Structure-Guided Design and Synthesis of a Pyridazinone Series of Trypanosoma cruzi Proteasome Inhibitors.

There is an urgent need for new treatments for Chagas disease, a parasitic infection which mostly impacts South and Central America. We previously reported on the discovery of GSK3494245/DDD01305143, a preclinical candidate for visceral leishmaniasis which acted through inhibition of the Leishmania proteasome. A related analogue, active against Trypanosoma cruzi, showed suboptimal efficacy in an animal model of Chagas disease, so alternative proteasome inhibitors were investigated. Screening a library of phenotypically active analogues against the T. cruzi proteasome identified an active, selective pyridazinone, the development of which is described herein. We obtained a cryo-EM co-structure of proteasome and a key inhibitor and used this to drive optimization of the compounds. Alongside this, optimization of the absorption, distribution, metabolism, and excretion (ADME) properties afforded a suitable compound for mouse efficacy studies. The outcome of these studies is discussed, alongside future plans to further understand the series and its potential to deliver a new treatment for Chagas disease.

Potent antiplasmodial alkaloids from the rhizobacterium Pantoea agglomerans as hemozoin modulators.

Global health concern regarding malaria has increased since the first report of artemisinin-resistant Plasmodium falciparum (Pf) two decades ago. The current therapies suffer various drawbacks such as low efficacy and significant side effects, alarming for an urgent need of more effective and less toxic drugs with higher patient compliance. Chemical entities with natural origins become progressively attractive as new drug leads due to their structural diversity and bio-compatibility. This study initially aimed at the targeted isolation of hydroxyquinoline derivatives following our published genomics and metabolomics study of Pantoea agglomerans (Pa). Fermentation of Pa on a pre-selected medium followed by chromatographic isolation, NMR and HRMS analyses led to the characterisation of one new hydroxyquinoline alkaloid together with another six known congeners and two known hydroxyquinolone derivatives. When screened for their antimalarial activity by high throughput screening against asexual blood-stage parasites, almost all compounds showed potent and selective sub-micromolar activities. Computational investigation was performed to identify the antiplasmodial potential targets. Ligand-based similarity search predicted the tested compounds to act as hemozoin inhibitors. Computational target identification results were further validated by competitive hemozoin inhibitory properties of hydroxyquinoline and hydroxyquinolone derivatives in vitro. The overall results suggest this natural scaffold is of potential to be developed as antimalarial drug lead.

Impact of a Rapid-Access Ambulatory Psychiatry Encounter on Subsequent Emergency Department Utilization.

The authors sought to determine whether providing a rapid-access ambulatory psychiatry encounter correlated with emergency department utilization during a 6-month follow-up period. Electronic medical records of patients who accessed ambulatory psychiatric care through an urgent care psychiatry clinic that offers treatment exclusively on a walk-in basis over a 1-year period (N = 157) were reviewed retrospectively to track emergency department encounters with and without a psychiatric chief complaint in the 6 months before and after the initial psychiatry evaluation. Among patients who had not previously received ambulatory psychiatric care (N = 88), emergency department utilization decreased from 0.68 visits per patient to 0.36, and this difference was statistically significant (p = 0.0147). No statistically significant differences were found between the average number of emergency department encounters in the 6 months before and after the rapid-access ambulatory psychiatry encounter, regardless of chief complaint, when all patients were included in the analysis. Providing a rapid-access ambulatory psychiatry encounter may reduce subsequent emergency department utilization among patients who have not previously received ambulatory psychiatric care.

Identification of 6-amino-1H-pyrazolo[3,4-d]pyrimidines with in vivo efficacy against visceral leishmaniasis.

Visceral leishmaniasis (VL) affects millions of people across the world, largely in developing nations. It is fatal if left untreated and the current treatments are inadequate. As such, there is an urgent need for new, improved medicines. In this paper, we describe the identification of a 6-amino-N-(piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidine scaffold and its optimization to give compounds which showed efficacy when orally dosed in a mouse model of VL.

Longitudinal Urgent Care Psychiatry as a Unique Access Point for Underserved Patients.

OBJECTIVE: The authors sought to determine whether a walk-in psychiatry model with longitudinal follow-up capability could improve access for patients who traditionally miss appointments. METHODS: An urgent care clinic that offers treatment exclusively on a walk-in basis was opened within an adult psychiatry practice to accommodate patients who missed prior scheduled appointments. Electronic health records for patients who received an initial psychiatry evaluation at the practice during a 6-month period (N=355) were reviewed retrospectively to track the clinic's productivity and patient demographic characteristics. RESULTS: Eighty patients (23%) accessed their initial psychiatry encounters through the walk-in clinic. Medicaid recipients (odds ratio [OR]=1.89, 95% confidence interval [CI]=1.10-3.24) and individuals without a college degree (OR=1.86, 95% CI=1.04-3.32) were more likely than patients with other insurance carriers and those with a college degree, respectively, to access care through a walk-in encounter versus a scheduled appointment. CONCLUSIONS: Longitudinal walk-in psychiatry services can feasibly be offered through the longitudinal urgent care psychiatry model. This model may serve as a unique access point for patients from historically underserved groups.

Identification of GSK3186899/DDD853651 as a Preclinical Development Candidate for the Treatment of Visceral Leishmaniasis.

The leishmaniases are diseases that affect millions of people across the world, in particular visceral leishmaniasis (VL) which is fatal unless treated. Current standard of care for VL suffers from multiple issues and there is a limited pipeline of new candidate drugs. As such, there is a clear unmet medical need to identify new treatments. This paper describes the optimization of a phenotypic hit against Leishmania donovani, the major causative organism of VL. The key challenges were to balance solubility and metabolic stability while maintaining potency. Herein, strategies to address these shortcomings and enhance efficacy are discussed, culminating in the discovery of preclinical development candidate GSK3186899/DDD853651 (1) for VL.

The Relationship between NALP3 and Autoinflammatory Syndromes.

The nucleotide-binding domain, leucine-rich repeat/pyrin domain-containing-3 (NALP3) inflammasome, which is required for synthesis of interleukin-1ß, has been implicated in the pathogenesis of several autoinflammatory syndromes. This review of the literature summarizes the interconnectedness of NALP3 inflammasome with some of these disorders. Familial Mediterranean fever results from a mutation in the Mediterranean fever (MEFV) gene, which encodes the pyrin protein. Previous study results suggest that pyrin suppresses caspase-1 activation, perhaps by competing for the adaptor protein, termed, pyrin domain of apoptosis/speck-like protein containing a caspase-recruitment domain (ACS) which therefore interferes with NALP3 inflammasome activation. The nucleotide-binding domain, leucine-rich repeat/pyrin domain-containing-3 (NALP3) inflammasome is constitutively activated in cryopyrin-associated periodic syndromes due to gain-of-function mutations resulting from point mutations within the neuronal apoptosis inhibitor protein/class 2 transcription factor/heterokaryon incompatibility/telomerase-associated protein-1 (NACHT) domain of the NALP3 protein. Pyogenic arthritis, pyoderma gangrenosum and acne (PAPA) syndrome is caused by mutations in the genes encoding proline-serine-threonine phosphatase interacting protein 1 (PSTPIP1). These PSTPIP1 mutants are thought to bind to pyrin causing an increase in the pyrin domain of apoptosis/speck-like protein containing a caspase-recruitment domain (ASC) pyroptosome assembly leading to procaspase-1 recruitment and therefore its activation. Hyperimmunoglublinemia D syndrome is caused by mevalonate kinase (MVK) deficiency, which may be affected by protein accumulation that leads to NALP3 inflammasome activation. Tumor necrosis factor receptor-associated periodic syndrome is associated with mutations in the tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A) gene which decreases the level of soluble tumor necrosis factor receptor-1 (TNFR1) leading to neutralization of tumor necrosis factor (TNF)-α. In general, these autoinflammatory disorders have shown a clinical response to interleukin-1 (IL-1) antagonists, suggesting that the NALP3 inflammasome serves a critical role in their pathogenesis.

Discovery of Indoline-2-carboxamide Derivatives as a New Class of Brain-Penetrant Inhibitors of Trypanosoma brucei.

There is an urgent need for new, brain penetrant small molecules that target the central nervous system second stage of human African trypanosomiasis (HAT). We report that a series of novel indoline-2-carboxamides have been identified as inhibitors of Trypanosoma brucei from screening of a focused protease library against Trypanosoma brucei brucei in culture. We describe the optimization and characterization of this series. Potent antiproliferative activity was observed. The series demonstrated excellent pharmacokinetic properties, full cures in a stage 1 mouse model of HAT, and a partial cure in a stage 2 mouse model of HAT. Lack of tolerability prevented delivery of a fully curative regimen in the stage 2 mouse model and thus further progress of this series.

Comparison of a high-throughput high-content intracellular Leishmania donovani assay with an axenic amastigote assay.

Visceral leishmaniasis is a neglected tropical disease with significant health impact. The current treatments are poor, and there is an urgent need to develop new drugs. Primary screening assays used for drug discovery campaigns have typically used free-living forms of the Leishmania parasite to allow for high-throughput screening. Such screens do not necessarily reflect the physiological situation, as the disease-causing stage of the parasite resides inside human host cells. Assessing the drug sensitivity of intracellular parasites on scale has recently become feasible with the advent of high-content screening methods. We describe here a 384-well microscopy-based intramacrophage Leishmania donovani assay and compare it to an axenic amastigote system. A panel of eight reference compounds was tested in both systems, as well as a human counterscreen cell line, and our findings show that for most clinically used compounds both axenic and intramacrophage assays report very similar results. A set of 15,659 diverse compounds was also screened using both systems. This resulted in the identification of seven new antileishmanial compounds and revealed a high false-positive rate for the axenic assay. We conclude that the intramacrophage assay is more suited as a primary hit-discovery platform than the current form of axenic assay, and we discuss how modifications to the axenic assay may render it more suitable for hit-discovery.

A static-cidal assay for Trypanosoma brucei to aid hit prioritisation for progression into drug discovery programmes.

Human African Trypanosomiasis is a vector-borne disease of sub-Saharan Africa that causes significant morbidity and mortality. Current therapies have many drawbacks, and there is an urgent need for new, better medicines. Ideally such new treatments should be fast-acting cidal agents that cure the disease in as few doses as possible. Screening assays used for hit-discovery campaigns often do not distinguish cytocidal from cytostatic compounds and further detailed follow-up experiments are required. Such studies usually do not have the throughput required to test the large numbers of hits produced in a primary high-throughput screen. Here, we present a 384-well assay that is compatible with high-throughput screening and provides an initial indication of the cidal nature of a compound. The assay produces growth curves at ten compound concentrations by assessing trypanosome counts at 4, 24 and 48 hours after compound addition. A reduction in trypanosome counts over time is used as a marker for cidal activity. The lowest concentration at which cell killing is seen is a quantitative measure for the cidal activity of the compound. We show that the assay can identify compounds that have trypanostatic activity rather than cidal activity, and importantly, that results from primary high-throughput assays can overestimate the potency of compounds significantly. This is due to biphasic growth inhibition, which remains hidden at low starting cell densities and is revealed in our static-cidal assay. The assay presented here provides an important tool to follow-up hits from high-throughput screening campaigns and avoid progression of compounds that have poor prospects due to lack of cidal activity or overestimated potency.

Comparison of conventional and nonconventional strictureplasties in Crohn's disease: a systematic review and meta-analysis.

BACKGROUND: The Heineke-Mikulicz and Finney techniques are conventional strictureplasties that have been used to manage short (<10 cm) and medium-length (>10 cm and <20 cm) strictures from Crohn's disease. Nonconventional strictureplasty techniques have emerged to facilitate bowel conservation for atypical strictures. These techniques include the modified Finney, combined Heineke-Mikulicz and Finney, modified Heineke-Mikuliczs, Michelassi, and modifications of it and others. OBJECTIVE: The aim of this study is to compare conventional vs nonconventional strictureplasties with respect to short-term complications and long-term results. DATA SOURCES AND STUDY SELECTION: A MEDLINE search was performed using "Crohn's disease", "surgical therapy", "strictureplasty", "complications", "reoperation", and "recurrence" as medical subject headings. Studies conducted between 1975 and June 31, 2010 were found via PubMed, Ovid, Embase, and Cochrane databases and categorized into 3 groups. These groups consist of centers performing conventional strictureplasties, nonconventional strictureplasties, or both. Studies with at least 3 patients were reviewed. INTERVENTIONS: A mixed-effects meta-analysis for each outcome was performed by use of Supermix software by SSI Scientific Software International. MAIN OUTCOME MEASURES: We focused on immediate and long-term complication rates among the groups. The 6 immediate complications include small-bowel obstructions, sepsis, other infections, reoperations, early postoperative GI bleeds, and other early complications. The 5 long-term complications include recurrent strictures, small-bowel obstructions, reoperations, carcinoma, and deaths. RESULTS: We reviewed 32 studies with 1616 patients who underwent 4538 strictureplasties. One thousand one hundred fifty-seven patients underwent conventional strictureplasties with an early complication rate of 15%; 459 patients underwent nonconventional strictureplasties with an early complication rate of 8%. A late complication rate of 29% for the conventional strictureplasty group and 17% for the nonconventional strictureplasty group was noted. LIMITATIONS: We are limited by the data published with the inherent risk of finding and analyzing mostly articles with positive results. CONCLUSION: The nonconventional strictureplasty techniques were noninferior to the conventional strictureplasty procedures with respect to all prespecified outcomes.

A comprehensive review of strictureplasty techniques in Crohn's disease: types, indications, comparisons, and safety.

INTRODUCTION: Crohn's disease is one of the chronic inflammatory diseases of the gastrointestinal tract that is often complicated by stricture formation with resulting obstructive symptoms. The technical repertoire of strictureplasty procedures has increased over the years in an effort to manage the diverse presentations of this condition while limiting the need for bowel resection. In this comprehensive review, we describe, compare, categorize, and appraise the strengths and weaknesses of 15 unique strictureplasty techniques. METHODS: To identify all unique strictureplasty procedures, a Medline search utilizing "Crohn's disease," "surgical therapy," "strictureplasty," "enteroenterostomy," "Heineke-Mikulicz," and "side-to-side isoperistaltic" strictureplasty as medical subject headings was completed. PubMed, Ovid, Embase, and Cochrane database searches were conducted. Relevant articles between 1980 to December 2010 were reviewed. We initially selected 58 articles, but only 18 introduced novel surgical procedures related to 15 types of strictureplasty in Crohn's disease. RESULTS: We identified 15 types of strictureplasty techniques. These were categorized into three main groups. The revised nomenclature will facilitate the reader to understand the differences and utility of each technique. These groups include the Heineke-Mikulicz-like strictureplasties, the intermediate procedures, and the enteroenterostomies. Heineke-Mikulicz strictureplasty was the most frequently used technique. CONCLUSION: Various techniques of strictureplasty have been reported in the published literature. Strictureplasty has been shown to be a safe and efficacious technique that is comparable to bowel resection for stricturing Crohn's disease. This technique spares bowel length and puts the Crohn's disease patient at a lower risk of developing short bowel syndrome with repeated resections.

The extent of working memory deficits associated with Williams syndrome: exploration of verbal and spatial domains and executively controlled processes.

The present study investigated verbal and spatial working memory (WM) functioning in individuals with the neuro-developmental disorder Williams syndrome (WS) using WM component tasks. While there is strong evidence of WM impairments in WS, previous research has focused on short-term memory and has neglected assessment of executive components of WM. There is a particular lack of consensus concerning the profile of verbal WM functioning in WS. Here, WS participants were compared to typically developing participants matched for (1) verbal ability and (2) spatial ability (N=14 in each of the 3 groups). Individuals with WS were impaired on verbal WM tasks, both those involving short-term maintenance of information and executive manipulation, in comparison to verbal-matched controls. Surprisingly, individuals with WS were not impaired on a spatial task assessing short-term maintenance of information in memory (remembering spatial locations) compared to spatial-matched controls. They were, however, impaired on a spatial executive WM task requiring the manipulation of spatial information in memory. The present study suggests that individuals with WS show WM impairments that extend to both verbal and spatial domains, although spatial deficits are selective to executive aspects of WM function.

Executive neuropsychological functioning in individuals with Williams syndrome.

The present study investigated executive neuropsychological functioning in individuals with the neuro-developmental disorder Williams syndrome (WS) using a set of validated standardized neuropsychological tasks. Relatively few studies have examined frontal lobe related executive functions within the cognitive phenotype associated with the disorder. The present study compared participants with WS to typically developing participants who were individually matched for (1) chronological age and (2) verbal mental age (N=19 each group) on tasks of attention-set shifting, planning and working memory from the Cambridge Neuropsychological Test Automated Battery (CANTAB). To address the specificity of executive function impairment, non-executive tasks of delayed short-term memory and short-term memory span were also administered. Individuals with WS (mean age 18 years) showed impaired executive functioning on tasks of attention set-shifting, working memory, and planning. Non-executive deficits were also observed in short-term delayed memory and memory span. Neuropsychological impairments were correlated with a range of behavioural problems assessed using parent-rated Questionnaires. Overall, these findings point to the role of a range of executive function impairments in WS but further suggest that cognitive impairments extend beyond executive dysfunction.

Small molecule antagonists of the sigma-1 receptor cause selective release of the death program in tumor and self-reliant cells and inhibit tumor growth in vitro and in vivo.

The acquisition of resistance to apoptosis, the cell's intrinsic suicide program, is essential for cancers to arise and progress and is a major reason behind treatment failures. We show in this article that small molecule antagonists of the sigma-1 receptor inhibit tumor cell survival to reveal caspase-dependent apoptosis. sigma antagonist-mediated caspase activation and cell death are substantially attenuated by the prototypic sigma-1 agonists (+)-SKF10,047 and (+)-pentazocine. Although several normal cell types such as fibroblasts, epithelial cells, and even sigma receptor-rich neurons are resistant to the apoptotic effects of sigma antagonists, cells that can promote autocrine survival such as lens epithelial and microvascular endothelial cells are as susceptible as tumor cells. Cellular susceptibility appears to correlate with differences in sigma receptor coupling rather than levels of expression. In susceptible cells only, sigma antagonists evoke a rapid rise in cytosolic calcium that is inhibited by sigma-1 agonists. In at least some tumor cells, sigma antagonists cause calcium-dependent activation of phospholipase C and concomitant calcium-independent inhibition of phosphatidylinositol 3'-kinase pathway signaling. Systemic administration of sigma antagonists significantly inhibits the growth of evolving and established hormone-sensitive and hormone-insensitive mammary carcinoma xenografts, orthotopic prostate tumors, and p53-null lung carcinoma xenografts in immunocompromised mice in the absence of side effects. Release of a sigma receptor-mediated brake on apoptosis may offer a new approach to cancer treatment.

Mapping and identification of essential gene functions on the X chromosome of Drosophila.

The Drosophila melanogaster genome consists of four chromosomes that contain 165 Mb of DNA, 120 Mb of which are euchromatic. The two Drosophila Genome Projects, in collaboration with Celera Genomics Systems, have sequenced the genome, complementing the previously established physical and genetic maps. In addition, the Berkeley Drosophila Genome Project has undertaken large-scale functional analysis based on mutagenesis by transposable P element insertions into autosomes. Here, we present a large-scale P element insertion screen for vital gene functions and a BAC tiling map for the X chromosome. A collection of 501 X-chromosomal P element insertion lines was used to map essential genes cytogenetically and to establish short sequence tags (STSs) linking the insertion sites to the genome. The distribution of the P element integration sites, the identified genes and transcription units as well as the expression patterns of the P-element-tagged enhancers is described and discussed.