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Deep serum proteomics reveals biomarkers for prevalent and incident T2DM
No full textThe prevalence of type 2 diabetes mellitus (T2DM) is expected to increase rapidly in the next decades, posing a major challenge to societies worldwide1. The emerging era of precision medicine calls for the discovery of biomarkers of clinical value for prediction of disease onset, where causal biomarkers can furthermore provide actionable targets. Blood-based factors like serum proteins are in contact with every organ in the body to mediate global homeostasis and may thus directly regulate complex processes such as aging and the development of common chronic diseases. We applied a data-driven proteomics approach measuring serum levels of 4192 proteins in 5457 Icelanders to discover novel biomarkers for incident T2DM and further describe the serum protein profile of prevalent T2DM. We identified 537 proteins associated with incident or prevalent T2DM and, through LASSO penalized logistic regression analysis combined with bootstrap resampling, a panel of 20 protein biomarkers that accurately predicted incident T2DM was identified with a significant incremental improvement over traditional risk factors. Finally, a Mendelian randomization analysis provided additional support for a causal role of 51 proteins in the development of T2DM, which could be of particular interest as novel therapeutic targets
Cutaneous Immune Cell-Microbiota Interactions Are Controlled by Epidermal JunB/AP-1
No full textAtopic dermatitis is a complex, multi-factorial inflammatory skin disease. Uluçkan et al. show that JunBΔep mice display several hallmarks of AD, including S. aureus colonization. The authors characterize the immune responses to S. aureus and show that the adaptive immune system is required for protection against S. aureus colonization.
Atopic dermatitis (AD) is a multi-factorial skin disease with a complex inflammatory signature including type 2 and type 17 activation. Although colonization by S. aureus is common in AD, the mechanisms rendering an organism prone to dysbiosis, and the role of IL-17A in the control of S. aureus-induced skin inflammation, are not well understood. Here, we show several pathological aspects of AD, including type 2/type 17 immune responses, elevated IgE, barrier dysfunction, pruritus, and importantly, spontaneous S. aureus colonization in JunBΔep mice, with a large transcriptomic overlap with AD. Additionally, using Rag1−/− mice, we demonstrate that adaptive immune cells are necessary for protection against S. aureus colonization. Prophylactic antibiotics, but not antibiotics after established dysbiosis, reduce IL-17A expression and skin inflammation, examined using Il17a-eGFP reporter mice. Mechanistically, keratinocytes lacking JunB exhibit higher MyD88 levels in vitro and in vivo, previously shown to regulate S. aureus colonization. In conclusion, our data identify JunB as an upstream regulator of microbiota-immune cell interactions and characterize the IL-17A response upon spontaneous dysbiosis
Use of Alternative Designs and Data Sources for Pediatric Trials
No full textChildren are considered a vulnerable group and as such are granted additional protection as research subjects. Research projects using children as research subjects are justifiable if the answer to the scientific question of the study cannot be obtained by enrolling adult subjects (cf. scientific necessity). Thus, there is an ethical obligation to explore innovative analytical strategies that seek balance between the feasibility of conducting a trial and maximizing the utilization of data on efficacy and safety. On this note, there is enthusiasm for implementing some less popular but efficient alternative designs for confirmatory pediatric trials. Within the pediatric extrapolation paradigm, examples of such designs, other than purely based on pharmacokinetic/pharmacodynamic data, are described in this article along with their advantages and disadvantages. This article will also discuss how to incorporate alternative data sources in the analysis of pediatric clinical trials. A discussion of existing approaches and a road-map to their utilization will be provided. Real case examples on the use of the approaches are provided
Extrinsic and intrinsic apoptosis activate pannexin-1 to drive NLRP3 inflammasome assembly
No full textPyroptosis is a form of lytic inflammatory cell death driven by inflammatory caspase-1, caspase-4, caspase-5 and caspase-11. These caspases cleave and activate the pore-forming protein gasdermin D (GSDMD) to induce membrane damage. By contrast, apoptosis is driven by apoptotic caspase-8 or caspase-9 and has traditionally been classified as an immunologically silent form of cell death. Emerging evidence suggests that therapeutics designed for cancer chemotherapy or inflammatory disorders such as SMAC mimetics, TAK1 inhibitors and BH3 mimetics promote caspase-8 or caspase-9-dependent inflammatory cell death and NLRP3 inflammasome activation. However, the mechanism by which caspase-8 or caspase-9 triggers cell lysis and NLRP3 activation is still undefined. Here, we demonstrate that during extrinsic apoptosis, caspase-1 and caspase-8 cleave GSDMD to promote lytic cell death. By engineering a novel Gsdmd D88A knock-in mouse, we further demonstrate that this proinflammatory function of caspase-8 is counteracted by caspase-3-dependent cleavage and inactivation of GSDMD at aspartate 88, and is essential to suppress GSDMD-dependent cell lysis during caspase-8-dependent apoptosis. Lastly, we provide evidence that channel-forming glycoprotein pannexin-1, but not GSDMD or GSDME promotes NLRP3 inflammasome activation during caspase-8 or caspase-9-dependent apoptosis
Coolade. A Low-Foaming Surfactant for Organic Synthesis in Water
No full textSeveral types of reduction reactions in organic synthesis run under aqueous micellar catalysis conditions (i.e., in water at ambient temperatures) produce significant volumes of foam due to the combination of the surfactant being used in the presence of gas evolution. The newly engineered surfactant “Coolade” minimizes this important technical issue due to its low-foaming properties. Coolade is the latest in a series of designer surfactants specifically tailored to enable organic synthesis in water. We herein report the synthesis of this new surfactant along with its applications to gas-involving reactions
Fragment-based screening of programmed death ligand 1 (PD-L1).
No full textThe PD-1 immune checkpoint pathway is a highly validated target for cancer immunotherapy. Despite the potential advantages of small molecule inhibitors over antibodies, the discovery of small molecule checkpoint inhibitors has lagged behind. To discover small molecule inhibitors of the PD-1 pathway, we have utilized a fragment-based approach. Small molecules were identified that bind to PD-L1 and crystal structures of these compounds bound to PD-L1 were obtained
A conditional inducible JAK2V617F transgenic mouse model reveals myeloproliferative disease that is reversible upon switching off transgene expression
No full textAberrant activation of the JAK/STAT pathway is thought to be the critical event in the pathogenesis of the chronic myeloproliferative neoplasms (MPNs) polycythemia vera, essential thrombocythemia and primary myelofibrosis. The most frequent genetic alteration in these pathologies is the activating JAK2V617F mutation, and expression of the mutant gene in mouse models was shown to cause a phenotype resembling the human diseases. Given the body of genetic evidence, it has come as a sobering finding that JAK inhibitor therapy only modestly suppresses the JAK2V617F allele burden, despite showing clear benefits in terms of reducing splenomegaly and constitutional symptoms in patients. To gain a better understanding if JAK2V617F is required for maintenance of myeloproliferative disease once it has evolved, we generated a conditional inducible transgenic JAK2V617F mouse model using the SCL-tTA-2S tet-off system. Our model corroborates that expression of JAK2V617F in hematopoietic stem and progenitor cells recapitulates key hallmarks of human MPNs, and exhibits gender differences in disease manifestation. The disease was found to be transplantable, and importantly, reversible when transgenic JAK2V617F expression was switched off. Our results indicate that mutant JAK2V617F-specific inhibitors should result in profound disease modification by disabling the MPN clone bearing mutant JAK2
All-Assay-Max2 pQSAR: Activity predictions as accurate as 4-concentration IC50s for nearly 9,000 Novartis assays
No full textProfile-QSAR (pQSAR) is a massively multi-task, 2-step machine learning method with unprecedented scope, accuracy and applicability domain. In step one, a “profile” of conventional single-assay random forest regression (RFR) models are trained on a very large number of biochemical and cellular pIC50 assays using Morgan 2 sub-structural fingerprints as compound descriptors. In step 2, a panel of PLS models are built using the profile of pIC50 predictions from those RFR models as compound descriptors. Hence the name. Previously described for a panel of 728 biochemical and cellular kinase assays, we have now built an enormous pQSAR from 11,805 diverse Novartis IC50 and EC50 assays. This large number of assays, and hence of compound descriptors for PLS, dictated reducing the profile by only including RFR models whose predictions correlate with the assay being modeled. We evaluate both the RFR and pQSAR models with our “realistically novel” held-out test set whose median, average similarity to the nearest training set member across the 11,805 assays was only 0.34, thus testing a realistically large applicability domain. For the 11,805 single-assay RFR models, the median correlation of prediction with experiment was only R2ext=0.05, virtually random, and only 8% of the models achieved our standard success threshold of R2ext=0.30. For pQSAR, the median correlation was R2ext=0.53, comparable to 4-concentration experimental IC50s, and 72% of the models met our R2ext>0.30 standard, totaling 8558 successful models. The successful models included assays from all of the 51 annotated target sub-classes, as well as 4196 phenotypic assays, indicating that pQSAR can be applied to virtually any disease area. Every month, all models are updated to include new measurements, and predictions are made for 5.5 million Novartis compounds, totaling 50 billion predictions. Common uses have included virtual screening, selectivity design, toxicity and promiscuity prediction, mechanism-of-action prediction, and others
Behavior of mice aboard the International Space Station
No full textInterest in space habitation has grown dramatically with planning underway for the first human transit to Mars. Despite a robust history of domestic and international spaceflight research, understanding behavioral adaptation to the space environment for extended durations is scant. Here we report the first detailed behavioral analysis of mice flown in the NASA Rodent Habitat on the International Space Station (ISS). Following 4-day transit from Earth to ISS, video images were acquired on orbit from 16- and 32-week-old female mice. Spaceflown mice engaged in a full range of species-typical behaviors. Physical activity was greater in younger flight mice as compared to identically-housed ground controls, and followed the circadian cycle. Within 7-10 days after launch, younger (but not older), mice began to exhibit distinctive circling or 'race-tracking' behavior that evolved into coordinated group activity. Organized group circling behavior unique to spaceflight may represent stereotyped motor behavior, rewarding effects of physical exercise, or vestibular sensation produced via self-motion. Affording mice the opportunity to grab and run in the RH resembles physical activities that the crew participate in routinely. Our approach yields a useful analog for better understanding human responses to spaceflight, providing the opportunity to assess how physical movement influences responses to microgravity
Evaluation of the Effect of 5 QT-Positive Drugs on the JTpeak Interval — An Analysis of ECGs From the IQ-CSRC Study
No full textThe JTpeak interval has been proposed as a new biomarker to demonstrate mixed ion channel effects, potentially leading to reduced late-stage electrocardiogram (ECG) monitoring for mildly QT-prolonging drugs. ECG waveforms from the IQ-CSRC study were used. Twenty healthy subjects were enrolled with 6 subjects on placebo and 9 subjects on each of 5 mildly QT-prolonging drugs — moxifloxacin, dofetilide, ondansetron, dolasetron, and quinine — and 1 negative drug, levocetirizine. A vector magnitude lead was derived from 12-lead ECGs, and measurements were made on a median beat from three 10-second replicates. Data were analyzed using a linear concentration-response model with QTcF and heart rate corrected JTpeak (JTpeak_c) as dependent variables. For moxifloxacin, dofetilide, and ondansetron, all pure hERG blockers, slopes of the concentration (C)-QTcF and C-JTpeak_c relationships were positive and statistically significant. With the prespecified linear model, the predicted effects on ΔΔQTcF and ΔΔJTpeak_c were 11.4 and 9.4 milliseconds for moxifloxacin at the geometric mean Cmax on day 1, 9.0 and 11.7 milliseconds for dofetilide and 11.5, and 7.9 milliseconds for ondansetron, respectively. In contrast, dolasetron and quinine, both with additional ion channel effects, prolonged QTcF with a positive C-ΔQTcF slope and predicted ΔΔQTcF effect on day 1 of 6.2 and 11.4 milliseconds, whereas the C-ΔJTpeak_c slope and the predicted ΔΔJTpeak on day 1 were negative (−0.3 and −7.5 milliseconds per ng/mL). Pure hERG-blocking drugs prolonged both the QTc and the JTpeak_c intervals, whereas drugs with mixed ion channel effects, including peak sodium inhibition, prolonged QTcF but not the JTpeak_c interval