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Among 143 cases of National Healthcare Safety Network (NHSN) catheter-associated urinary tract infections (CAUTI), 40% were considered catheter-associated asymptomatic bacteriuria (CA-ASB), and 18% clinical CAUTI. An alternative source of fever was present in 70% of CA-ASB. NHSN CAUTI may not be an effective metric for tracking hospital-level infection prevention efforts., Competing Interests: None., (© The Author(s) 2024.)

We describe our experience implementing an intensive quality improvement cohort pilot focused on managing asymptomatic bacteriuria in 19 critical access hospitals. Participation in the pilot was high, and almost all sites identified an improvement goal and collected clinical data. Barriers to implementation included staffing shortages, turnover, and lack of bandwidth., (© The Author(s) 2024.)

There are limited control measures for the disease schistosomiasis, despite the fact that infection with parasitic blood flukes affects hundreds of millions of people worldwide. The current treatment, praziquantel, has been in use since the 1980's and there is a concern that drug resistance may emerge with continued monotherapy. Given the need for additional antischistosomal drugs, we have re-visited an old lead, meclonazepam. In comparison to praziquantel, there has been relatively little work on its antiparasitic mechanism. Recent findings indicate that praziquantel and meclonazepam act through distinct receptors, making benzodiazepines a promising chemical series for further exploration. Previous work has profiled the transcriptional changes evoked by praziquantel treatment. Here, we examine in detail schistosome phenotypes evoked by in vitro and in vivo meclonazepam treatment. These data confirm that meclonazepam causes extensive tegument damage and directly kills parasites, as measured by pro-apoptotic caspase activation. In vivo meclonazepam exposure results in differential expression of many genes that are divergent in parasitic flatworms, as well as several gene products implicated in blood feeding and regulation of hemostasis in other parasites. Many of these transcripts are also differentially expressed with praziquantel exposure, which may reflect a common schistosome response to the two drugs. However, despite these similarities in drug response, praziquantel-resistant parasites retain susceptibility to meclonazepam's schistocidal effects. These data provide new insight into the mechanism of antischistosomal benzodiazepines, resolving similarities and differences with the current frontline therapy, praziquantel.

Chimeric antigen receptor (CAR) T cell therapy has transformed the treatment of haematological malignancies such as acute lymphoblastic leukaemia, B cell lymphoma and multiple myeloma 1-4 , but the efficacy of CAR T cell therapy in solid tumours has been limited 5 . This is owing to a number of factors, including the immunosuppressive tumour microenvironment that gives rise to poorly persisting and metabolically dysfunctional T cells. Analysis of anti-CD19 CAR T cells used clinically has shown that positive treatment outcomes are associated with a more 'stem-like' phenotype and increased mitochondrial mass 6-8 . We therefore sought to identify transcription factors that could enhance CAR T cell fitness and efficacy against solid tumours. Here we show that overexpression of FOXO1 promotes a stem-like phenotype in CAR T cells derived from either healthy human donors or patients, which correlates with improved mitochondrial fitness, persistence and therapeutic efficacy in vivo. This work thus reveals an engineering approach to genetically enforce a favourable metabolic phenotype that has high translational potential to improve the efficacy of CAR T cells against solid tumours., (© 2024. The Author(s).)

Schistosomiasis is a disease caused by trematodes of the genus Schistosoma that affects over 200 million people worldwide. For decades, praziquantel (PZQ) has been the only available drug to treat the disease. Despite recent discoveries that identified a transient receptor ion channel as the target of PZQ, schistosome response to this drug remains incompletely understood, since effectiveness relies on other factors that may trigger a complex regulation of parasite gene expression. Long non-coding RNAs (lncRNAs) are transcripts longer than 200 nucleotides with low or no protein-coding potential that play important roles in S. mansoni homeostasis, reproduction, and fertility. Here, we show that in vivo PZQ treatment modulates lncRNA levels in S. mansoni . We re-analyzed public RNA-Seq data from mature and immature S. mansoni worms treated in vivo with PZQ and detected hundreds of lncRNAs differentially expressed following drug exposure, many of which are shared among mature and immature worms. Through RT-qPCR, seven out of ten selected lncRNAs were validated as differentially expressed; interestingly, we show that these lncRNAs are not adult worm stage-specific and are co-expressed with PZQ-modulated protein-coding genes. By demonstrating that parasite lncRNA expression levels alter in response to PZQ, this study unravels an important step toward elucidating the complex mechanisms of S. mansoni response to PZQ.

The ability to provide feedback to a colleague is a key skill required for professional growth and patient safety. However, these conversations are limited by time constraints, differences in values, and a culture of "noninterference." This advocacy-inquiry-identify-teach framework creates an organized approach to initiating successful "challenging" conversations with peers., Competing Interests: The authors report no conflicts of interest relevant to this work., (© The Author(s) 2023.)

CXCL9 expression is a strong predictor of response to immune checkpoint blockade therapy. Accordingly, we sought to develop therapeutic strategies to enhance the expression of CXCL9 and augment antitumor immunity. To perform whole-genome CRISPR-Cas9 screening for regulators of CXCL9 expression, a CXCL9-GFP reporter line is generated using a CRISPR knockin strategy. This approach finds that IRF1 limits CXCL9 expression in both tumor cells and primary myeloid cells through induction of SOCS1, which subsequently limits STAT1 signaling. Thus, we identify a subset of STAT1-dependent genes that do not require IRF1 for their transcription, including CXCL9. Targeting of either IRF1 or SOCS1 potently enhances CXCL9 expression by intratumoral macrophages, which is further enhanced in the context of immune checkpoint blockade therapy. We hence show a non-canonical role for IRF1 in limiting the expression of a subset of STAT1-dependent genes through induction of SOCS1., Competing Interests: Declaration of interests P.A.B. declares the following conflicts: research funding from AstraZeneca, Bristol-Myers-Squibb, and Gilead Sciences. P.K.D. declares the following conflicts: research funding from Myeloid Therapeutics, Prescient Therapeutics, Bristol-Myers-Squibb, and Juno Therapeutics., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Bacterial superinfection and antibiotic prescribing in the setting of the current mpox outbreak are not well described in the literature. This retrospective observational study revealed low prevalence (11%) of outpatient antibiotic prescribing for bacterial superinfection of mpox lesions; at least 3 prescriptions (23%) were unnecessary., Competing Interests: R.J. serves as a clinical consultant for Wolters-Kluwer for their online content. All other authors report no conflicts related to this article., (© The Author(s) 2023.)

Macrocyclic lactones are front-line therapies for parasitic roundworm infections; however, there are no comprehensive studies on the activity of this drug class against parasitic flatworms. Ivermectin is well known to be inactive against flatworms. However, the structure-activity relationship of macrocyclic lactones may vary across phyla, and it is entirely possible other members of this drug class do in fact show antiparasitic activity on flatworms. For example, there are several reports hinting at the anti-schistosomal activity of doramectin and moxidectin. To explore this class further, we developed an automated imaging assay combined with measurement of lactate levels from worm media. This assay was applied to the screening of 21 macrocyclic lactones (avermectins, milbemycins, and others such as spinosyns) against adult schistosomes. These in vitro assays identified several macrocyclic lactones (emamectin, milbemycin oxime, and the moxidectin metabolite 23-ketonemadectin) that caused contractile paralysis and lack of lactate production. Several of these were also active against miracidia, which infect the snail intermediate host. Hits prioritized from these in vitro assays were administered to mice harboring patent schistosome infections. However, no reduction in worm burden was observed. Nevertheless, these data show the utility of a multiplexed in vitro screening platform to quantitatively assess drug action and exclude inactive compounds from a chemical series before proceeding to in vivo studies. While the prototypical macrocyclic lactone ivermectin displays minimal activity against adult Schistosoma mansoni, this family of compounds does contain schistocidal compounds which may serve as a starting point for development of new anti-flatworm chemotherapies.

Development of direct acting macrofilaricides for the treatment of human filariases is hampered by limitations in screening throughput imposed by the parasite life cycle. In vitro adult screens typically assess single phenotypes without prior enrichment for chemicals with antifilarial potential. We developed a multivariate screen that identified dozens of compounds with submicromolar macrofilaricidal activity, achieving a hit rate of >50% by leveraging abundantly accessible microfilariae. Adult assays were multiplexed to thoroughly characterize compound activity across relevant parasite fitness traits, including neuromuscular control, fecundity, metabolism, and viability. Seventeen compounds from a diverse chemogenomic library elicited strong effects on at least one adult trait, with differential potency against microfilariae and adults. Our screen identified five compounds with high potency against adults but low potency or slow-acting microfilaricidal effects, at least one of which acts through a novel mechanism. We show that the use of microfilariae in a primary screen outperforms model nematode developmental assays and virtual screening of protein structures inferred with deep learning. These data provide new leads for drug development, and the high-content and multiplex assays set a new foundation for antifilarial discovery., (© 2023. The Author(s).)

Ligand-gated ion channels (LGICs) are important regulators of neuromuscular function, making them attractive antiparasitic drug targets. While roundworm LGICs are targeted by several anthelmintic classes, flatworm LGICs are less studied. Chromosome-level genome assemblies have recently been released for Schistosoma flatworm species that cause the disease schistosomiasis. These have allowed us to comprehensively predict schistosome LGICs, adding to prior annotations. Analysis of LGIC sequences revealed a clade of receptors lacking cysteines at the eponymous Cys-loop region of the channel. Since these atypical channels are divergent from mammalian LGICs, they may be promising targets to treat diseases caused by parasitic flatworms., (Copyright: © 2023 by the authors.)

Infection with Schistosoma parasitic flatworms ( Schistosoma haematobium, Schistosoma mansoni and Schistosoma japonicum ) causes the neglected tropical disease schistosomiasis. There is a need to identify new chemotherapies to treat these parasites, and G-protein coupled receptors (GPCRs) are a logical druggable targets to explore given they control key aspects of schistosome biology such as neuromuscular function and reproduction. Updated chromosome level genome assemblies for each of the three major species have recently been released. However, studies on these GPCRs require accurate, updated genome annotations. Here, we have re-annotated the GPCRs present in each of the three major schistosome species., (Copyright: © 2023 by the authors.)

Advances in high-throughput and high-content imaging technologies require concomitant development of analytical software capable of handling large datasets and generating relevant phenotypic measurements. Several tools have been developed to analyze drug response phenotypes in parasitic and free-living worms, but these are siloed and often limited to specific instrumentation, worm species, and single phenotypes. No unified tool exists to analyze diverse high-content phenotypic imaging data of worms and provide a platform for future extensibility. We have developed wrmXpress, a unified framework for analyzing a variety of phenotypes matched to high-content experimental assays of free-living and parasitic nematodes and flatworms. We demonstrate its utility for analyzing a suite of phenotypes, including motility, development/size, fecundity, and feeding, and establish the package as a platform upon which to build future custom phenotypic modules. We show that wrmXpress can serve as an analytical workhorse for anthelmintic screening efforts across schistosomes, filarial nematodes, and free-living model nematodes and holds promise for enabling collaboration among investigators with diverse interests., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Wheeler et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Purpose: In this article, we describe a combination chimeric antigen receptor (CAR) T-cell therapy that eradicated the majority of tumors in two immunocompetent murine pancreatic cancer models and a human pancreatic cancer xenograft model., Experimental Design: We used a dual-specific murine CAR T cell that expresses a CAR against the Her2 tumor antigen, and a T-cell receptor (TCR) specific for gp100. As gp100 is also known as pMEL, the dual-specific CAR T cells are thus denoted as CARaMEL cells. A vaccine containing live vaccinia virus coding a gp100 minigene (VV-gp100) was administered to the recipient mice to stimulate CARaMEL cells. The treatment also included the histone deacetylase inhibitor panobinostat (Pano)., Results: The combination treatment enabled significant suppression of Her2 + pancreatic cancers leading to the eradication of the majority of the tumors. Besides inducing cancer cell apoptosis, Pano enhanced CAR T-cell gene accessibility and promoted CAR T-cell differentiation into central memory cells. To test the translational potential of this approach, we established a method to transduce human T cells with an anti-Her2 CAR and a gp100-TCR. The exposure of the human T cells to Pano promoted a T-cell central memory phenotype and the combination treatment of human CARaMEL cells and Pano eradicated human pancreatic cancer xenografts in mice., Conclusions: We propose that patients with pancreatic cancer could be treated using a scheme that contains dual-specific CAR T cells, a vaccine that activates the dual-specific CAR T cells through their TCR, and the administration of Pano., (©2021 American Association for Cancer Research.)

Inflammation early in life can prime the local immune milieu of peripheral tissues, which can cause lasting changes in immunological tone that confer disease protection or susceptibility 1 . The cellular and molecular mechanisms that prompt changes in immune tone in many nonlymphoid tissues remain largely unknown. Here we find that time-limited neonatal inflammation induced by a transient reduction in neonatal regulatory T cells causes a dysregulation of subcutaneous tissue in mouse skin. This is accompanied by the selective accumulation of type 2 helper T (T H 2) cells within a distinct microanatomical niche. T H 2 cells are maintained into adulthood through interactions with a fibroblast population in skin fascia that we refer to as T H 2-interacting fascial fibroblasts (TIFFs), which expand in response to T H 2 cytokines to form subcutaneous fibrous bands. Activation of the T H 2-TIFF niche due to neonatal inflammation primes the skin for altered reparative responses to wounding. Furthermore, we identify fibroblasts in healthy human skin that express the TIFF transcriptional signature and detect these cells at high levels in eosinophilic fasciitis, an orphan disease characterized by inflammation and fibrosis of the skin fascia. Taken together, these data define a previously unidentified T H 2 cell niche in skin and functionally characterize a disease-associated fibroblast population. The results also suggest a mechanism of immunological priming whereby inflammation early in life creates networks between adaptive immune cells and stromal cells to establish an immunological set-point in tissues that is maintained throughout life., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

The function of MR1-restricted mucosal-associated invariant T (MAIT) cells in tumor immunity is unclear. Here we show that MAIT cell-deficient mice have enhanced NK cell-dependent control of metastatic B16F10 tumor growth relative to control mice. Analyses of this interplay in human tumor samples reveal that high expression of a MAIT cell gene signature negatively impacts the prognostic significance of NK cells. Paradoxically, pre-pulsing tumors with MAIT cell antigens, or activating MAIT cells in vivo, enhances anti-tumor immunity in B16F10 and E0771 mouse tumor models, including in the context of established metastasis. These effects are associated with enhanced NK cell responses and increased expression of both IFN-γ-dependent and inflammatory genes in NK cells. Importantly, activated human MAIT cells also promote the function of NK cells isolated from patient tumor samples. Our results thus describe an activation-dependent, MAIT cell-mediated regulation of NK cells, and suggest a potential therapeutic avenue for cancer treatment., (© 2021. Crown.)

Control of the neglected tropical disease schistosomiasis relies almost entirely on praziquantel (PZQ) monotherapy. How PZQ clears parasite infections remains poorly understood. Many studies have examined the effects of PZQ on worms cultured in vitro, observing outcomes such as muscle contraction. However, conditions worms are exposed to in vivo may vary considerably from in vitro experiments given the short half-life of PZQ and the importance of host immune system engagement for drug efficacy in animal models. Here, we investigated the effects of in vivo PZQ exposure on Schistosoma mansoni. Measurement of pro-apoptotic caspase activation revealed that worm death occurs only after parasites shift from the mesenteric vasculature to the liver, peaking 24 hours after drug treatment. This indicates that PZQ is not directly schistocidal, since PZQ's half-life is ~2 hours in humans and ~30 minutes in mice, and focuses attention on parasite interactions with the host immune system following the shift of worms to the liver. RNA-Seq of worms harvested from mouse livers following sub-lethal PZQ treatment revealed drug-evoked changes in the expression of putative immunomodulatory and anticoagulant gene products. Several of these gene products localized to the schistosome esophagus and may be secreted into the host circulation. These include several Kunitz-type protease inhibitors, which are also found in the secretomes of other blood feeding animals. These transcriptional changes may reflect mechanisms of parasite immune-evasion in response to chemotherapy, given the role of complement-mediated attack and the host innate/humoral immune response in parasite elimination. One of these isoforms, SmKI-1, has been shown to exhibit immunomodulatory and anti-coagulant properties. These data provide insight into the effect of in vivo PZQ exposure on S. mansoni, and the transcriptional response of parasites to the stress of chemotherapy., Competing Interests: The authors have declared that no competing interests exist.

Herein is described the strategy to debrominate different aryl bromides selectively, using polymethylhydrosiloxane (PMHS) which tolerates a variety of functional groups. Key elements of this approach include the use of catalytic Pd(OAc) 2 and the correct equivalents of polymethylhydrosiloxane (PMHS), in conjunction with aqueous KF. The present reaction process provides a strategic tool for the synthesis of a number of medicinally important molecules.

Background: Inappropriate testing for Clostridioides difficile leads to overdiagnosis of C difficile infection (CDI). We determined the effect of a computerized clinical decision support (CCDS) order set on C difficile polymerase chain reaction (PCR) test utilization and clinical outcomes., Methods: This study is an interrupted time series analysis comparing C difficile PCR test utilization, hospital-onset CDI (HO-CDI) rates, and clinical outcomes before and after implementation of a CCDS order set at 2 academic medical centers: University of Washington Medical Center (UWMC) and Harborview Medical Center (HMC)., Results: Compared with the 20-month preintervention period, during the 12-month postimplementation of the CCDS order set, there was an immediate and sustained reduction in C difficile PCR test utilization rates at both hospitals (HMC, -28.2% [95% confidence interval {CI}, -43.0% to -9.4%], P  = .005; UWMC, -27.4%, [95% CI, -37.5% to -15.6%], P  < .001). There was a significant reduction in rates of C difficile tests ordered in the setting of laxatives (HMC, -60.8% [95% CI, -74.3% to -40.1%], P  < .001; UWMC, -37.3%, [95% CI, -58.2% to -5.9%], P  = .02). The intervention was associated with an increase in the C difficile test positivity rate at HMC ( P  = .01). There were no significant differences in HO-CDI rates or in the proportion of patients with HO-CDI who developed severe CDI or CDI-associated complications including intensive care unit transfer, extended length of stay, 30-day mortality, and toxic megacolon., Conclusions: Computerized clinical decision support tools can improve C difficile diagnostic test stewardship without causing harm. Additional studies are needed to identify key elements of CCDS tools to further optimize C difficile testing and assess their effect on adverse clinical outcomes., (© The Author(s) 2020. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Objectives: Adoptive transfer of chimeric antigen receptor (CAR)-modified T cells is a form of cancer immunotherapy that has achieved remarkable efficacy in patients with some haematological cancers. However, challenges remain in CAR T-cell treatment of solid tumours because of tumour-mediated immunosuppression., Methods: We have demonstrated that CAR T-cell stimulation through T-cell receptors (TCRs) in vivo can generate durable responses against solid tumours in a variety of murine models. Since Clec9A-targeting tailored nanoemulsion (Clec9A-TNE) vaccine enhances antitumour immune responses through selective activation of Clec9A + cross-presenting dendritic cells (DCs), we hypothesised that Clec9A-TNE could prime DCs for antigen presentation to CAR T cells through TCRs and thus improve CAR T-cell responses against solid tumours. To test this hypothesis, we used CAR T cells expressing transgenic TCRs specific for ovalbumin (OVA) peptides SIINFEKL (CAROTI) or OVA 323-339 (CAROTII)., Results: We demonstrated that the Clec9A-TNEs encapsulating full-length recombinant OVA protein (OVA-Clec9A-TNE) improved CAROT T-cell proliferation and inflammatory cytokine secretion in vitro . Combined treatment using the OVA-Clec9A-TNE and CAROT cells resulted in durable responses and some rejections of tumours in immunocompetent mice. Tumour regression was accompanied by enhanced CAROT cell proliferation and infiltration into the tumours., Conclusion: Our study presents Clec9A-TNE as a prospective avenue to enhance CAR T-cell efficacy for solid cancers., Competing Interests: The authors declare no conflict of interest., (© 2020 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

In this issue of Cancer Cell, Gurusamy et al. use a CRISPR-Cas9 screening approach to demonstrate that deletion of p38 increases multiple phenotypic qualities of effective anti-tumor T cells. Preconditioning T cells with a p38 inhibitor enhances anti-tumor efficacy of adoptive immunotherapy., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Responses of solid tumors to chimeric antigen receptor (CAR) T cell therapy are often minimal. This is potentially due to a lack of sustained activation and proliferation of CAR T cells when encountering antigen in a profoundly immunosuppressive tumor microenvironment. In this study, we investigate if inducing an interaction between CAR T cells and antigen-presenting cells (APCs) in lymphoid tissue, away from an immunosuppressive microenvironment, could enhance solid-tumor responses. We combined CAR T cell transfer with the bacterial enterotoxin staphylococcal enterotoxin-B (SEB), which naturally links a proportion of T cell receptor (TCR) Vβ subtypes to MHC-II, present on APCs. CAR T cell proliferation and function was significantly enhanced by SEB. Solid tumor-growth inhibition in mice was increased when CAR T cells were administered in combination with SEB. CAR T cell expansion in lymphoid tissue was demonstrated, and inhibition of lymphocyte egress from lymph nodes using FTY720 abrogated the benefit of SEB. We also demonstrate that a bispecific antibody, targeting a c-Myc tag on CAR T cells and cluster of differentiation 40 (CD40), could also enhance CAR T cell activity and mediate increased antitumor activity of CAR T cells. These model systems serve as proof-of-principle that facilitating the interaction of CAR T cells with APCs can enhance their ability to mediate antitumor activity., Competing Interests: The authors declare no competing interest.

The anthelmintic drug praziquantel (PZQ) is used to treat schistosomiasis, a neglected tropical disease that affects over 200 million people worldwide. PZQ causes Ca 2+ influx and spastic paralysis of adult worms and rapid vacuolization of the worm surface. However, the mechanism of action of PZQ remains unknown even after 40 years of clinical use. Here, we demonstrate that PZQ activates a schistosome transient receptor potential (TRP) channel, christened Sm TRPM PZQ , present in parasitic schistosomes and other PZQ-sensitive parasites. Several properties of Sm TRPM PZQ were consistent with known effects of PZQ on schistosomes, including (i) nanomolar sensitivity to PZQ; (ii) stereoselectivity toward ( R )-PZQ; (iii) mediation of sustained Ca 2+ signals in response to PZQ; and (iv) a pharmacological profile that mirrors the well-known effects of PZQ on muscle contraction and tegumental disruption. We anticipate that these findings will spur development of novel therapeutic interventions to manage schistosome infections and broader interest in PZQ, which is finally unmasked as a potent flatworm TRP channel activator., (© 2019 Park et al.)

Subversion of parasite neuromuscular function is a key strategy for anthelmintic drug development. Schistosome Ca 2+ signaling has been an area of particular interest for decades, with a specific focus on L-type voltage-gated Ca 2+ channels (Ca v s). However, the study of these channels has been technically challenging. One barrier is the lack of pharmacological probes that are active on flatworms, since the dihydropyridine (DHP) based ligands typically used to study Ca v s are relatively ineffective on schistosomes. Here, we have characterized the effect of a structurally distinct putative L-type Ca v agonist, FPL-64176, on schistosomes cultured ex vivo and in an in vivo murine model of infection. Unlike DHPs, FPL-64176 evokes rapid and sustained contractile paralysis of adult Schistosoma mansoni reminiscent of the anthelmintic praziquantel. This is accompanied by tegument disruption and an arrest of mitotic activity in somatic stem cells and germ line tissues. Interestingly, this strong ex vivo phenotype was temperature dependent, with FPL-64176 treatment being less potent at 37 °C than 23 °C. However, FPL-64176 caused intra-tegument lesions at the basement membrane of worms cultured ex vivo under both conditions, as well as an in vivo hepatic shift of parasites from the mesenteric vasculature of infected mice to the liver. Gene expression profiling of worms harvested following in vivo FPL-64176 exposure reveals differences in transcripts associated with muscle and extracellular matrix function, as well as female reproduction, which is consistent with the worm phenotypes observed following ex vivo drug treatment. These data advance FPL-64176 as a useful tool to study schistosome Ca 2+ signaling, and the benzoyl pyrrole core as a hit compound that may be optimized to develop new parasite-selective leads., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: Investigation of variable response rates to cancer immunotherapies has exposed the immunosuppressive tumor microenvironment (TME) as a limiting factor of therapeutic efficacy. A determinant of TME composition is the tumor location, and clinical data have revealed associations between certain metastatic sites and reduced responses. Preclinical models to study tissue-specific TMEs have eliminated genetic heterogeneity, but have investigated models with limited clinical relevance., Methods: We investigated the TMEs of tumors at clinically relevant sites of metastasis (liver and lungs) and their impact on αPD-1/αCTLA4 and trimAb (αDR5, α4-1BB, αCD40) therapy responses in the 67NR mouse breast cancer and Renca mouse kidney cancer models., Results: Tumors grown in the lungs were resistant to both therapies whereas the same tumor lines growing in the mammary fat pad (MFP), liver or subcutaneously could be completely eradicated, despite greater tumor burden. Assessment of tumor cells and drug delivery in 67NR lung or MFP tumors revealed no differences and prompted investigation into the immune TME. Lung tumors had a more immunosuppressive TME with increased myeloid-derived suppressor cell infiltration, decreased T cell infiltration and activation, and decreased NK cell activation. Depletion of various immune cell subsets indicated an equivalent role for NK cells and CD8 + T cells in lung tumour control. Thus, targeting T cells with αPD-1/αCTLA4 or trimAb was not sufficient to elicit a robust antitumor response in lung tumors., Conclusion: Taken together, these data demonstrate that tissue-specific TMEs influence immunotherapy responses and highlight the importance in defining tissue-specific response patterns in patients., Competing Interests: The authors declare no conflict of interest., (© 2019 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

Parasitic flatworm infections (e.g. tapeworms and fluke worms) are treated by a limited number of drugs. In most cases, control is reliant upon praziquantel (PZQ) monotherapy. However, PZQ is ineffective against sexually immature parasites, and there have also been several concerning reports on cestode and trematode infections with poor PZQ cure-rates, emphasizing the need for alternative therapies to treat these infections. We have revisited a series of benzodiazepines given the anti-schistosomal activity of meclonazepam (MCLZ). MCLZ was discovered in the 1970's but was not brought to market due to dose-limiting sedative side effects. However, in the decades since there have been advances in our understanding of the benzodiazepine GABAA receptor sub-types that drive sedation and the development of sub-type selective, non-sedating ligands. Additionally, the sequencing of flatworm genomes reveals that parasitic trematodes and cestodes have lost GABAAR-like ligand gated anion channels, indicating that MCLZ's anti-parasitic target is distinct from the human receptors that drive sedation. Therefore, we have screened a library of classical and non-sedating 1,4-benzodiazepines against Schistosoma mansoni and identified a series of imidazobenzodiazepines that immobilize worms in vitro. One of these hits, Xhe-II-048 also disrupted the parasite tegument, resulting in extensive vacuole formation beneath the apical membrane. The hit compound series identified has a dramatically lower (~1000×) affinity for the human central benzodiazepine binding site and is a promising starting point for the development of novel anti-schistosomal benzodiazepines with minimal host side-effects., Competing Interests: The authors have declared that no competing interests exist.

We aim to describe the characteristics, risk factors, and clinical outcomes associated with NAP1 strain Clostridioides difficile infection (CDI) in this single-center, retrospective, case-control (1:1) study. We found that the NAP1 strain accounted for 19.7% of CDI, and risk factors for acquisition included residence in skilled nursing facilities, previous CDI, and proton pump inhibitor use., (© The Author(s) 2019. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Conventional treatments for pancreatic cancer are largely ineffective, and the prognosis for the vast majority of patients is poor. Clearly, new treatment options are desperately needed. Immunotherapy offers hope for the development of treatments for pancreatic cancer. A central requirement for the efficacy of this approach is the existence of cancer antigen-specific T cells, but these are often not present or difficult to isolate for most pancreatic tumors. Nevertheless, specific T cells can be generated using genetic modification to express chimeric antigen receptors (CAR), which can enable T cell responses against pancreatic tumor cells. CAR T cells can be produced ex vivo and expanded in vitro for infusion into patients. Remarkable responses have been documented using CAR T cells against several malignancies, including leukemias and lymphomas. Based on these successes, the extension of CAR T cell therapy for pancreatic cancer holds great promise. However, there are a number of challenges that limit the full potential of CAR T cell therapies for pancreatic cancer, including the highly immunosuppressive tumor microenvironment (TME). In this article, we will review the recent progress in using CAR T cells in pancreatic cancer preclinical and clinical settings, discuss hurdles for utilizing the full potential of CAR T cell therapy and propose research strategies and future perspectives. Research into the use of CAR T cell therapy in pancreatic cancer setting is rapidly gaining momentum and understanding strategies to overcome the current challenges in the pancreatic cancer setting will allow the development of effective CAR T cell therapies, either alone or in combination with other treatments to benefit pancreatic cancer patients.

We retrospectively evaluated off-label use of dalbavancin as secondary therapy in 32 patients with serious Staphylococcus aureus infections (endocarditis, osteomyelitis, septic thrombophlebitis, epidural infection) who were also persons who use drugs. The majority of patients (56%) had a clinical response to treatment. Only 1 patient who completed the intended dalbavancin course experienced a treatment failure.

Serotonin (5-HT) is an important regulator of numerous aspects of flatworm biology, ranging from neuromuscular function to sexual maturation and egg laying. In the parasitic blood fluke Schistosoma mansoni, 5-HT targets several G-protein coupled receptors (GPCRs), one of which has been demonstrated to couple to cAMP and regulate parasite movement. This receptor, Sm.5HTR L , has been successfully co-expressed in mammalian cells alongside a luminescent cAMP-biosensor, enabling pharmacological profiling for candidate anti-schistosomal drugs. Here, we have utilized this assay to perform structure-activity investigations of 143 compounds containing previously identified alkaloid natural product pharmacophores (tryptamines, aporphines and protoberberines) shown to regulate Sm.5HTR L . These experiments mapped regions of the tryptamine pharmacophore amenable and intolerant to substitution, highlighting differences relative to orthologous mammalian 5-HT receptors. Potent Sm.5HTR L antagonists were identified, and the efficacy of these compounds were evaluated against live adult parasites cultured ex vivo. Such structure-activity profiling, characterizing the effect of various modifications to these core ring systems on Sm.5HTR L responses, provides greater understanding of pharmacophores selective for this target to aid future drug development efforts., (Copyright © 2018. Published by Elsevier Ltd.)

Conventional approaches for antiparasitic drug discovery center upon discovering selective agents that adversely impact parasites with minimal host side effects. Here, we show that agents with a broad polypharmacology, often considered 'dirtier' drugs, can have unique efficacy if they combine deleterious effects on the parasite with beneficial actions in the host. This principle is evidenced through a screen for drugs to treat schistosomiasis, a parasitic flatworm disease that impacts over 230 million people. A target-based screen of a Schistosoma serotoninergic G protein coupled receptor yielded the potent agonist, ergotamine, which disrupted worm movement. In vivo, ergotamine decreased mortality, parasite load and intestinal egg counts but also uniquely reduced organ pathology through engagement of host GPCRs that repressed hepatic stellate cell activation, inflammatory damage and fibrosis. The unique ability of ergotamine to engage both host and parasite GPCRs evidences a future strategy for anthelmintic drug design that coalesces deleterious antiparasitic activity with beneficial host effects., Competing Interests: JC, TD, JM No competing interests declared, (© 2018, Chan et al.)

Background: Cestodes are a diverse group of parasites, some of them being agents of neglected diseases. In cestodes, little is known about the functional properties of G protein coupled receptors (GPCRs) which have proved to be highly druggable targets in other organisms. Notably, serotoninergic G-protein coupled receptors (5-HT GPCRs) play major roles in key functions like movement, development and reproduction in parasites., Methodology/principal Findings: Three 5-HT GPCRs from Echinococcus granulosus and Mesocestoides corti were cloned, sequenced, bioinformatically analyzed and functionally characterized. Multiple sequence alignment with other GPCRs showed the presence of seven transmembrane segments and conserved motifs but interesting differences were also observed. Phylogenetic analysis grouped these new sequences within the 5-HT7 clade of GPCRs. Molecular modeling showed a striking resemblance in the spatial localization of key residues with their mammalian counterparts. Expression analysis using available RNAseq data showed that both E. granulosus sequences are expressed in larval and adult stages. Localization studies performed in E. granulosus larvae with a fluorescent probe produced a punctiform pattern concentrated in suckers. E. granulosus and M. corti larvae showed an increase in motility in response to serotonin. Heterologous expression revealed elevated levels of cAMP production in response to 5-HT and two of the GPCRs showed extremely high sensitivity to 5-HT (picomolar range). While each of these GPCRs was activated by 5-HT, they exhibit distinct pharmacological properties (5-HT sensitivity, differential responsiveness to ligands)., Conclusions/significance: These data provide the first functional report of GPCRs in parasitic cestodes. The serotoninergic GPCRs characterized here may represent novel druggable targets for antiparasitic intervention.

Schistosomiasis is a debilitating tropical disease caused by infection with parasitic blood flukes. Approximately 260 million people are infected worldwide, underscoring the clinical and socioeconomic impact of this chronic infection. Schistosomiasis is treated with the drug praziquantel (PZQ), which has proved the therapeutic mainstay for over three decades of clinical use. However, the molecular target(s) of PZQ remain undefined. Here we identify a molecular target for the antischistosomal eutomer - (R)-PZQ - which functions as a partial agonist of the human serotoninergic 5HT 2B receptor. (R)-PZQ modulation of serotoninergic signaling occurs over a concentration range sufficient to regulate vascular tone of the mesenteric blood vessels where the adult parasites reside within their host. These data establish (R)-PZQ as a G-protein-coupled receptor ligand and suggest that the efficacy of this clinically important anthelmintic is supported by a broad, cross species polypharmacology with PZQ modulating signaling events in both host and parasite.

The robust regenerative capacity of planarian flatworms depends on the orchestration of signaling events from early wounding responses through the stem cell enacted differentiative outcomes that restore appropriate tissue types. Acute signaling events in excitable cells play an important role in determining regenerative polarity, rationalized by the discovery that sub-epidermal muscle cells express critical patterning genes known to control regenerative outcomes. These data imply a dual conductive (neuromuscular signaling) and instructive (anterior-posterior patterning) role for Ca 2+ signaling in planarian regeneration. Here, to facilitate study of acute signaling events in the excitable cell niche, we provide a de novo transcriptome assembly from the planarian Dugesia japonica allowing characterization of the diverse ionotropic portfolio of this model organism. We demonstrate the utility of this resource by proceeding to characterize the individual role of each of the planarian voltage-operated Ca 2+ channels during regeneration, and demonstrate that knockdown of a specific voltage operated Ca 2+ channel (Ca v 1B) that impairs muscle function uniquely creates an environment permissive for anteriorization. Provision of the full transcriptomic dataset should facilitate further investigations of molecules within the planarian voltage-gated channel portfolio to explore the role of excitable cell physiology on regenerative outcomes. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The diversity and uniqueness of flatworm G protein coupled receptors (GPCRs) provides impetus for identifying ligands useful as tools for studying flatworm biology, or as therapeutics for treating diseases caused by parasitic flatworm infections. To catalyse this discovery process, technologies optimized for mammalian GPCR high throughput screening need be transposed for screening flatworm GPCRs. Here, we demonstrate the utility of a genetically encoded cAMP biosensor for resolving the properties of an abundantly expressed planarian serotonergic GPCR (S7.1R). Application of this methodology resolved the real time kinetics of GPCR modulation by ligands and demonstrated a marked difference in the kinetic action of antagonists at S7.1R. Notably, bromocriptine caused a protracted inhibition of S7.1R activity in vitro and a protracted paralysis of planarian movement, replicating the effect of S7.1R in vivo RNAi. The lengthy inhibition of function caused by bromocriptine at this abundantly expressed GPCR provides a useful tool to ablate serotonergic signaling in vivo, and is a noteworthy feature for exploitation as an anthelmintic vulnerability., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

5-hydroxytryptamine (5-HT) is a key regulator of muscle contraction in parasitic flatworms. In Schistosoma mansoni, the myoexcitatory action of 5-HT is effected through activation of a serotonergic GPCR (Sm.5HTR L ), prioritizing pharmacological characterization of this target for anthelmintic drug discovery. Here, we have examined the effects of several aporphine alkaloids on the signaling activity of a heterologously expressed Sm.5HTR L construct using a cAMP biosensor assay. Four structurally related natural products - nuciferine, D-glaucine, boldine and bulbocapnine - were demonstrated to block Sm.5HTR L evoked cAMP generation with the potency of GPCR blockade correlating well with the ability of each drug to inhibit contractility of schistosomule larvae. Nuciferine was also effective at inhibiting both basal and 5-HT evoked motility of adult schistosomes. These data advance our understanding of structure-affinity relationships at Sm.5HTR L , and demonstrate the effectiveness of Sm.5HTR L antagonists as hypomotility-evoking drugs across different parasite life cycle stages., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

This data article provides a transcriptomic resource for the free living planarian flatworm Dugesia japonica related to the research article entitled 'Utilizing the planarian voltage-gated ion channel transcriptome to resolve a role for a Ca 2+ channel in neuromuscular function and regeneration (J.D. Chan, D. Zhang, X. Liu, M. Zarowiecki, M. Berriman, J.S. Marchant, 2016) [1]. Data provided in this submission comprise sequence information for the unfiltered de novo assembly, the filtered assembly and a curated analysis of voltage-gated like (VGL) ion channel sequences mined from this resource. Availability of this data should facilitate further adoption of this model by laboratories interested in studying the role of individual genes of interest in planarian physiology and regenerative biology.

We present a 71-year old woman treated with 14 days of 5 mg intraventricular caspofungin for Scedosporium apiospermum complex meningoencephalitis diagnosed after spinal fusion and instrumentation. Cerebrospinal fluid studies improved during therapy and intraventricular administration was well tolerated. Within weeks of discontinuation, the patient experienced clinical deterioration with disease progression. There are sparse data on the efficacy and safety of administering intraventricular caspofungin. While apparently safe, intraventricular caspofungin was insufficient for disease control in this case.

Schistosomiasis is a tropical parasitic disease afflicting ~200 million people worldwide and current therapy depends on a single drug (praziquantel) which exhibits several non-optimal features. These shortcomings underpin the need for next generation anthelmintics, but the process of validating physiologically relevant targets ('target selection') and pharmacologically profiling them is challenging. Remarkably, even though over a quarter of current human therapeutics target rhodopsin-like G protein coupled receptors (GPCRs), no library screen of a flatworm GPCR has yet been reported. Here, we have pharmacologically profiled a schistosome serotonergic GPCR (Sm.5HTR) implicated as a downstream modulator of PZQ efficacy, in a miniaturized screening assay compatible with high content screening. This approach employs a split luciferase based biosensor sensitive to cellular cAMP levels that resolves the proximal kinetics of GPCR modulation in intact cells. Data evidence a divergent pharmacological signature between the parasitic serotonergic receptor and the closest human GPCR homolog (Hs.5HTR7), supporting the feasibility of optimizing parasitic selective pharmacophores. New ligands, and chemical series, with potency and selectivity for Sm.5HTR over Hs.5HTR7 are identified in vitro and validated for in vivo efficacy against schistosomules and adult worms. Sm.5HTR also displayed a property resembling irreversible inactivation, a phenomenon discovered at Hs.5HTR7, which enhances the appeal of this abundantly expressed parasite GPCR as a target for anthelmintic ligand design. Overall, these data underscore the feasibility of profiling flatworm GPCRs in a high throughput screening format competent to resolve different classes of GPCR modulators. Further, these data underscore the promise of Sm.5HTR as a chemotherapeutically vulnerable node for development of next generation anthelmintics.

Praziquantel (PZQ) is a key therapy for treatment of parasitic flatworm infections of humans and livestock, but the mechanism of action of this drug is unresolved. Resolving PZQ-engaged targets and effectors is important for identifying new druggable pathways that may yield novel antiparasitic agents. Here we use functional, genetic and pharmacological approaches to reveal that serotonergic signals antagonize PZQ action in vivo. Exogenous 5-hydroxytryptamine (5-HT) rescued PZQ-evoked polarity and mobility defects in free-living planarian flatworms. In contrast, knockdown of a prevalently expressed planarian 5-HT receptor potentiated or phenocopied PZQ action in different functional assays. Subsequent screening of serotonergic ligands revealed that several ergot alkaloids possessed broad efficacy at modulating regenerative outcomes and the mobility of both free living and parasitic flatworms. Ergot alkaloids that phenocopied PZQ in regenerative assays to cause bipolar regeneration exhibited structural modifications consistent with serotonergic blockade. These data suggest that serotonergic activation blocks PZQ action in vivo, while serotonergic antagonists phenocopy PZQ action. Importantly these studies identify the ergot alkaloid scaffold as a promising structural framework for designing potent agents targeting parasitic bioaminergic G protein coupled receptors.

Stem cells hold great promise in the treatment of diseases ranging from cancer to dementia. However, as rapidly as the field of stem cell biology has emerged, heated political debate has followed, scrutinizing the ethical implications of stem cell use. It is therefore imperative to promote scientific literacy by educating students about stem cell biology. Yet, there is a definite lack of material to engage students in this subject at the basic science level. Therefore, we have developed and implemented a hands-on introductory laboratory module that introduces students to stem cell biology and can be easily incorporated into existing curricula. Students learn about stem cell biology using an in vivo planarian model system in which they down-regulate two genes important in stem cell differentiation using RNA interference and then observe the regenerative phenotype. The module was piloted at the high school, community college, and university levels. Here, we report that introductory biology students enrolled at a community college were able to demonstrate gains in learning after completion of a one-hour lecture and four 45-minute laboratory sessions over the course of three weeks. These gains in learning outcomes were objectively evaluated both before and after its execution using a student quiz and experimental results. Furthermore, students' self-assessments revealed increases in perceived knowledge as well as a general interest in stem cells. Therefore, these data suggest that this module is a simple, useful way to engage and to teach students about stem cell biology.