action Profiling Studies Demonstrate No Opioid Agonist Properties Implicated in Opioid-Related Abuse

Preclinical evaluation of the individualized approach for chronic non-healing wounds management

Chronic non‑healing wounds (CNHW) are very common and often incorrectly treated, the morbidity and associated costs of chronic wounds management highlight the need to implement wound prevention and treatment concepts. Objective — to evaluate the possibility of different metal nanooxide polymer nanofilms use for CNHW’ local treatment. Materials and methods. The study design is based on evaluation of various types of dressing materials considering their option for use in CNHW local treatment. Samples of biodegradable polymer films (with an optimal composition of gelatin, polyvinyl alcohol, lactic acid, glycerin and distilled water) saturated with nanoparticles of several oxides with expected antibacterial and pro‑regenerative feature — nZnO, nMgO in concentrations of 1%, 5%, and 10% were used in the study of antimicrobial action and substance release profiling. Quarterly ammonium antiseptic decamethoxin 0.02% was used for control. Results. Obtained data shows that polymer based biodegradable films incorporating optimal component composition (gelatin, polyvinyl alcohol, lactic acid and glycerin) enriched with 5% and 10% zinc nanooxide have potent antimicrobial activity against both gram‑positive and gram‑negative microorganisms, the most common causative agents of CNHW’s. The ion release capacity analysis showed that the Zinc impregnated wound‑healing biodegradable polymer film gradually releases the active substance in a time dependent manner, and the nano‑sized particles of nanoZinc oxide are released from the polymer composition faster than ordinary zinc oxide. Conclusions. Complex natural biodegradable polymer based nanofilms are composite materials impregnated with metal nanooxides showing high potential in local treatment of chronic non‑healing wounds. Polymer film with 5% nanoZnO showed up to the 58% higher antimicrobial activity, comparable or exceeding the one of quarterly ammonium compound decamethoxin. Furthermore, nanoZnO impregnated polymer films compared to standard ZnO impregnated polymer films showed up to 63.2% faster substance release profile with rapid and more unified curve.

A semiconductor 96-microplate platform for electrical-imaging based high-throughput phenotypic screening

High-content imaging for compound and genetic profiling is popular for drug discovery but limited to endpoint images of fixed cells. Conversely, electronic-based devices offer label-free, live cell functional information but suffer from limited spatial resolution or throughput. Here, we introduce a semiconductor 96-microplate platform for high-resolution, real-time impedance imaging. Each well features 4096 electrodes at 25 µm spatial resolution and a miniaturized data interface allows 8× parallel plate operation (768 total wells) for increased throughput. Electric field impedance measurements capture >20 parameter images including cell barrier, attachment, flatness, and motility every 15 min during experiments. We apply this technology to characterize 16 cell types, from primary epithelial to suspension cells, and quantify heterogeneity in mixed co-cultures. Screening 904 compounds across 13 semiconductor microplates reveals 25 distinct responses, demonstrating the platform’s potential for mechanism of action profiling. The scalability and translatability of this semiconductor platform expands high-throughput mechanism of action profiling and phenotypic drug discovery applications.

225. NMRA-140, a Novel and Selective Kappa Opioid Receptor Antagonist: Mode of Action Profiling Studies Demonstrate No Opioid Agonist Properties Implicated in Opioid-Related Abuse

225. NMRA-140, a Novel and Selective Kappa Opioid Receptor Antagonist: Mode of Action Profiling Studies Demonstrate No Opioid Agonist Properties Implicated in Opioid-Related Abuse

Meropenem in combination with baicalein exhibits synergism against extensively drug resistant and pan-drug-resistant Acinetobacter baumannii clinical isolates in vitro.

Several studies have demonstrated that the effectiveness of carbapenems against drug-resistant Acinetobacter baumannii infections has been decreasing. Combination therapy with two or more drugs is currently under investigation to overcome the emerging resistance against carbapenems. In this study, we tested the possible synergistic interactions of a potent antibacterial flavonoid, baicalein, with meropenem to illustrate this duo's antibacterial and antibiofilm effects on 15 extensively drug resistant or pan-drug-resistant (XDR/PDR) A. baumannii clinical isolates in vitro. Isolates included in the study were identified with MALDI-TOF MS, and antibiotic resistance patterns were studied according to EUCAST protocols. Carbapenem resistance was confirmed with the modified Hodge test, and resistance genes were also analyzed with genotypical methods. Then, checkerboard and time-kill assays were performed to analyze antibacterial synergism. Additionally, a biofilm inhibition assay was performed for screening the antibiofilm activity. To provide structural and mechanistic insights into baicalein action, protein-ligand docking, and interaction profiling calculations were conducted. Our study shed light on the remarkable potential of the baicalein-meropenem combination, since either synergistic or additive antibacterial activity was observed against every XDR/PDR A. baumannii strain in question. Furthermore, the baicalein-meropenem combination displayed significantly better antibiofilm activity in contrast to standalone use. In silico studies predicted that these positive effects arose from inhibition by baicalein of A. baumannii beta-lactamases and/or penicillin-binding proteins. Overall, our findings highlight the prospective potential benefits of baicalein in combination with meropenem for the treatment of carbapenem-resistant A. baumannii infections.

Abstract LB-007: The emergence of tumor organoid engineering as a powerful tool for drug efficacy and mechanism of action profiling

Abstract Introduction: Patient-derived models such as organoids are a powerful translational tool for preclinical research with several advantages over current conventional models: 1) they are derived from stem cells with self-organizing and renewal properties reflecting the 3D structure and morphology of original tissue; 2) they preserve the original genomic, physiological, and multicellular identity thereby representing patient diversity and heterogeneity in a dish; 3) they provide in vitro convenience for greater throughput for screening large numbers of compounds and/or patients; 4) they are amenable to genetic engineering. Methods: We have used Hubrecht Organoid Technology (HUB) protocols to systematically create a biobank of organoids derived from a well-annotated patient-derived xenograft (PDX) library (the world's largest commercially available PDX library with over 2,500 models, covering a variety of carcinomas, with extensive pathology, genomic, and treatment information), referred to as PDX-derived organoids or PDXOs. Results: We have currently developed more than 200 PDXOs covering many cancer types, including colorectal, gastric, liver, lung, bladder, breast, ovarian, pancreatic cancer, etc. Genomic analyses have demonstrated that PDXOs largely preserve the histo- and molecular pathology of the original PDX. In vitro maintenance of PDX models as organoids enables genetic manipulation to be performed, which is not possible for in vivo PDX models, thereby creating new models to facilitate drug development. We have employed both lentivirus-based and CRISPR/Cas9-based approaches to modify various tumor organoid models with luciferase, tumor associated antigens (TAA), and some EGFR mutants unavailable in current preclinical model panels. These engineered organoid models, alongside corresponding in vivo matched models, have been incorporated into a new drug efficacy platform. Preliminary results show that these engineered organoid models greatly expand the capacity and application of our current organoid bank. Conclusions: In summary, engineered cancer organoids are a great tool to enrich and diversify current tumor organoid biobanks, particularly for unavailable and clinically relevant models to be used for drug testing, as well as for studies on mechanisms of action and drug resistance. Citation Format: Fei Wang, Rongyun Zhou, Xinlong Dong, Yuqing Ma, Jun Zhou, Xiaoxi Xu, Yujun Huang, Henry Li, Phillip S-Z Wang. The emergence of tumor organoid engineering as a powerful tool for drug efficacy and mechanism of action profiling [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-007.

Integrating QSAR models predicting acute contact toxicity and mode of action profiling in honey bees (A. mellifera): Data curation using open source databases, performance testing and validation

Honey bees (Apis mellifera) provide key ecosystem services as pollinators bridging agriculture, the food chain and ecological communities, thereby ensuring food production and security. Ecological risk assessment of single Plant Protection Products (PPPs) requires an understanding of the exposure and toxicity. In silico tools such as QSAR models can play a major role for the prediction of structural, physico-chemical and pharmacokinetic properties of chemicals as well as toxicity of single and multiple chemicals. Here, the first integrative honey bee QSAR model has been developed for PPPs using EFSA's OpenFoodTox, US-EPA ECOTOX and Pesticide Properties DataBase i) to predict acute contact toxicity (LD50) and ii) to profile the Mode of Action (MoA) of pesticides active substances. Three different classification-based and four regression-based models were developed and tested for their performance, thus identifying two models providing the most reliable predictions based on k-NN algorithm. The two-category QSAR model (toxic/non-toxic; n = 411) was validated using sensitivity (=0.93), specificity (=0.85), balanced accuracy (=0.90), and Matthews correlation coefficient (MCC = 0.78) as statistical parameters. The regression-based model (n = 113) was validated for its reliability and robustness (R2 = 0.74; MAE = 0.52). Current study proposes the MoA profiling for 113 pesticides active substances and the first harmonised MoA classification scheme for acute contact toxicity in honey bees, including LD50s data points from three different databases. The classification allows to further define MoAs and the target site of PPPs active substances, thus enabling regulators and scientists to refine chemical grouping and toxicity extrapolations for single chemicals and component-based mixture risk assessment of multiple chemicals. Relevant future perspectives are briefly addressed to integrate MoA, adverse outcome pathways (AOPs) and toxicokinetic information for the refinement of single-chemical/combined toxicity predictions and risk estimates at different levels of biological organization in the bee health context.

Impact of methyl salicylate on storage quality, ethylene action, and protein profiling of ‘Zaosu’ pear (Pyrus bretschneideri)

‘Zaosu’ pear, an early-maturing cultivar harvested in summer in China, readily exhibits quality deterioration and senescence owing to the lack of refrigeration and effective postharvest technology. To explore possible molecular mechanisms of methyl salicylate (MeSA) in delaying senescence, ‘Zaosu’ pear was treated with 0.05 mmol L―1 MeSA and stored at 25 ± 2 °C. The effects on quality, ethylene synthesis and signaling elements, and protein profiling were evaluated. MeSA maintained fruit postharvest quality. In the treated fruit, the yellowing index, weight loss, and color L* value retained lower levels, and the yellowing index was 69 % of that of the control after 20 days, whereas the hue angle and texture properties (fracturability, firmness, and chewiness) had increased levels. The respiration rate and ethylene content, and ethylene biosynthesis and signaling components ACS4, ACO1, ACO4, ETR1, ETR2, ERS2, CTR1, EIN2, and EIL1, were inhibited and downregulated by MeSA, respectively, while ERS1 was upregulated. Thirty-eight different expression proteins were identified by 2-DE and MALDI-TOF/TOF MS analysis in the treated and untreated fruits. Among them, related ripening and senescence proteins 2-methylene-furan-3-one reductase, abscisic stress-ripening protein, annexin RJ4, and mitochondrial outer membrane protein porin were down-accumulated during storage, while electron transfer flavoprotein was up-accumulated later in the storage period. These results indicated that MeSA postponed the ripening and senescence of postharvest ‘Zaosu’ pear.

Developing high through put screening strategies to identify the next generation of antimicrobials

Area of expertise: My research interests focus on identifying novel compounds that can disrupt key phenotypes associated with antibiotic resistance such as biofilm formation and motility in the pathogens Acinetobacter baumannii and Pseudomonas aeruginosa. To this end we have developed a range of screens and assays to facilitate compound identification and mechanism of action profiling in these pathogens. These include: Antibiotic Resistance Inhibitor Screens, Biofilm Inhibition Screens, Biofilm Disruption Screens, Signalling Pathway Specific Biosensor Screens , Mechanism of Action Assays, Resistance Development Screens, Antibiotic Resistance Inhibitor Screens, In vivo biofilm and pathogenicity models to validate lead compounds and perform toxicity profiling. Future goals: Future goals include being part of a collaborative multidisciplinary drug discovery network that has access to diverse and bespoke extract/compound collections and the expertise to identify the bioactive leads, characterise their mechanism of action and ultimately take them forward to clinical trials. Further to this the goal is to target national and international funding opportunities that will facilitate taking active compounds right through the drug discovery pipeline from the bench to the bedside. I am hoping to identify collaborators across all aspects of drugs discovery, particularly collaborators with: Access to compound or extract collections, Expertise in compound identification ,Clinical trial expertise.

Design, synthesis and biological evaluation of uncharged catechol derivatives as selective inhibitors of PTP1B

Protein tyrosine phosphatases 1B (PTP1B) is a promising and validated therapeutic target to effectively treat T2DM and obesity. However, the development of charged PTP1B inhibitors was restricted due to their low cell permeability and poor bioavailability. Based on active natural products, two series of uncharged catechol derivatives were identified as PTP1B inhibitors by targeting a secondary aryl phosphate-binding site as well as the catalytic site. The most potent inhibitor 22 showed an IC50 of 0.487 μM against PTP1B and strong selectivity (27-fold) over TCPTP. Kinetic studies were also performed that 22 act as a competitive PTP1B inhibitor. The treatment of C2C12 myotubes with 22 markedly increased the phosphorylation levels of IRβ, Akt and IRS1 phosphorylation. The similarity of its action profiling with that produced by insulin suggested its potential as a new non-insulin-dependent drug candidate.

The First Principles of Standing

This Article examines the indeterminacy of standing doctrine by deconstructing recent desegregation, affirmative action, and racial profiling cases. This examination is an attempt to uncover the often unstated meta-principles that guide standing jurisprudence. The Article contends that the inherent indeterminacy of standing law can be understood as reflecting an unstated desire to protect racial and class privilege, which is accomplished through the dogma of individualism, equal opportunity (liberty), and “white innocence.” Relying on insights from System Justification Theory, a burgeoning field of social psychology, the Article argues that the seemingly incoherent results in racial standing cases can be understood as unconscious attempts to preserve the status quo. The Article proposes moving “beyond the transcendental nonsense” of standing doctrine and its inevitable replication of economic and racial privilege by completely eliminating all standing limitations to the access of justice.

Comparison of similarity measures to differentiate players' actions and decision-making profiles in serious games analytics

Three Gameplay Action-Decision (GAD) profiles: Explorer, Fulfiller, and Quitter, have been identified based on individual's decision-making actions and navigational behaviors in situ serious games. The ability to profile trainees using serious games can yield new analytics and insights towards training and learning performance improvement, including the identification of weaknesses or potential training needs in the players towards adaptive training, and the creation of new diagnostics for prescriptive training, retraining, and remediation. Similarity measures of players' in-game course of actions (COAs) have been shown to be a viable approach in differentiating novices from experts in serious games.In this study, we examined and compared several popular similarity measures to see if any measure, or combination of measures, would be viable in differentiating players based on their GAD profiles in serious games. Our findings revealed that similarity measures, while significant in their predicting abilities individually, could gain more strength from one another in combination. More research is needed to create or develop new metrics and methods for players’ action and behavioral profiling in Serious Games Analytics.

ADP GloTM: A One Platform Ideal for Assay Development, Screening and Profiling of All Phosphotransferases

Drug safety is of significant important importance in the pharmaceutical industry indicating that minimal side effects constitute a major requirement in drug development. Thus, after rigorous screening and lead optimization of the lead compounds, profiling of the compound against a broad panel of kinases is important for providing a better understanding of its activity and obviating any off‐target activities. The luminescent ADP‐GloTM kinase assay is a universal and robust platform that measures kinase activity by quantifying the amount of ADP produced during the enzymatic reaction. We have tested the utility of this platform against panels of different families of the human kinome here we show that using the new kinase profiling format we could easily generate selectivity profiles using small or large kinase panels, as well as identifying compound promiscuity towards members of a single kinase subfamily or different subfamilies of the kinome including lipid kinases. The ADP‐Glo™ KES platforms now address all the needs of basic kinase characterizations, kinase screening, mode of action (MOA) studies and profiling in an affordable manner using one assay format during the drug discovery process

IM(V)ERSÕES CORPO AMBIENTE E A CRIAÇÃO COREO-VIDEOGRÁFICA

O texto apresenta e discute a terminologia Imersão Corpo Ambiente, parte do projeto de pesquisa “Corpo, Ritmo, Imagem: A Criação Coreo-Videográfica Transcultural” (PPGAC/UFBA e CNPq, 2010-2014), e suas implicações somático-performativas, bem como sua aplicação na criação do video-dança Im(v)ersões Corpo Ambiente, de Uirá Meneses. A partir de alguns termos, como intervenção urbana, site specific art, site functional art, composição coreo-videográfica, sintonia somática e pulsão espacial, e possibilidades associativas de diferentes abordagens, como a Laban/Bartenieff Movement Analysis, o Movement Pattern Analysis (Action Profiling) e o Body-Mind Centering, o processo criativo interartístico de base somático-performativa é enfatizado como pesquisa acadêmica sensível e atual.

Evaluation of licorice flavonoids as protein tyrosine phosphatase 1B inhibitors

Protein tyrosine phosphatase 1B (PTP1B) is a major negative regulator in insulin- and leptin-signaling cascades as well as a positive regulator in tumorigenesis, and much attention has been paid to PTP1B inhibitors as potential therapies for diabetes, obesity, and cancer. In the present study, the screening of a compound library of licorice flavonoids allowed for the discovery of several compounds, including licoagrone (3), licoagrodin (4), licoagroaurone (5), and isobavachalcone (6), as new PTP1B inhibitors. It was revealed that these compounds inhibit the activity of PTP1B in different modes and with different selectivities and that they exhibit different cellular activity in the insulin-signaling pathway. Glycybenzofuran (1), a competitive PTP1B inhibitor, showed both excellent inhibitory selectivity against PTP1B and cellular activity on the insulin-stimulated Akt phosphorylation level. The similarity of its action profiling in the insulin-signaling pathway suggested its potential as a new anti-insulin-resistant drug candidate.

“Function-First” Lead Discovery: Mode of Action Profiling of Natural Product Libraries Using Image-Based Screening

Cytological profiling is a high-content image-based screening technology that provides insight into the mode of action (MOA) for test compounds by directly measuring hundreds of phenotypic cellular features. We have extended this recently reported technology to the mechanistic characterization of unknown natural products libraries for the direct prediction of compound MOAs at the primary screening stage. By analyzing a training set of commercial compounds of known mechanism and comparing these profiles to those obtained from natural product library members, we have successfully annotated extracts based on MOA, dereplicated known compounds based on biological similarity to the training set, and identified and predicted the MOA of a unique family of iron siderophores. Coupled with traditional analytical techniques, cytological profiling provides an avenue for the creation of "function-first" approaches to natural products discovery.

Targeted estrogen delivery reverses the metabolic syndrome

We report the development of a new combinatorial approach that allows for peptide-mediated selective tissue targeting of nuclear hormone pharmacology while eliminating adverse effects in other tissues. Specifically, we report the development of a glucagon-like peptide-1 (GLP-1)-estrogen conjugate that has superior sex-independent efficacy over either of the individual hormones alone to correct obesity, hyperglycemia and dyslipidemia in mice. The therapeutic benefits are driven by pleiotropic dual hormone action to improve energy, glucose and lipid metabolism, as shown by loss-of-function models and genetic action profiling. Notably, the peptide-based targeting strategy also prevents hallmark side effects of estrogen in male and female mice, such as reproductive endocrine toxicity and oncogenicity. Collectively, selective activation of estrogen receptors in GLP-1-targeted tissues produces unprecedented efficacy to enhance the metabolic benefits of GLP-1 agonism. This example of targeting the metabolic syndrome represents the discovery of a new class of therapeutics that enables synergistic co-agonism through peptide-based selective delivery of small molecules. Although our observations with the GLP-1-estrogen conjugate justify translational studies for diabetes and obesity, the multitude of other possible combinations of peptides and small molecules may offer equal promise for other diseases.

Development of Antibiotic Activity Profile Screening for the Classification and Discovery of Natural Product Antibiotics

Despite recognition of the looming antibiotic crisis by healthcare professionals, the number of new antibiotics reaching the clinic continues to decline sharply. This study aimed to establish an antibiotic profiling strategy using a panel of clinically relevant bacterial strains to create unique biological fingerprints for all major classes of antibiotics. Antibiotic mode of action profile (BioMAP) screening has been shown to effectively cluster antibiotics by structural class based on these fingerprints. Using this approach, we have accurately predicted the presence of known antibiotics in natural product extracts and have discovered a naphthoquinone-based antibiotic from our marine natural product library that possesses a unique carbon skeleton. We have demonstrated that bioactivity fingerprinting is a successful strategy for profiling antibiotic lead compounds and that BioMAP can be applied to the discovery of new natural product antibiotics leads.

Antibiotic mode of action profiling (Biomap) as a platform for the discovery of novel natural product antibiotics

Antibiotic mode of action profiling (Biomap) as a platform for the discovery of novel natural product antibiotics

Studies on the antimicrobial properties of N-acylated ciprofloxacins

Fluoroquinolone antibiotics have been a mainstay in the treatment of bacterial diseases. The most notable representative, ciprofloxacin, possesses potent antimicrobial activity; however, a rise in resistance to this agent necessitates development of novel derivatives to prolong the clinical lifespan of these antibiotics. Herein we have synthesized and analyzed the antimicrobial properties of a library of N-acylated ciprofloxacin analogues. We find that these compounds are broadly effective against Gram-positive and Gram-negative bacteria, with many proving more effective than the parental drug, and several possessing MICs ⩽1.0μg/ml against methicillin-resistant Staphylococcus aureus and Bartonella species. An analysis of spontaneous mutation frequencies reveals very low potential for resistance in MRSA compared to existing fluoroquinolones. Mode of action profiling reveals that modification of the piperazinyl nitrogen by acylation does not alter the effect of these molecules towards their bacterial target. We also present evidence that these N-acylated compounds are highly effective at killing intracellular bacteria, suggesting the suitability of these antibiotics for therapeutic treatment.