Discovery of the sEH Inhibitor Epoxykynin as a Potent Kynurenine Pathway Modulator.

Disease-related phenotypic assays enable unbiased discovery of novel bioactive small molecules and may provide novel insights into physiological systems and unprecedented molecular modes of action (MMOA). Herein, we report the identification and characterization of epoxykynin, a potent inhibitor of the soluble epoxide hydrolase (sEH). Epoxykynin was discovered by means of a cellular assay monitoring modulation of kynurenine (Kyn) levels in BxPC-3 cells upon stimulation with the cytokine interferon-γ (IFN-γ) and subsequent target identification employing affinity-based chemical proteomics. Increased Kyn levels are associated with immune suppression in the tumor microenvironment and, thus, the Kyn pathway and its key player indoleamine 2,3-dioxygenase 1 (IDO1) are appealing targets in immuno-oncology. However, targeting IDO1 directly has led to limited success in clinical investigations, demonstrating that alternative approaches to reduce Kyn levels are in high demand. We uncover a cross-talk between sEH and the Kyn pathway that may provide new opportunities to revert cancer-induced immune tolerance.

Transmembrane protein 97 is a potential synaptic amyloid beta receptor in human Alzheimer's disease.

Synapse loss correlates with cognitive decline in Alzheimer's disease, and soluble oligomeric amyloid beta (Aß) is implicated in synaptic dysfunction and loss. An important knowledge gap is the lack of understanding of how Aß leads to synapse degeneration. In particular, there has been difficulty in determining whether there is a synaptic receptor that binds Aß and mediates toxicity. While many candidates have been observed in model systems, their relevance to human AD brain remains unknown. This is in part due to methodological limitations preventing visualization of Aß binding at individual synapses. To overcome this limitation, we combined two high resolution microscopy techniques: array tomography and Förster resonance energy transfer (FRET) to image over 1 million individual synaptic terminals in temporal cortex from AD (n = 11) and control cases (n = 9). Within presynapses and post-synaptic densities, oligomeric Aß generates a FRET signal with transmembrane protein 97. Further, Aß generates a FRET signal with cellular prion protein, and post-synaptic density 95 within post synapses. Transmembrane protein 97 is also present in a higher proportion of post synapses in Alzheimer's brain compared to controls. We inhibited Aß/transmembrane protein 97 interaction in a mouse model of amyloidopathy by treating with the allosteric modulator CT1812. CT1812 drug concentration correlated negatively with synaptic FRET signal between transmembrane protein 97 and Aß. In human-induced pluripotent stem cell derived neurons, transmembrane protein 97 is present in synapses and colocalizes with Aß when neurons are challenged with human Alzheimer's brain homogenate. Transcriptional changes are induced by Aß including changes in genes involved in neurodegeneration and neuroinflammation. CT1812 treatment of these neurons caused changes in gene sets involved in synaptic function. These data support a role for transmembrane protein 97 in the synaptic binding of Aß in human Alzheimer's disease brain where it may mediate synaptotoxicity.

Collective Synthesis of Sarpagine and Macroline Alkaloid-Inspired Compounds.

Design strategies that can access natural-product-like chemical space in an efficient manner may facilitate the discovery of biologically relevant compounds. We have employed a divergent intermediate strategy to construct an indole alkaloid-inspired compound collection derived from two different molecular design principles, i.e. biology-oriented synthesis and pseudo-natural products. The divergent intermediate was subjected to acid-catalyzed or newly discovered Sn-mediated conditions to selectively promote intramolecular C- or N-acylation, respectively. After further derivatization, a collection totalling 84 compounds representing four classes was obtained. Morphological profiling via the cell painting assay coupled with a subprofile analysis showed that compounds derived from different design principles have different bioactivity profiles. The subprofile analysis suggested that a pseudo-natural product class is enriched in modulators of tubulin, and subsequent assays led to the identification of compounds that suppress in vitro tubulin polymerization and mitotic progression.

Apolipoprotein E isoform does not influence trans-synaptic spread of tau pathology in a mouse model.

A key hallmark of Alzheimer's disease (AD) is the accumulation of hyperphosphorylated tau in neurofibrillary tangles. This occurs alongside neuroinflammation and neurodegeneration. Pathological tau propagates through the AD brain in a defined manner, which correlates with neuron and synapse loss and cognitive decline. One proposed mechanism of tau spread is through synaptically connected brain structures. Apolipoprotein E4 (APOE4) genotype is the strongest genetic risk factor for late-onset AD and is associated with increased tau burden. Whether the apolipoprotein E (APOE) genotype influences neurodegeneration via tau spread is currently unknown. Here, we demonstrate that virally expressed human tau (with the P301L mutation) injected into mouse entorhinal cortex at 5-6 months or 15-16 months of age spreads trans-synaptically to the hippocampus by 14 weeks post-injection. Injections of tau in mice expressing human APOE2, APOE3 or APOE4, as well as APOE knock-outs, showed that tau can spread trans-synaptically in all genotypes and that APOE genotype and age do not affect the spread of tau. These data suggest that APOE genotype is not directly linked to synaptic spread of tau in our model, but other mechanisms involving non-cell autonomous manners of tau spread are still possible.

Human astrocytes and microglia show augmented ingestion of synapses in Alzheimer's disease via MFG-E8.

Synapse loss correlates with cognitive decline in Alzheimer's disease (AD). Data from mouse models suggests microglia are important for synapse degeneration, but direct human evidence for any glial involvement in synapse removal in human AD remains to be established. Here we observe astrocytes and microglia from human brains contain greater amounts of synaptic protein in AD compared with non-disease controls, and that proximity to amyloid-ß plaques and the APOE4 risk gene exacerbate this effect. In culture, mouse and human astrocytes and primary mouse and human microglia phagocytose AD patient-derived synapses more than synapses from controls. Inhibiting interactions of MFG-E8 rescues the elevated engulfment of AD synapses by astrocytes and microglia without affecting control synapse uptake. Thus, AD promotes increased synapse ingestion by human glial cells at least in part via an MFG-E8 opsonophagocytic mechanism with potential for targeted therapeutic manipulation.

Circulating tumour cells for early detection of clinically relevant cancer.

Given that cancer mortality is usually a result of late diagnosis, efforts in the field of early detection are paramount to reducing cancer-related deaths and improving patient outcomes. Increasing evidence indicates that metastasis is an early event in patients with aggressive cancers, often occurring even before primary lesions are clinically detectable. Metastases are usually formed from cancer cells that spread to distant non-malignant tissues via the blood circulation, termed circulating tumour cells (CTCs). CTCs have been detected in patients with early stage cancers and, owing to their association with metastasis, might indicate the presence of aggressive disease, thus providing a possible means to expedite diagnosis and treatment initiation for such patients while avoiding overdiagnosis and overtreatment of those with slow-growing, indolent tumours. The utility of CTCs as an early diagnostic tool has been investigated, although further improvements in the efficiency of CTC detection are required. In this Perspective, we discuss the clinical significance of early haematogenous dissemination of cancer cells, the potential of CTCs to facilitate early detection of clinically relevant cancers, and the technological advances that might improve CTC capture and, thus, diagnostic performance in this setting.

Synaptic oligomeric tau in Alzheimer's disease - A potential culprit in the spread of tau pathology through the brain.

In Alzheimer's disease, fibrillar tau pathology accumulates and spreads through the brain and synapses are lost. Evidence from mouse models indicates that tau spreads trans-synaptically from pre- to postsynapses and that oligomeric tau is synaptotoxic, but data on synaptic tau in human brain are scarce. Here we used sub-diffraction-limit microscopy to study synaptic tau accumulation in postmortem temporal and occipital cortices of human Alzheimer's and control donors. Oligomeric tau is present in pre- and postsynaptic terminals, even in areas without abundant fibrillar tau deposition. Furthermore, there is a higher proportion of oligomeric tau compared with phosphorylated or misfolded tau found at synaptic terminals. These data suggest that accumulation of oligomeric tau in synapses is an early event in pathogenesis and that tau pathology may progress through the brain via trans-synaptic spread in human disease. Thus, specifically reducing oligomeric tau at synapses may be a promising therapeutic strategy for Alzheimer's disease.

Unusual phosphatidylcholine lipid phase behavior in the ionic liquid ethylammonium nitrate.

HYPOTHESIS: The forces that govern lipid self-assembly ionic liquids are similar to water, but their different balance can result in unexpected behaviour. EXPERIMENTS: The self-assembly behaviour and phase equilibria of two phospholipids, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), in the most common protic ionic liquid, ethylammonium nitrate (EAN) have been investigated as function of composition and temperature by small- and wide-angle X-ray scattering (SAXS/WAXS) and small-angle neutron scattering (SANS). FINDINGS: Both lipids form unusual self-assembly structures and show complex and unexpected phase behaviour unlike that seen in water; DSPC undergoes a gel Lß to crystalline Lc phase transition on warming, while POPC forms worm-like micelles L1 upon dilution. This surprising phase behaviour is attributed to the large size of the EAN ions that solvate the lipid headgroup compared to water changing amphiphile packing. Weaker H-bonding between EAN and lipid headgroups also contributes. These results provide new insight for the design of lipid based nanostructured materials in ionic liquids with atypical properties.

UK nurses' and midwives' experiences of healthful leadership practices during the COVID-19 pandemic: A rapid realist review.

AIM: We aim to explore healthful leadership practices in nursing and midwifery evident within the COVID-19 pandemic in the United Kingdom, the contextual facilitators, barriers and outcomes. BACKGROUND: Globally, the health and care sectors are under pressure and despite nurses and other professionals, demonstrating resilience and resourcefulness in the COVID-19 pandemic; this has negatively impacted on their health and wellbeing and on patient care. EVALUATION: Two searches were conducted in July 2021 and December 2021. Inclusion/exclusion criteria were identified to refine the search, including papers written since the beginning of the pandemic in 2020. A total of 38 papers were included principally from the United States and United Kingdom. Ten were research papers; the others were commentaries, opinion pieces and editorials. MS Teams literature repository was created. A unique critical appraisal tool was devised to capture contexts, mechanisms and outcomes whilst reflecting more standardized tools, that is, the Critical Appraisal Skills Programme and the Authority, Accuracy, Coverage, Objectivity and Date tool for reviewing grey literature to refine the search further. KEY ISSUES: Six tentative theories of healthful leadership emerged from the literature around leadership strategies, which are relational, being visible and present; being open and engaging; caring for self and others; embodying values; being prepared and preparing others; and using available information and support. Contextual factors that enable healthful leadership practices are in the main, created by leaders' values, attributes and style. The literature suggests that leaders who embody values of compassion, empathy, courage and authenticity create conditions for positive and healthful relations between leaders and others. Nurse and midwives' voices are however absent from the literature in this review. CONCLUSION: Current available literature would suggest healthful leadership practices are not prioritized by nurse leaders. Perspectives of nurses' and midwives' about the impact of such practices on their wellbeing is also missing. Tentative theories are offered as a means of identifying healthful leadership strategies, the context that enable these and potential outcomes for nurses and midwives. These will be explored in phase two of this study. IMPLICATIONS FOR NURSING MANAGEMENT: Nurse leaders must be adequately prepared to create working environments that support nurses' and midwives' wellbeing, so that they may be able to provide high-quality care. Ensuring a supportive organizational culture, which embodies the values of healthfulness, may help to mitigate the impact of the COVID-19 pandemic on nurses' and midwives' wellbeing in the immediate aftermath and going forward.

Identification of a Novel Pseudo-Natural Product Type IV IDO1 Inhibitor Chemotype.

Natural product (NP)-inspired design principles provide invaluable guidance for bioactive compound discovery. Pseudo-natural products (PNPs) are de novo combinations of NP fragments to target biologically relevant chemical space not covered by NPs. We describe the design and synthesis of apoxidoles, a novel pseudo-NP class, whereby indole- and tetrahydropyridine fragments are linked in monopodal connectivity not found in nature. Apoxidoles are efficiently accessible by an enantioselective [4+2] annulation reaction. Biological evaluation revealed that apoxidoles define a new potent type IV inhibitor chemotype of indoleamine 2,3-dioxygenase 1 (IDO1), a heme-containing enzyme considered a target for the treatment of neurodegeneration, autoimmunity and cancer. Apoxidoles target apo-IDO1, prevent heme binding and induce unique amino acid positioning as revealed by crystal structure analysis. Novel type IV apo-IDO1 inhibitors are in high demand, and apoxidoles may provide new opportunities for chemical biology and medicinal chemistry research.

Cysteine string protein alpha accumulates with early pre-synaptic dysfunction in Alzheimer's disease.

In Alzheimer's disease, synapse loss causes memory and cognitive impairment. However, the mechanisms underlying synaptic degeneration in Alzheimer's disease are not well understood. In the hippocampus, alterations in the level of cysteine string protein alpha, a molecular co-chaperone at the pre-synaptic terminal, occur prior to reductions in synaptophysin, suggesting that it is a very sensitive marker of synapse degeneration in Alzheimer's. Here, we identify putative extracellular accumulations of cysteine string alpha protein, which are proximal to beta-amyloid deposits in post-mortem human Alzheimer's brain and in the brain of a transgenic mouse model of Alzheimer's disease. Cysteine string protein alpha, at least some of which is phosphorylated at serine 10, accumulates near the core of beta-amyloid deposits and does not co-localize with hyperphosphorylated tau, dystrophic neurites or glial cells. Using super-resolution microscopy and array tomography, cysteine string protein alpha was found to accumulate to a greater extent than other pre-synaptic proteins and at a comparatively great distance from the plaque core. This indicates that cysteine string protein alpha is most sensitive to being released from pre-synapses at low concentrations of beta-amyloid oligomers. Cysteine string protein alpha accumulations were also evident in other neurodegenerative diseases, including some fronto-temporal lobar dementias and Lewy body diseases, but only in the presence of amyloid plaques. Our findings are consistent with suggestions that pre-synapses are affected early in Alzheimer's disease, and they demonstrate that cysteine string protein alpha is a more sensitive marker for early pre-synaptic dysfunction than traditional synaptic markers. We suggest that cysteine string protein alpha should be used as a pathological marker for early synaptic disruption caused by beta-amyloid.

Reducing voltage-dependent potassium channel Kv3.4 levels ameliorates synapse loss in a mouse model of Alzheimer's disease.

Synapse loss is associated with cognitive decline in Alzheimer's disease, and owing to their plastic nature, synapses are an ideal target for therapeutic intervention. Oligomeric amyloid beta around amyloid plaques is known to contribute to synapse loss in mouse models and is associated with synapse loss in human Alzheimer's disease brain tissue, but the mechanisms leading from Aß to synapse loss remain unclear. Recent data suggest that the fast-activating and -inactivating voltage-gated potassium channel subtype 3.4 (Kv3.4) may play a role in Aß-mediated neurotoxicity. Here, we tested whether this channel could also be involved in Aß synaptotoxicity. Using adeno-associated virus and clustered regularly interspaced short palindromic repeats technology, we reduced Kv3.4 expression in neurons of the somatosensory cortex of APP/PS1 mice. These mice express human familial Alzheimer's disease-associated mutations in amyloid precursor protein and presenilin-1 and develop amyloid plaques and plaque-associated synapse loss similar to that observed in Alzheimer's disease brain. We observe that reducing Kv3.4 levels ameliorates dendritic spine loss and changes spine morphology compared to control virus. In support of translational relevance, Kv3.4 protein was observed in human Alzheimer's disease and control brain and is associated with synapses in human induced pluripotent stem cell-derived cortical neurons. We also noted morphological changes in induced pluripotent stem cell neurones challenged with human Alzheimer's disease-derived brain homogenate containing Aß but, in this in vitro model, total mRNA levels of Kv3.4 were found to be reduced, perhaps as an early compensatory mechanism for Aß-induced damage. Overall, our results suggest that approaches to reduce Kv3.4 expression and/or function in the Alzheimer's disease brain could be protective against Aß-induced synaptic alterations.

Inhibitory synapse loss and accumulation of amyloid beta in inhibitory presynaptic terminals in Alzheimer's disease.

BACKGROUND AND PURPOSE: Synapse degeneration in Alzheimer's disease (AD) correlates strongly with cognitive decline. There is well-established excitatory synapse loss in AD with known contributions of pathological amyloid beta (Aß) to excitatory synapse dysfunction and loss. Despite clear changes in circuit excitability in AD and model systems, relatively little is known about pathology in inhibitory synapses. METHODS: Here human postmortem brain samples (n = 5 control, 10 AD cases) from temporal and occipital cortices were examined to investigate whether inhibitory synapses and neurons are lost in AD and whether Aß may contribute to inhibitory synapse degeneration. Inhibitory neurons were counted in all six cortical layers using stereology software, and array tomography was used to examine synapse density and the accumulation of Aß in synaptic terminals. RESULTS: Differing inhibitory neuron densities were observed in the different cortical layers. The highest inhibitory neuron density was observed in layer 4 in both brain regions and the visual cortex had a higher inhibitory neuron density than the temporal cortex. There was significantly lower inhibitory neuron density in AD than in control cases in all six cortical layers. High-resolution array tomography imaging revealed plaque-associated loss of inhibitory synapses and accumulation of Aß in a small subset of inhibitory presynaptic terminals with the most accumulation near amyloid plaques. CONCLUSIONS: Inhibitory neuron and synapse loss in AD may contribute to disrupted excitatory/inhibitory balance and cognitive decline. Future work is warranted to determine whether targeting inhibitory synapse loss could be a useful therapeutic strategy.

Potential neurobiological links between social isolation and Alzheimer's disease risk.

It is estimated that 40% of dementia cases could be prevented by modification of lifestyle factors that associate with disease risk. One of these potentially modifiable lifestyle factors is social isolation. In this review, we discuss what is known about associations between social isolation and Alzheimer's disease, the most common cause of dementia. This is particularly relevant in the time of the COVID-19 pandemic when social isolation has been enforced with potential emerging negative impacts on cognition. While there are neurobiological mechanisms emerging that may account for the observed epidemiological associations between social isolation and Alzheimer's disease, more fundamental research is needed to fully understand the brain changes induced by isolation that may make people vulnerable to disease.

The potential of using circulating tumour cells and their gene expression to predict docetaxel response in metastatic prostate cancer.

Background: Docetaxel improves overall survival (OS) in castration-resistant prostate cancer (PCa) (CRPC) and metastatic hormone-sensitive PCa (mHSPC). However, not all patients respond due to inherent and/or acquired resistance. There remains an unmet clinical need for a robust predictive test to stratify patients for treatment. Liquid biopsy of circulating tumour cell (CTCs) is minimally invasive, can provide real-time information of the heterogeneous tumour and therefore may be a potentially ideal docetaxel response prediction biomarker. Objective: In this study we investigate the potential of using CTCs and their gene expression to predict post-docetaxel tumour response, OS and progression free survival (PFS). Methods: Peripheral blood was sampled from 18 mCRPC and 43 mHSPC patients, pre-docetaxel treatment, for CTC investigation. CTCs were isolated using the epitope independent Parsortix® system and gene expression was determined by multiplex RT-qPCR. We evaluated CTC measurements for post-docetaxel outcome prediction using receiver operating characteristics and Kaplan Meier analysis. Results: Detection of CTCs pre-docetaxel was associated with poor patient outcome post-docetaxel treatment. Combining total-CTC number with PSA and ALP predicted lack of partial response (PR) with an AUC of 0.90, p= 0.037 in mCRPC. A significantly shorter median OS was seen in mCRPC patients with positive CTC-score (12.80 vs. 37.33 months, HR= 5.08, p= 0.0005), ≥3 total-CTCs/7.5mL (12.80 vs. 37.33 months, HR= 3.84, p= 0.0053), ≥1 epithelial-CTCs/7.5mL (14.30 vs. 37.33 months, HR= 3.89, p= 0.0041) or epithelial to mesenchymal transitioning (EMTing)-CTCs/7.5mL (11.32 vs. 32.37 months, HR= 6.73, p= 0.0001). Significantly shorter PFS was observed in patients with ≥2 epithelial-CTCs/7.5mL (7.52 vs. 18.83 months, HR= 3.93, p= 0.0058). mHSPC patients with ≥5 CTCs/7.5mL had significantly shorter median OS (24.57 vs undefined months, HR= 4.14, p= 0.0097). In mHSPC patients, expression of KLK2, KLK4, ADAMTS1, ZEB1 and SNAI1 was significantly associated with shorter OS and/or PFS. Importantly, combining CTC measurements with clinical biomarkers increased sensitivity and specificity for prediction of patient outcome. Conclusion: While it is clear that CTC numbers and gene expression were prognostic for PCa post-docetaxel treatment, and CTC subtype analysis may have additional value, their potential predictive value for docetaxel chemotherapy response needs to be further investigated in large patient cohorts.

Lowering Synaptogyrin-3 expression rescues Tau-induced memory defects and synaptic loss in the presence of microglial activation.

Tau is a major driver of neurodegeneration and is implicated in over 20 diseases. Tauopathies are characterized by synaptic loss and neuroinflammation, but it is unclear if these pathological events are causally linked. Tau binds to Synaptogyrin-3 on synaptic vesicles. Here, we interfered with this function to determine the role of pathogenic Tau at pre-synaptic terminals. We show that heterozygous knockout of synaptogyrin-3 is benign in mice but strongly rescues mutant Tau-induced defects in long-term synaptic plasticity and working memory. It also significantly rescues the pre- and post-synaptic loss caused by mutant Tau. However, Tau-induced neuroinflammation remains clearly upregulated when we remove the expression of one allele of synaptogyrin-3. Hence neuroinflammation is not sufficient to cause synaptic loss, and these processes are separately induced in response to mutant Tau. In addition, the pre-synaptic defects caused by mutant Tau are enough to drive defects in cognitive tasks.

The impact of chronic daily nicotine exposure and its overnight withdrawal on the structure of anxiety-related behaviors in rats: Role of the lateral habenula.

Tobacco smoking is a serious health problem worldwide and a leading cause of mortality. Nicotine, the addictive component of tobacco, affects a range of emotional responses, including anxiety-related behaviors. Although perceived by smokers to be anxiolytic, evidence suggests that smoking increases anxiety and that mood fluctuates with nicotine intake. Thus, nicotine addiction may depend on easing the psychobiological distress caused by its abuse. The lateral habenula (LHb) has been implicated as a neural substrate for acute nicotine-induced anxiety, but its role in anxiety-like behaviors associated with chronic nicotine exposure has not been explored. Here, we assessed the effect of chronic nicotine exposure and its subsequent overnight withdrawal on anxiety-like behavior using both quantitative and multivariate T-pattern analysis in rats tested using the hole-board apparatus. Additionally, we explored the role of the LHb by comparing the behavioral effects of short-term nicotine withdrawal in chronically treated LHb-lesioned rats. Quantitative analysis revealed increased anxiety-like behavior in chronically treated overnight nicotine-deprived rats, as manifested in reduced general and focused exploratory behaviors, which was eased in animals that received nicotine. Quantitative analysis failed to reveal a role of the LHb in overnight nicotine deprivation-induced anxiety. Conversely, T-pattern analysis of behavioral outcomes revealed that chronic nicotine-treated rats still show anxiety-like behavior following nicotine challenge. Moreover, it demonstrated that the LHb lesion induced a stronger anxiolytic-like response to the acute challenge of nicotine in chronically nicotine-exposed animals, implicating the LHb in the anxiogenic effect of chronic nicotine exposure. These data further highlight the LHb as a promising target for smoking cessation therapies and support the importance of T-pattern analysis for behavioral analysis.

Translational neuroscience: the state of the nation (a PhD student perspective).

Many brain disorders are currently untreatable. It has been suggested that taking a 'translational' approach to neuroscientific research might change this. We discuss what 'translational neuroscience' is and argue for the need to expand the traditional translational model if we are to make further advances in treating brain disorders.