SMYD5 methylation of rpL40 links ribosomal output to gastric cancer.

Dysregulated transcription due to disruption in histone lysine methylation dynamics is an established contributor to tumorigenesis1,2. However, whether analogous pathologic epigenetic mechanisms act directly on the ribosome to advance oncogenesis is unclear. Here we find that trimethylation of the core ribosomal protein L40 (rpL40) at lysine 22 (rpL40K22me3) by the lysine methyltransferase SMYD5 regulates mRNA translation output to promote malignant progression of gastric adenocarcinoma (GAC) with lethal peritoneal ascites. A biochemical-proteomics strategy identifies the monoubiquitin fusion protein partner rpL40 (ref. 3) as the principal physiological substrate of SMYD5 across diverse samples. Inhibiting the SMYD5-rpL40K22me3 axis in GAC cell lines reprogrammes protein synthesis to attenuate oncogenic gene expression signatures. SMYD5 and rpL40K22me3 are upregulated in samples from patients with GAC and negatively correlate with clinical outcomes. SMYD5 ablation in vivo in familial and sporadic mouse models of malignant GAC blocks metastatic disease, including peritoneal carcinomatosis. Suppressing SMYD5 methylation of rpL40 inhibits human cancer cell and patient-derived GAC xenograft growth and renders them hypersensitive to inhibitors of PI3K and mTOR. Finally, combining SMYD5 depletion with PI3K-mTOR inhibition and chimeric antigen receptor T cell administration cures an otherwise lethal in vivo mouse model of aggressive GAC-derived peritoneal carcinomatosis. Together, our work uncovers a ribosome-based epigenetic mechanism that facilitates the evolution of malignant GAC and proposes SMYD5 targeting as part of a potential combination therapy to treat this cancer.

A second-generation eIF4A RNA helicase inhibitor exploits translational reprogramming as a vulnerability in triple-negative breast cancer.

In this study, we aimed to address the current limitations of therapies for macro-metastatic triple-negative breast cancer (TNBC) and provide a therapeutic lead that overcomes the high degree of heterogeneity associated with this disease. Specifically, we focused on well-documented but clinically underexploited cancer-fueling perturbations in mRNA translation as a potential therapeutic vulnerability. We therefore developed an orally bioavailable rocaglate-based molecule, MG-002, which hinders ribosome recruitment and scanning via unscheduled and non-productive RNA clamping by the eukaryotic translation initiation factor (eIF) 4A RNA helicase. We demonstrate that MG-002 potently inhibits mRNA translation and primary TNBC tumor growth without causing overt toxicity in mice. Importantly, given that metastatic spread is a major cause of mortality in TNBC, we show that MG-002 attenuates metastasis in pre-clinical models. We report on MG-002, a rocaglate that shows superior properties relative to existing eIF4A inhibitors in pre-clinical models. Our study also paves the way for future clinical trials exploring the potential of MG-002 in TNBC and other oncological indications.

PABP prevents the untimely decay of select mRNA populations in human cells.

Gene expression is tightly regulated at the levels of both mRNA translation and stability. The poly(A)-binding protein (PABP) is thought to play a role in regulating these processes by binding the mRNA 3' poly(A) tail and interacting with both the translation and mRNA deadenylation machineries. In this study, we directly investigate the impact of PABP on translation and stability of endogenous mRNAs in human cells. Remarkably, our transcriptome-wide analysis only detects marginal mRNA translation changes in PABP-depleted cells. In contrast, rapidly depleting PABP alters mRNA abundance and stability, albeit non-uniformly. Otherwise stable transcripts, including those encoding proteins with constitutive functions, are destabilized in PABP-depleted cells. In contrast, many unstable mRNAs, including those encoding proteins with regulatory functions, decay at similar rates in presence or absence of PABP. Moreover, PABP depletion-induced cell death can partially be suppressed by disrupting the mRNA decapping and 5'-3' decay machinery. Finally, we provide evidence that the LSM1-7 complex promotes decay of "stable" mRNAs in PABP-depleted cells. Taken together, these findings suggest that PABP plays an important role in preventing the untimely decay of select mRNA populations.

Distinct, opposing functions for CFIm59 and CFIm68 in mRNA alternative polyadenylation of Pten and in the PI3K/Akt signalling cascade.

Precise maintenance of PTEN dosage is crucial for tumor suppression across a wide variety of cancers. Post-transcriptional regulation of Pten heavily relies on regulatory elements encoded by its 3'UTR. We previously reported the important diversity of 3'UTR isoforms of Pten mRNAs produced through alternative polyadenylation (APA). Here, we reveal the direct regulation of Pten APA by the mammalian cleavage factor I (CFIm) complex, which in turn contributes to PTEN protein dosage. CFIm consists of the UGUA-binding CFIm25 and APA regulatory subunits CFIm59 or CFIm68. Deep sequencing analyses of perturbed (KO and KD) cell lines uncovered the differential regulation of Pten APA by CFIm59 and CFIm68 and further revealed that their divergent functions have widespread impact for APA in transcriptomes. Differentially regulated genes include numerous factors within the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signalling pathway that PTEN counter-regulates. We further reveal a stratification of APA dysregulation among a subset of PTEN-driven cancers, with recurrent alterations among PI3K/Akt pathway genes regulated by CFIm. Our results refine the transcriptome selectivity of the CFIm complex in APA regulation, and the breadth of its impact in PTEN-driven cancers.

Fragment-type 4-azolylcoumarin derivatives with anticancer properties.

Several coumarin derivatives with a directly attached azole substituent at C-4 were synthesized and biologically studied for their anticancer properties. The cell lines used for this investigation (HeLa, K-562, MDA-MB-53, and MCF-7) demonstrated different sensitivities. The best response in the MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) assay was shown by K-562 cells, with compounds displaying activity (3c, IC50 3.06 µM; 4a, IC50 5.24 µM; 4c, IC50 4.7 µM) similar to that of cisplatin (IC50 ~6 µM), which was used as the standard. The studied azole-substituted coumarins demonstrated weaker activity toward other cell lines, except for compound 4c, which was equally potent in the case of MCF-7 cells. Additional biological evaluations supported interference with the cell cycle as a potential mechanism of action and confirmed the absence of toxicity in zebrafish embryos. On the basis of these initial results, 4-azole coumarins should be explored further. Although their activity would need additional optimization, the fact that these compounds are fragment-like structures with MW <300 and clog P <3 offers enough flexibility to fine-tune their drug-like properties.

Melatonin treatment affects changes in adrenal gene expression of catecholamine biosynthesizing enzymes and norepinephrine transporter in the rat model of chronic-stress-induced depression.

This study investigated the effects of melatonin treatment on adrenal catecholamine content, synthesis, uptake, and vesicular transport induced by the chronic unpredictable mild stress (CUMS) model of depression in rats. This entailed quantifying the norepinephrine, epinephrine, mRNA, and protein levels of tyrosine hydroxylase (TH), dopamine-ß-hydroxylase (DBH), phenylethanolamine N-methyltransferase (PNMT), norepinephrine transporter (NET), and vesicular monoamine transporter 2 (VMAT2) in the adrenal medulla. CUMS caused a significant depletion of norepinephrine stores and protein levels of TH, DBH, and NET, whereas the gene expression of PNMT was increased. It was observed that melatonin treatment in the CUMS rats prevented the stress-induced decrease in norepinephrine content and the protein expression of TH, DBH, and NET in the adrenal medulla of chronically stressed rats. The present study demonstrates the stimulatory effect of melatonin on adrenomedullary synthesis, the uptake and content of catecholamine in the rat model of chronic stress-induced depression.

Preparation of pyrrolizinone derivatives via sequential transformations of cyclic allyl imides: synthesis of quinolactacide and marinamide.

A facile synthetic route has been developed for the preparation of pyrrolizinone derivatives employing N-allyl imides as starting materials. The nucleophilic addition of a vinyl Grignard reagent/RCM/elimination sequence afforded pyrrolizinones in good yields and has been applied for the preparation of naturally occurring quinolactacide and marinamide.

Ganglion Cell-Inner Plexiform Layer Thickness in Different Glaucoma Stages Measured by Optical Coherence Tomography.

AIM: To compare ganglion cell (GCL) and inner plexiform layer (IPL) thickness in patients at different stages of primary open-angle glaucoma (POAG), determine their sensitivity and specificity values, and correlate thickness values with mean deviations (MD). METHODS: This prospective, cross- sectional study was conducted in a group of patients with confirmed POAG who were compared to an age- and gender-matched control group. Glaucomatous damage was classified according to the Hodapp-Parrish-Anderson scale: glaucoma stage 1 (early), glaucoma stage 2 (moderate), and glaucoma stage 3 (severe). The average, minimum, and all 6 sectoral (superotemporal, superior, superonasal, inferonasal, inferior, and inferotemporal) GCL + IPL thicknesses were measured and compared between groups. RESULTS: The average GCL + IPL thickness of 154 eyes of 93 patients in glaucoma stages 1, 2, 3, and 94 eyes of 47 persons in the control group were 76.79 ± 8.05, 65.90 ± 7.92, 57.38 ± 10.00, and 86.01 ± 3.68 µm, respectively. There were statistically significant differences in the average, minimum, and all 6 sectoral GCL + IPL values among the groups. The areas under the receiver operating characteristic curve for average and minimum GCL + IPL thickness values were 0.93 and 0.94, respectively, sensitivity 91.5 and 88.3%, and specificity 98.9 and 100%, respectively. Both thickness values showed significant correlations with MD. Each micrometer decrease in the average GCL + IPL thickness was associated with a 0.54-dB loss in MD. CONCLUSION: GCL + IPL layer thickness is a highly specific and sensitive parameter in differentiating glaucomatous from healthy eyes showing progressive damage as glaucoma worsens. Loss of this layer is highly correlated with overall loss of visual field sensitivity.

A novel thiourea type organocatalyst possessing a single NH functionality.

A novel thiourea organocatalyst was rationally designed by altering a typical H-bonding pattern of thiourea derivatives and utilising the potential of the 3,5-bis(trifluoromethyl)phenyl motif to participate in the H-bond formation. This unique catalyst afforded the products of the α-amination and Michael reaction in excellent yields and with a high level of stereoselectivity. Although additional studies are necessary to establish the full potential of the catalyst and to broaden its application further, the presented results may indicate alternative routes for further exploration of the thiourea class of organocatalysts.

Effects of oxytocin on adreno-medullary catecholamine synthesis, uptake and storage in rats exposed to chronic isolation stress.

PURPOSE/AIM: The adreno-medullar system represents one of the main systems involved in the response to stressful events. The neuropeptide oxytocin, is highly sensitive to the social environment, and regulates autonomic function. Adreno-medullary activity is dependent on the synthesis of catecholamine, its reuptake, release, degradation and vesicular transport. A direct influence of oxytocin on catecholamine synthesizing enzyme and transports in animals exposed to chronic social isolation stress has not been studied yet. MATERIALS AND METHODS: In the present study, we examined the effect of chronic oxytocin treatment on the level of plasma catecholamine and its content, mRNA and protein levels of tyrosine hydroxylase (TH), noradrenaline transporter (NET) as well as vesicular monoamine transporter 2 (VMAT2) in the adrenal medulla of socially isolated rats. RESULTS: Our results show that, by the end of 12 weeks, social isolation did not produce any significant changes in catecholamine content but increased plasma catecholamine level and synthesis in the adrenal medulla. Oxytocin treatment had no further effect either on catecholamine synthesis or content in socially stressed animals whereas a significant elevation of plasma norepinephrine and epinephrine were reduced. On the other hand, chronic isolation caused a significant increase in VMAT2 and decrease in NET protein levels. Oxytocin treatment brought about an increase in protein levels of NET and its return to the levels of control group. Besides, it further increases VMAT2 protein levels in the adrenal medulla of individually housed rats. CONCLUSION: The present results show that peripheral oxytocin treatment enhances catecholamine uptake and storage in the adrenal medulla of chronically isolated animals.

Differential regulation of catecholamine synthesis and transport in rat adrenal medulla by fluoxetine treatment.

We have recently shown that chronic fluoxetine treatment acted significantly increasing plasma norepinephrine and epinephrine concentrations both in control and chronically stressed adult male rats. However, possible effects of fluoxetine on catecholamine synthesis and re-uptake in adrenal medulla have been largely unknown. In the present study the effects of chronic fluoxetine treatment on tyrosine hydroxylase, a rate-limiting enzyme in catecholamine synthesis, as well as a norepinephrine transporter and vesicular monoamine transporter 2 gene expressions in adrenal medulla of animals exposed to chronic unpredictable mild stress (CUMS) for 4 weeks, were investigated. Gene expression analyses were performed using a real-time quantitative reverse transcription-PCR. Chronically stressed animals had increased tyrosine hydroxylase mRNA levels and decreased expression of both transporters. Fluoxetine increased tyrosine hydroxylase and decreased norepinephrine transporter gene expression in both unstressed and CUMS rats. These findings suggest that chronic fluoxetine treatment increased plasma catecholamine levels by affecting opposing changes in catecholamine synthesis and uptake.

FibroTest/Fibrosure for significant liver fibrosis and cirrhosis in chronic hepatitis B: a meta-analysis.

OBJECTIVES: Extent of liver fibrosis is one of the most important factors in determining prognosis and the need for active treatment in chronic hepatitis B (CHB). Noninvasive alternatives such as FibroTest/Fibrosure (FT) have been developed in order to overcome the shortcomings of liver biopsy (LB). We aimed to systematically review studies describing the diagnostic accuracy of FT for predicting CHB-related fibrosis. METHODS: MEDLINE and EMBASE searches and hand searching methods were performed to identify studies that assessed the diagnostic accuracy of FibroTest in HB patients using LB as a reference standard. We used a hierarchical summary receiver operating curves model and the bivariate model to produce summary receiver operating characteristic curves and pooled estimates of sensitivity and specificity. RESULTS: We included 16 studies (N=2494) and 13 studies (N=1754) in the heterogenous meta-analysis for liver fibrosis and cirrhosis, respectively. The area under the hierarchical summary receiver operating curve for significant liver fibrosis and for all included studies was 0.84 (95% confidence interval (CI): 0.78-0.88). At the FT threshold of 0.48, the sensitivity, specificity, and diagnostic odds ratio (DOR) of FT for significant fibrosis were 61 (48-72%), 80 (72-86%), and 6.2% (3.3-11.9), respectively. The area under the hierarchical summary receiver operating curve for liver cirrhosis and for all included studies was 0.87 (95% CI: 0.85-0.90). At the FT threshold of 0.74, the sensitivity, specificity, and DOR of FT for cirrhosis were 62 (47-75%), 91 (88-93%), and 15.7% (8.6-28.8), respectively. CONCLUSIONS: FibroTest is of value in exclusion of patients with CHB-related cirrhosis, but has suboptimal accuracy in the detection of significant fibrosis and cirrhosis. It is necessary to further improve the test or combine it with other noninvasive modalities in order to improve accuracy.

Artificial neural network predicts the need for therapeutic ERCP in patients with suspected choledocholithiasis.

BACKGROUND: Selection of patients with the highest probability for therapeutic ERCP remains an important task in a clinical workup of patients with suspected choledocholithiasis (CDL). OBJECTIVE: To determine whether an artificial neural network (ANN) model can improve the accuracy of selecting patients with a high probability of undergoing therapeutic ERCP among those with strong clinical suspicion of CDL and to compare it with our previously reported prediction model. DESIGN: Prospective, observational study. SETTING: Single, tertiary-care endoscopy center. PATIENTS: Between January 2010 and September 2012, we prospectively recruited 291 consecutive patients who underwent ERCP after being referred to our center with firm suspicion for CDL. INTERVENTIONS: Predictive scores for CDL based on a multivariate logistic regression model and ANN model. MAIN OUTCOME MEASUREMENTS: The presence of common bile duct stones confirmed by ERCP. RESULTS: There were 80.4% of patients with positive findings on ERCP. The area under the receiver-operating characteristic curve for our previously established multivariate logistic regression model was 0.787 (95% CI, 0.720-0.854; P < .001), whereas area under the curve for the ANN model was 0.884 (95% CI, 0.831-0.938; P < .001). The ANN model correctly classified 92.3% of patients with positive findings on ERCP and 69.6% patients with negative findings on ERCP. LIMITATIONS: Only those variables believed to be related to the outcome of interest were included. The majority of patients in our sample had positive findings on ERCP. CONCLUSIONS: An ANN model has better discriminant ability and accuracy than a multivariate logistic regression model in selecting patients for therapeutic ERCP.

Machine Learning for Short-Term Mortality in Acute Decompensation of Liver Cirrhosis: Better than MELD Score.

Prediction of short-term mortality in patients with acute decompensation of liver cirrhosis could be improved. We aimed to develop and validate two machine learning (ML) models for predicting 28-day and 90-day mortality in patients hospitalized with acute decompensated liver cirrhosis. We trained two artificial neural network (ANN)-based ML models using a training sample of 165 out of 290 (56.9%) patients, and then tested their predictive performance against Model of End-stage Liver Disease-Sodium (MELD-Na) and MELD 3.0 scores using a different validation sample of 125 out of 290 (43.1%) patients. The area under the ROC curve (AUC) for predicting 28-day mortality for the ML model was 0.811 (95%CI: 0.714- 0.907; p < 0.001), while the AUC for the MELD-Na score was 0.577 (95%CI: 0.435-0.720; p = 0.226) and for MELD 3.0 was 0.600 (95%CI: 0.462-0.739; p = 0.117). The area under the ROC curve (AUC) for predicting 90-day mortality for the ML model was 0.839 (95%CI: 0.776- 0.884; p < 0.001), while the AUC for the MELD-Na score was 0.682 (95%CI: 0.575-0.790; p = 0.002) and for MELD 3.0 was 0.703 (95%CI: 0.590-0.816; p < 0.001). Our study demonstrates that ML-based models for predicting short-term mortality in patients with acute decompensation of liver cirrhosis perform significantly better than MELD-Na and MELD 3.0 scores in a validation cohort.

Short-term efficacy of vedolizumab in patients with inflammatory bowel disease in real-life settings in Bosnia and Herzegovina.

Inflammatory bowel disease (IBD), encompassing Crohn's disease (CD) and ulcerative colitis (UC), necessitates effective management strategies. This study aims to evaluate the real-world efficacy of vedolizumab, a newer biological therapy, in treating IBD in Bosnia and Herzegovina. A retrospective observational study was conducted across 6 medical centers, involving 139 IBD patients, 76 with UC and 63 with CD. Patients were assessed for clinical remission and other outcomes at the 26-week mark post vedolizumab treatment initiation. At 26 weeks, clinical remission was achieved in 82.9% of UC patients and 85.7% of CD patients. Mucosal healing was observed in 38.1% of CD patients. The efficacy of vedolizumab did not significantly differ based on prior anti-tumor necrosis factor (TNF) exposure. Notably, the clinical scoring tools for predicting vedolizumab response showed limited applicability in this cohort. Vedolizumab demonstrated high efficacy in treating both UC and CD in a real-world settings in Bosnia and Herzegovina, underscoring its potential as a significant therapeutic option in IBD management.

MTOR modulation induces selective perturbations in histone methylation which influence the anti-proliferative effects of mTOR inhibitors.

Emerging data suggest a significant cross-talk between metabolic and epigenetic programs. However, the relationship between the mechanistic target of rapamycin (mTOR), which is a pivotal metabolic regulator, and epigenetic modifications remains poorly understood. Our results show that mTORC1 activation caused by the abrogation of its negative regulator tuberous sclerosis complex 2 (TSC2) coincides with increased levels of the histone modification H3K27me3 but not H3K4me3 or H3K9me3. This selective H3K27me3 induction was mediated via 4E-BP-dependent increase in EZH2 protein levels. Surprisingly, mTOR inhibition also selectively induced H3K27me3. This was independent of TSC2, and was paralleled by reduced EZH2 and increased EZH1 protein levels. Notably, the ability of mTOR inhibitors to induce H3K27me3 levels was positively correlated with their anti-proliferative effects. Collectively, our findings demonstrate that both activation and inhibition of mTOR selectively increase H3K27me3 by distinct mechanisms, whereby the induction of H3K27me3 may potentiate the anti-proliferative effects of mTOR inhibitors.

A sex-specific thermogenic neurocircuit induced by predator smell recruiting cholecystokinin neurons in the dorsomedial hypothalamus.

Olfactory cues are vital for prey animals like rodents to perceive and evade predators. Stress-induced hyperthermia, via brown adipose tissue (BAT) thermogenesis, boosts physical performance and facilitates escape. However, many aspects of this response, including thermogenic control and sex-specific effects, remain enigmatic. Our study unveils that the predator odor trimethylthiazoline (TMT) elicits BAT thermogenesis, suppresses feeding, and drives glucocorticoid release in female mice. Chemogenetic stimulation of olfactory bulb (OB) mitral cells recapitulates the thermogenic output of this response and associated stress hormone corticosterone release in female mice. Neuronal projections from OB to medial amygdala (MeA) and dorsomedial hypothalamus (DMH) exhibit female-specific cFos activity toward odors. Cell sorting and single-cell RNA-sequencing of DMH identify cholecystokinin (CCK)-expressing neurons as recipients of predator odor cues. Chemogenetic manipulation and neuronal silencing of DMHCCK neurons further implicate these neurons in the propagation of predator odor-associated thermogenesis and food intake suppression, highlighting their role in female stress-induced hyperthermia.

Myeloid cell ACE shapes cellular metabolism and function in PCSK-9 induced atherosclerosis.

The pathogenesis of atherosclerosis is defined by impaired lipid handling by macrophages which increases intracellular lipid accumulation. This dysregulation of macrophages triggers the accumulation of apoptotic cells and chronic inflammation which contributes to disease progression. We previously reported that mice with increased macrophage-specific angiotensin-converting enzyme, termed ACE10/10 mice, resist atherosclerosis in an adeno-associated virus-proprotein convertase subtilisin/kexin type 9 (AAV-PCSK9)-induced model. This is due to increased lipid metabolism by macrophages which contributes to plaque resolution. However, the importance of ACE in peripheral blood monocytes, which are the primary precursors of lesional-infiltrating macrophages, is still unknown in atherosclerosis. Here, we show that the ACE-mediated metabolic phenotype is already triggered in peripheral blood circulating monocytes and that this functional modification is directly transferred to differentiated macrophages in ACE10/10 mice. We found that Ly-6Clo monocytes were increased in atherosclerotic ACE10/10 mice. The monocytes isolated from atherosclerotic ACE10/10 mice showed enhanced lipid metabolism, elevated mitochondrial activity, and increased adenosine triphosphate (ATP) levels which implies that ACE overexpression is already altered in atherosclerosis. Furthermore, we observed increased oxygen consumption (VO2), respiratory exchange ratio (RER), and spontaneous physical activity in ACE10/10 mice compared to WT mice in atherosclerotic conditions, indicating enhanced systemic energy consumption. Thus, ACE overexpression in myeloid lineage cells modifies the metabolic function of peripheral blood circulating monocytes which differentiate to macrophages and protect against atherosclerotic lesion progression due to better lipid metabolism.

Trigeminal artery: a review of normal and pathological features.

OBJECTS: Carotid-vertebrobasilar anastomoses-the trigeminal, otic, hypoglossal, and proatlantal intersegmental arteries-serve as transitory channels between primitive internal carotid arteries and bilateral longitudinal neural arterial plexus, which is the precursor of future basilar artery, when the human embryo reaches about 4-mm length. MATERIAL AND METHODS: Normal and/or abnormal morphofunctional aspects of the prenatal and postnatal forms of the trigeminal artery are described according to personal and literature data. Many arteries of similar origin and course are also noted in the differential diagnosis of the trigeminal artery. CONCLUSIONS: The persistent primitive trigeminal artery, as the most commonly carotid-vertebrobasilar anastomosis, has a reported incidence of 0.03-2.2% in the literature. There is female sex predilection, and it may be discovered in patients of any age, on either side, and in association with many vascular variants. Although the significance of persistent primitive trigeminal artery regarding the development of an aneurysm or association with another pathological condition may not be clear, its (ab)normal morphology is the inspiration for anatomists, especially for neurosurgeons, before planning diagnostic and therapeutic procedures.