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Case Presentation A 62-year-old nonsmoking woman with no medical history initially presented with a 3-month history of rash. A painful, erythematous exanthem had progressed from her forehead, cheeks, and upper chest to her eyes (heliotrope rash) and hands, primarily involving the extensor surface finger joints with prominent digital ulceration.

IntroductionProcalcitonin expression is thought to be stimulated by bacteria and suppressed by viruses via interferon signalling. Consequently, during respiratory viral illness, clinicians often interpret elevated procalcitonin as evidence of bacterial coinfection, prompting antibiotic administration. We sought to evaluate the validity of this practice and the underlying assumption that viral infection inhibits procalcitonin synthesis.MethodsWe conducted a retrospective cohort study of patients hospitalised with pure viral infection (n=2075) versus bacterial coinfection (n=179). The ability of procalcitonin to distinguish these groups was assessed. In addition, procalcitonin and interferon gene expression were evaluated in murine and cellular models of influenza infection.ResultsPatients with bacterial coinfection had higher procalcitonin than those with pure viral infection, but also more severe disease and higher mortality (pDiscussionThese studies reveal that procalcitonin rises during pure viral infection in proportion to disease severity and is not suppressed by interferon signalling, in contrast to prior models of procalcitonin regulation. Applied clinically, our data suggest that procalcitonin represents a better indicator of disease severity than bacterial coinfection during viral respiratory infection.

Neurogenic cardiac impairment can occur after acute ischemic stroke (AIS), but the mapping of the neuroanatomic correlation of stroke-related myocardial injury remains uncertain. This study aims to identify the association between cardiac outcomes and middle cerebral artery (MCA) ischemic stroke, with or without insular cortex involvement, as well as the impact of new-onset atrial fibrillation (AF) after AIS on recurrent stroke.Serial measurements of high sensitivity troponin T (TnT), brain natriuretic peptide (BNP), electrocardiography (ECG), echocardiogram, and cardiac monitoring were performed on 415 patients with imaging confirmed MCA stroke, with or without insular involvement. Patients with renal failure, recent cardiovascular events, or congestive heart failure were excluded.One hundred fifteen patients (28%) had left MCA infarcts with insular involvement, 122 (29%) had right MCA infarcts involving insular cortex, and 178 (43%) had no insular involvement. Patients with left MCA stroke with insular involvement tended to exhibit higher BNP and TnI, and transient cardiac dysfunction, which mimicked Takotsubo cardiomyopathy in 10 patients with left ventricular ejection fraction (LVEF) of 20-40%. Incidence of new-onset AF was higher in right MCA stroke involving insula (39%) than left MCA involving insula (4%). Nine out of fifty-three patients with new-onset AF were not on anticoagulant therapy due to various reasons; none of them experienced recurrent AF or stroke during up to a 3-year follow-up period. Statistically significant correlations between BNP or TnT elevation and left insular infarcts, as well as the incidence of AF and right insular infarcts, were revealed using linear regression analysis.The present study demonstrated that acute left MCA stroke with insular involvement could cause transient cardiac dysfunction and elevated cardiac enzymes without persistent negative outcomes in the setting of health baseline cardiac condition. The incidence of new-onset AF was significantly higher in patients with right MCA stroke involving the insula. There was no increased risk of recurrent ischemic stroke in nine patients with newly developed AF who were not on anticoagulant therapy, which indicated a need for further research on presumed neurogenic AF and its management.

Adult-onset Still’s disease (AOSD) is an uncommon inflammatory condition characterised by a triad of fevers, arthralgias and a salmon-coloured rash. It is also strongly associated with high ferritin levels, whose role in its pathogenesis is not entirely clear. Central nervous system (CNS) manifestations are exceedingly rare in this disease, accounting for only a handful of reported cases. Herein, we describe a case of a 63-year-old woman who developed new-onset psychiatric symptoms in the months preceding her diagnosis. 2 months after her diagnosis, she experienced an exacerbation of psychiatric symptoms followed by new-onset seizures in conjunction with an acute lung infection. In addition, we discuss two other previously reported cases of AOSD patients with psychiatric symptoms as their initial presentation.

The past two decades have witnessed a resurgence in neutrophil research, inspired in part by the discovery of neutrophil extracellular traps (NETs) and their myriad roles in health and disease. Within the lung, dysregulation of neutrophils and NETosis have been linked to an array of diseases including pneumonia, cystic fibrosis, acute respiratory distress syndrome (ARDS), chronic obstructive pulmonary disease (COPD), and severe asthma. However, our understanding of pathologic neutrophil responses in the lung remains incomplete. Two methodologic issues have contributed to this gap: first, an emphasis on studying neutrophils from blood rather than the lung and second, the technical difficulties of interrogating neutrophil responses in mice, which has largely restricted basic murine research to specialized laboratories. To address these limitations, we have developed a suite of techniques for studying neutrophil effector functions specifically in the mouse lung. These include ex vivo assays for phagocytosis and NETosis using bronchoalveolar neutrophils and in situ evaluation of NETosis in a murine model of pneumonia. Throughout, we have prioritized technical ease and robust, quantitative readouts. We hope these assays will help to standardize research on lung neutrophils and improve accessibility to this burgeoning field.

Seasonal influenza (flu) is an underappreciated source of disease morbidity and mortality worldwide. While vaccination remains the cornerstone of influenza prevention, common measures practiced during the COVID-19 pandemic such as social distancing, the use of protective face masks, and frequent hand washing are rarely utilized during flu season. In this investigation, we examined the effect of these preventative measures in decreasing influenza burden this year. We examined three countries with major COVID-19 outbreaks i.e. China, Italy and the United States, and compared the flu activity this year to the average of the last 4 years (2015-2019). We found that this year in China and Italy, there was a significantly steeper decline of flu cases than average, which correlated with an increase in positive COVID-19 case reports in those countries. These "averted" cases can be translated into a substantial decrease in morbidity and mortality. As such, we conclude that the current COVID-19 pandemic is a reminder that behavioral measures can decrease the burden of communicable respiratory infections, and these measures should be adopted to an extent during normal influenza season.

Iterative structure–activity analyses in a class of highly functionalized furo[2,3-b]pyridines led to the identification of the second generation pan-genotypic hepatitis C virus NS5B polymerase primer grip inhibitor BMT-052 (14), a potential clinical candidate. The key challenge of poor metabolic stability was overcome by strategic incorporation of deuterium at potential metabolic soft spots. The preclinical profile and status of BMT-052 (14) is described.

The hepatitis C virus (HCV) NS5B replicase is a prime target for the development of direct-acting antiviral drugs for the treatment of chronic HCV infection. Inspired by the overlay of bound structures of three structurally distinct NS5B palm site allosteric inhibitors, the high-throughput screening hit anthranilic acid 4, the known benzofuran analogue 5, and the benzothiadiazine derivative 6, an optimization process utilizing the simple benzofuran template 7 as a starting point for a fragment growing approach was pursued. A delicate balance of molecular properties achieved via disciplined lipophilicity changes was essential to achieve both high affinity binding and a stringent targeted absorption, distribution, metabolism, and excretion profile. These efforts led to the discovery of BMS-929075 (37), which maintained ligand efficiency relative to early leads, demonstrated efficacy in a triple combination regimen in HCV replicon cells, and exhibited consistently high oral bioavailability and pharmacokinetic parameters across preclinical animal species. The human PK properties from the Phase I clinical studies of 37 were better than anticipated and suggest promising potential for QD administration.

This article explores the topic of mediated memory as expressed in the film I Am the Pretty Thing That Lives in the House (2016), chiefly concerned with the challenge of communicating memory from a spectral position. As the film’s narrative is related to audiences through the testimony of the ghost of Lily Saylor, it seems crucial to establish how one might commune with ghosts (or phan- toms). In the first section of the article, I compare the richly mined conception of hauntology as imparted by Jacques Derrida and the recently re-examined precursor to his thought in the psychoanalytic tradition, expressed by Nicolas Abraham and Mária Török. Following a com- parative analysis, I take up both as methods for interpret- ing the film, considering how each might speak to vari- ous hauntological themes in the film which correspond with their accounts (temporal indeterminacy, the role of memory, testimony and the past, trauma and the nature of ghosts/phantoms). A secondary concern of the film, explored through a feminist reading of the film, is the role of women and their confinement and silencing within the domestic. Finally, I consider the question of what is to be done with the specters in the film: should one follow Der- rida’s ethical injunction to make space for the spectral, or ought one exorcize the potentially malevolent influence of the spectral on our lives and break the recurring cycle of transgenerational trauma that it haunts us with?, Grant-Young, Josh. 2020. "'To Look at Oneself and All the While See Nothing': Haunting, Testimony and Mediated Memory in I Am the Pretty Thing That Lives in the House”. Revista de Comunicação e Linguagens, no. 53 (Outono/Inverno): 188- 205. ISSN 2183-7198

[Image: see text] In solving the P-gp and BCRP transporter-mediated efflux issue in a series of benzofuran-derived pan-genotypic palm site inhibitors of the hepatitis C virus NS5B replicase, it was found that close attention to physicochemical properties was essential. In these compounds, where both molecular weight (MW >579) and TPSA (>110 Å(2)) were high, attenuation of polar surface area together with weakening of hydrogen bond acceptor strength of the molecule provided a higher intrinsic membrane permeability and more desirable Caco-2 parameters, as demonstrated by trifluoroacetamide 11 and the benchmark N-ethylamino analog 12. In addition, the tendency of these inhibitors to form intramolecular hydrogen bonds potentially contributes favorably to the improved membrane permeability and absorption. The functional group minimization that resolved the efflux problem simultaneously maintained potent inhibitory activity toward a gt-2 HCV replicon due to a switching of the role of substituents in interacting with the Gln414 binding pocket, as observed in gt-2a NS5B/inhibitor complex cocrystal structures, thus increasing the efficiency of the optimization. Noteworthy, a novel intermolecular S=O···C=O n → π* type interaction between the ligand sulfonamide oxygen atom and the carbonyl moiety of the side chain of Gln414 was observed. The insights from these structure–property studies and crystallography information provided a direction for optimization in a campaign to identify second generation pan-genotypic NS5B inhibitors.

Ataxia telangiectasia mutated (ATM) is a serine/threonine kinase critical to the cellular DNA damage response, including DNA double strand breaks (DSBs). ATM activation results in the initiation of a complex cascade of events facilitating DNA damage repair, cell cycle checkpoint control, and survival. Traditionally, protein kinases have been analyzed in vitro using biochemical methods (kinase assays using purified proteins or immunological assays) requiring a large number of cells and cell lysis. Genetically encoded biosensors based on optical molecular imaging such as fluorescence or bioluminescence have been developed to enable interrogation of kinase activities in live cells with a high signal to background. We have genetically engineered a hybrid protein whose bioluminescent activity is dependent on the ATM-mediated phosphorylation of a substrate. The engineered protein consists of the split luciferase-based protein complementation pair with a CHK2 (a substrate for ATM kinase activity) target sequence and a phospho-serine/threonine-binding domain, FHA2, derived from yeast Rad53. Phosphorylation of the serine residue within the target sequence by ATM would lead to its interaction with the phospho-serine-binding domain, thereby preventing complementation of the split luciferase pair and loss of reporter activity. Bioluminescence imaging of reporter-expressing cells in cultured plates or as mouse xenografts provides a quantitative surrogate for ATM kinase activity and therefore the cellular DNA damage response in a noninvasive, dynamic fashion.

The Ser/Thr kinase BUB1 (budding uninhibited by benzimidazoles-1) is required for efficient signaling by the TGF-β receptor. Recruitment of BUB1 to the activated heteromeric TGF-β-receptor complex including SMAD2 and SMAD3, is required for ligand mediated canonical and non-canonical downstream cascades (Nyati et. al. 2015). In an effort to delineate the structure-function basis for this, we utilized in vitro kinase reactions followed by mass spectrometry to demonstrate that TGFBR2 phosphorylates BUB1 at Serine 318. To elucidate the functional significance of TGFBR2 mediated phosphorylation of BUB1, S318 was substituted with alanine (S318A) or aspartic acid (S318D). Co-immunoprecipitation studies of these phospho-deficient or phospho-mimicking BUB1 mutants with TGFBR1, and SMAD2 revealed that the S318A mutant interacted more efficiently to TGFBR1 and SMAD2 compared to the S318D BUB1. To investigate if the Ser318 residue was involved in mediating BUB1 interaction with TGF-β signaling components, we utilized deletion mutants of BUB1. Our findings reveal that the N-terminal (1-241) and C-terminal (482-723) domains of BUB1 independently interact with SMAD2, SMAD3, TGFBR1, or TGFBR2. The middle domain (241-482) having a S318A mutation interacted efficiently with TGFBR2 while the S318D mutant did not. Additionally, the 241-482 S318D mutant exhibited an increased interaction with SMAD2. Over-expression of 241-482 S318D resulted in a decrease in TGFBR1-TGFBR2 interaction as well as TGFBR1-SMAD2 interaction, suggesting that TGFBR2 mediated BUB1 phosphorylation at S318 maybe the trigger for the dissociation of the activated receptor complex, and therefore a key regulator of TGF-β signaling. Taken together, we show that TGFBR2 phosphorylates BUB1 at Ser318 and propose a model with the following steps: (i) BUB1 is recruited to TGFBR1-TGFBR2 complex in response to ligand, (ii) BUB1 participates in the recruitment of SMAD2/3 to the receptor, (iii) TGFBR2 phosphorylates BUB1 at S318, which triggers the disassembly of the activated complex. Citation Format: Shyam Nyati, Brandon Gregg, Jiaqi Xu, Grant Young, Lauren Kimmel, Nyati Mukesh, Dipankar Ray, Hongtao Yu, Alnawaz Rehemtulla. TGFBR2 mediated phosphorylation of BUB1 at Ser-318 is required for transforming growth factor-β signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3430.

Described herein are structure-activity relationship studies that resulted in the optimization of the activity of members of a class of cyclopropyl-fused indolobenzazepine HCV NS5B polymerase inhibitors. Subsequent iterations of analogue design and syntheses successfully addressed off-target activities, most notably human pregnane X receptor (hPXR) transactivation, and led to significant improvements in the physicochemical properties of lead compounds. Those analogues exhibiting improved solubility and membrane permeability were shown to have notably enhanced pharmacokinetic profiles. Additionally, a series of alkyl bridged piperazine carboxamides was identified as being of particular interest, and from which the compound BMS-791325 (2) was found to have distinguishing antiviral, safety, and pharmacokinetic properties that resulted in its selection for clinical evaluation.

Ataxia telangiectasia mutated (ATM) is a serine/threonine kinase critical to the cellular DNA-damage response, including DNA double-strand breaks (DSBs). ATM activation results in the initiation of a complex cascade of events facilitating DNA damage repair, cell cycle checkpoint control, and survival. Traditionally, protein kinases have been analyzed in vitro using biochemical methods (kinase assays using purified proteins or immunological assays) requiring a large number of cells and cell lysis. Genetically encoded biosensors based on optical molecular imaging such as fluorescence or bioluminescence have been developed to enable interrogation of kinase activities in live cells with a high signal to background. We have genetically engineered a hybrid protein whose bioluminescent activity is dependent on the ATM-mediated phosphorylation of a substrate. The engineered protein consists of the split luciferase-based protein complementation pair with a CHK2 (a substrate for ATM kinase activity) target sequence and a phospho-serine/threonine-binding domain, FHA2, derived from yeast Rad53. Phosphorylation of the serine residue within the target sequence by ATM would lead to its interaction with the phospho-serine-binding domain, thereby preventing complementation of the split luciferase pair and loss of reporter activity. Bioluminescence imaging of reporter expressing cells in cultured plates or as mouse xenografts provides a quantitative surrogate for ATM kinase activity and therefore the cellular DNA damage response in a noninvasive, dynamic fashion.

The development of a series of novel 7-azabenzofurans exhibiting pan-genotype inhibition of HCV NS5B polymerase via binding to the primer grip site is presented. Many challenges, including poor oral bioavailability, high clearance, bioactivation, high human serum shift, and metabolic stability were encountered and overcome through SAR studies. This work culminated in the selection of BMS-986139 (43) as a preclinical candidate.

Grazing livestock excrete large volumes of faecal material on pasture. Understanding the magnitude of this faecal burden is important for attributing sources of agricultural pollutants to different spatial and temporal scales. This field-based study evaluated the utility and transferability of a rapid approach often used in plant ecology (the line intercept method (LIM)) for estimating faecal burden from grazing cattle on pasture. Results from the LIM were of equivalent magnitude, with no significant difference observed, to those derived from more time-consuming sampling of faecal material from pasture using a quadrat-style methodology (herein termed burden sampling). However, the variability in estimates using the LIM was much larger (839–7,079 kg fresh weight faeces over the 50,000 m2 field) compared with estimates provided by the burden sampling of pasture at 0.2 % area sampled (1,616–3,979 kg/50,000 m2), 0.4 % area sampled (1,753–2,723 kg/50,000 m2) and 0.8 % area sampled (1,212–2,344 kg/50,000 m2). The LIM offers a rapid and cost-effective alternative to time-consuming sampling campaigns of faecal burden on pasture and provides estimations that are preferable to back-of-the-envelope calculations based on the over-simplification of livestock excretion rates.

An exploratory SAR study on a series of potent, non-apamin-displacing 4-(aminomethylaryl)pyrazolopyrimidine KCa channel blockers is described and their selectivity against KCa channel subtypes is reported. The most potent analog, 5-chloro-N-(thiophen-2-ylmethyl)pyrazolo[1,5-a]pyrimidin-7-amine (24) displayed sub-micromolar activity in both a thallium flux and whole-cell electrophysiology assay and did not displace apamin in a competitive binding study.

The application of parallel synthesis to lead optimization programs in drug discovery has been an ongoing challenge since the first reports of library synthesis. A number of approaches to the application of parallel array synthesis to lead optimization have been attempted over the years, ranging from widespread deployment by (and support of) individual medicinal chemists to centralization as a service by an expert core team. This manuscript describes our experience with the latter approach, which was undertaken as part of a larger initiative to optimize drug discovery. In particular, we highlight how concepts taken from the manufacturing sector can be applied to drug discovery and parallel synthesis to improve the timeliness and thus the impact of arrays on drug discovery.

The anti-influenza activity of a series of thiobenzamide fusion inhibitors derived from 1,3,3-trimethyl-5-hydroxy-cyclohexylmethylamine is profiled. Axial disposition of the thioamide moiety is essential for potent influenza inhibitory activity.

A novel protocol which employs commercially available, standard tools and hardware has been developed. This protocol allows for cleavage and collection of IRORI Microkan products in 96 well plate format. Typically, 640 compounds can be cleaved in a 4 hour time period using approximately 3 square feet of space. This protocol has been used successfully for the liberation of thousands of individual compounds, in single compound per well format from the solid phase. Additionally, this protocol is the first example of making IRORI Microkan technology directly compatible with standard 96 position deep well blocks.

N-dienyl-5-alkenyl-lactams can be prepared from succinimide. Heating leads to an intramolecular Diels-Alder cyclization that produces tricyclic products useful for the preparation of related alkaloids.

Comprehensive structure activity relationship (SAR) studies were conducted on a focused screening hit, 2-(methylthio)-3-(phenylsulfonyl)-4H-pyrido[1,2-a]pyrimidin-4-imine (1, IC50: 4.0 nM), as 5-HT6 selective antagonists. Activity was improved some 2-4 fold when small, electron-donating groups were added to the central core as observed in 19, 20 and 26. Molecular docking of key compounds in a homology model of the human 5-HT6 receptor was used to rationalize our structure-activity relationship (SAR) findings. In pharmacokinetic experiments, compound 1 displayed good brain uptake in rats following intra-peritoneal administration, but limited oral bioavailability.

The synthesis of novel ligands for the NPY 2 receptor using solid phase split pool methodology is described. One of the analogues, diamine 16 , was found to be a potent NPY 2 binder.

4-Thiazolidinones were synthesized and evaluated for their ability to inhibit the bacterial enzyme MurB. Selected 4-thiazolidinones displayed activity against the enzyme in vitro. This activity, coupled with the design principles of the thiazolidinones, supports the postulate that 4-thiazolidinones may be recognized as diphosphate mimics by a biological selector.

An initial SAR study on a series of apamin-displacing 2-aminothiazole K(Ca)2 channel blockers is described. Potent inhibitors such as N-(4-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine (13) are disclosed, and for select members of the series, the relationship between the observed activity in a thallium flux, a binding and a whole-cell electrophysiology assay is presented.