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Shortening of the 3′ untranslated regions (3′UTR) of mRNA is an important mechanism for oncogene activation. However, 3′UTR alteration events, their pathologic functions, and underlying mechanisms in human urothelial carcinoma of the bladder (UCB) are not clear. Here, we combine RNA sequencing, bioinformatics, and clinical studies in two independent cohorts of patients with UCB to identify a novel RAC1 shorter 3′UTR isoform that is frequently expressed in UCB and is critical in the tumorigenesis and acquisition of a poor prognostic phenotype in patients. Short 3′UTR isoform of RAC1 substantially upregulated RAC1 expression by escaping from miRNA-targeted repression and played an essential oncogenic role in UCB pathogenesis. An important cleavage/polyadenylation factor, cleavage stimulation factor 2 (CSTF2), induced 3′UTR shortening of RAC1 in UCB by mediating slow transcriptional elongation at RAC1. Cotranscriptional recruitment of CSTF2 on the GUAAU motif at proximal polyadenylation site of RAC1 attenuated the recruitment of two transcription factors AFF1 and AFF4, causing the defects in elongation. CSTF2 regulated the tumorigenic functions of the shorter RAC1 isoform in UCB cells, enhancing cell proliferation, migration, and invasion. The combination of high expression of CSTF2 and high usage of RAC1 short-3′UTR isoform may be used as a powerful biomarker to predict poor prognosis in UCB. Our findings also suggest a CSTF2-regulated RAC1-3′UTR shortening program as an exploitable therapeutic strategy for patients with UCB.Significance: These findings demonstrate that the short isoform of RAC1 is critical in UCB tumorigenesis and may have implications for developing new therapeutic strategies to treat this disease. Cancer Res; 78(20); 5848–62. ©2018 AACR.

Supplementary Figure S1. Identification of 3'UTR shortening of RALA and RAC1 from UCB, colorectal carcinoma, hepatocellular carcinoma and non-small cell lung cancer samples. Supplementary Figure S2. RAC1 with short 3'UTR promotes tumorigenesis and produces more protein due to evasion of miRNA-mediated repression. Supplementary Figure S3. Screening of APA factors that contribute to the PAS selection of RAC1 in UCB. Supplementary Figure S4. CSTF2 regulates colony formation and invasiveness abilities of shorter RAC1 isoform in UCB. Supplementary Table S1. PDUI value calculated from DaPars analysis of 15 pairs of UCB and adjacent normal bladder tissues. Supplementary Table S2. KEGG pathway analyses of APA genes identified from RNA-Seq. Supplementary Table S3. RAC1 genomic mutations identified from TCGA UCB database. Supplementary Table S4. Correlation between the expression of CSTF2 and clinicopathologic characteristics of UCB patients

Background Macrophages, including lung alveolar macrophages (AM), and monocytes are the first lines of defense against SARS-CoV-2. Several reports have suggested that SARS-CoV-2 can hijack AM and monocytes for replication and viral spread, which may, in turn, drive the cytokine storm associated with severe COVID-19. Herein, we describe one of many advantageous features that EDP-235, a novel and potent SARS-CoV-2 3C-like protease (3CLpro) inhibitor under development as a once-daily oral antiviral therapy for COVID-19, displays - excellent penetration into macrophages and monocytes. Methods Intracellular uptake of EDP-235 was tested side-by-side with nirmatrelvir in rat lung AM, human monocytes and human macrophages. To determine the in vivo drug distribution into lung AM, rats were dosed orally with 25 mg/kg of EDP-235 or nirmatrelvir and plasma and AM drug levels were analyzed by LC/MS/MS. Results The ratios of intracellular to extracellular concentrations of EDP-235 in rat lung AM, human monocytes and human macrophages were 22.8, 22.7 and 30.5, respectively. In contrast, nirmatrelvir had ratios of 1.2 to 1.5 in these cells. Consistent with the in vitro observations, EDP-235 showed favorable rat AM penetration with an AUC0-24 ratio of 28.4 (AM over plasma), and nirmatrelvir had much less rat AM penetration with an AUC0-24 ratio of 0.5 (AM over plasma). EDP-235 had respective AUC0-24 values of 9.6 and 271.9 h·μg/mL in rat plasma and AM, while the AUC0-24 values of nirmatrelvir in rat plasma and AM were 2.7 and 1.2 h·μg/mL, respectively. Conclusion EDP-235, a novel and potent SARS-CoV-2 3CL protease inhibitor, demonstrated excellent penetration into monocytes and macrophages, including lung AM. EDP-235 has the potential to eliminate the viral replication of SARS-CoV-2 in these critical immune cells, thus mitigating macrophage-mediated cytokine storm in high-risk COVID-19 patients. Clinical trials with EDP-235 for COVID-19 treatment and prevention are ongoing. Disclosures All Authors: No reported disclosures.

Background EDP‐305 is a novel and potent farnesoid X receptor (FXR) agonist, with no/minimal cross‐reactivity to TGR5 or other nuclear receptors. Herein we report therapeutic efficacy of EDP‐305, in direct comparison with the first‐in‐class FXR agonist obeticholic acid (OCA), in mouse models of liver disease. Methods EDP‐305 (10 and 30 mg/kg/day) or OCA (30mg/kg/day) was tested in mouse models of pre‐established biliary fibrosis (BALBc.Mdr2‐/‐, n = 9‐12/group) and steatohepatitis induced by methionine/choline‐deficient diet (MCD, n = 7‐12/group). Effects on biliary epithelium were evaluated in vivo and in primary EpCAM + hepatic progenitor cell (HPC) cultures. Results In a BALBc.Mdr2‐/‐ model, EDP‐305 reduced serum transaminases by up to 53% and decreased portal pressure, compared to untreated controls. Periportal bridging fibrosis was suppressed by EDP‐305 at both doses, with up to a 39% decrease in collagen deposition in high‐dose EDP‐305. In MCD‐fed mice, EDP‐305 treatment reduced serum ALT by 62% compared to controls, and profoundly inhibited perisinusoidal ‘chicken wire’ fibrosis, with over 80% reduction in collagen deposition. In both models, treatment with 30mg/kg OCA reduced serum transaminases up to 30%, but did not improve fibrosis. The limited impact on fibrosis was mediated by cholestasis‐independent worsening of ductular reaction by OCA in both disease models; OCA but not EDP‐305 at therapeutic doses promoted ductular proliferation in healthy mice and favoured differentiation of primary HPC towards cholangiocyte lineage in vitro. Conclusions EDP‐305 potently improved pre‐established liver injury and hepatic fibrosis in murine biliary and metabolic models of liver disease, supporting the clinical evaluation of EDP‐305 in fibrotic liver diseases including cholangiopathies and non‐alcoholic steatohepatitis.

An approach for the construction of furo[3,2-b]quinolines and furo[2,3-b:4,5-b′]diquinolines is developed through a metal-free [4 + 2] cycloaddition of easily available in situ generated aza-o-quinone methides and furans. The reaction tolerates a wide range of aza-o-quinone methides and substituted furans to afford the corresponding dihydro- or tetrahydrofuroquinolines in good to excellent yields. Mechanistic studies reveal that the reaction involves a concerted [4 + 2] cycloaddition pathway and shows a high regioselectivity of cycloaddition for a furan ring. The present method features mild reaction conditions, dearomatization of furans, high regio- and diastereoselectivity, gram-scalable preparations, and diversity of furoquinolines.

Farnesoid X receptor (FXR) is a nuclear receptor that has emerged as a key regulator in the maintenance of hepatic steatosis, inflammation, and fibrosis. However, the role of FXR in renal fibrosis remains to be established. Here, we investigate the effects of the FXR agonist EDP‐305 in a mouse model of tubulointerstitial fibrosis via unilateral ureteral obstruction (UUO). Male C57Bl/6 mice received a UUO on their left kidney. On postoperative d 4, mice received daily treatment by oral gavage with either vehicle control (0.5% methylcellulose) or 10 or 30 mg/kg EDP‐305. All animals were euthanized on postoperative d 12. EDP‐305 dose‐dependently decreased macrophage infiltration as measured by the F4/80 staining area and proinflammatory cytokine gene expression. EDP‐305 also dose‐dependently reduced interstitial fibrosis as assessed by morphometric quantification of the collagen proportional area and kidney hydroxyproline levels. Finally, yes‐associated protein (YAP) activation, a major driver of fibrosis, increased after UUO injury and was diminished by EDP‐305 treatment. Consistently, EDP‐305 decreased TGF‐β1—induced YAP nuclear localization in human kidney 2 cells by increasing inhibitory YAP phosphorylation. YAP inhibition may be a novel antifibrotic mechanism of FXR agonism, and EDP‐305 could be used to treat renal fibrosis.—Li, S., Ghoshal, S., Sojoodi, M., Arora, G., Masia, R., Erstad, D. J., Ferriera, D. S., Li, Y., Wang, G., Lanuti, M., Caravan, P., Or, Y. S., Jiang, L.‐J., Tanabe, K. K., Fuchs, B. C. The farnesoid X receptor agonist EDP‐305 reduces interstitial renal fibrosis in a mouse model of unilateral ureteral obstruction. FASEB J. 33, 7103–7112 (2019). www.fasebj.org

Background Pleural fibrosis is defined as excessive depositions of matrix components that result in pleural tissue architecture destruction and dysfunction. In severe cases, the progression of pleural fibrosis leads to lung entrapment, resulting in dyspnea and respiratory failure. However, the mechanism of pleural fibrosis is poorly understood. Methods miR-4739 levels were detected by miRNA array and real-time PCR. Real-time PCR, western blotting and immunofluorescence were used to identify the expression profile of indicators related to fibrosis. Target gene of miR-4739 and promoter activity assay was measured by using dual-luciferase reporter assay system. In vivo, pleural fibrosis was evaluated by Masson staining and miR-4739 level was detected by In situ hybridization histochemistry. Findings We found that bleomycin induced up-regulation of miR-4739 in pleural mesothelial cells (PMCs). Over-regulated miR-4739 mediated mesothelial-mesenchymal transition and increased collagen-I synthesis in PMCs. Investigation on the clinical specimens revealed that high levels of miR-4739 and low levels of bone morphogenetic protein 7 (BMP-7) associated with pleural fibrosis in patients. Then we next identified that miR-4739 targeted and down-regulated BMP-7 which further resulted in unbalance between Smad1/5/9 and Smad2/3 signaling. Lastly, in vivo studies revealed that miR-4739 over-expression induced pleural fibrosis, and exogenous BMP-7 prevented pleural fibrosis in mice. Interpretation Our data indicated that miR-4739 targets BMP-7 which mediates pleural fibrosis. The miR-4739/BMP-7 axis is a promising therapeutic target for the disease. Fund The National Natural Science Foundation of China.

Purpose There are limited therapeutic options for patients with advanced penile squamous cell carcinoma (PSCC) after chemotherapy failure. Thus, we evaluated the feasibility of salvage treatment using the epidermal growth factor receptor (EGFR) mono-antibody nimotuzumab in chemotherapy-failed PSCC patients and explored potential response or resistance biomarkers. Materials and methods Six chemotherapy-failed PSCC patients with locally advanced disease or distant metastasis were enrolled consecutively to nimotuzumab treatment. Clinical responses and side effects were evaluated, and genetic characteristics of cancer specimens were analyzed through the next-generation sequencing of hotspot regions in cancer-related genes. Results Two of 6 patients showed partial responses, one was identified as having stable disease, while the other 3 had disease progression after nimotuzumab therapy. Side effects were all welltolerated. Genetic analysis revealed that TP53, CDKN2A, RB1, SMAD4, FLT3, and PIK3CA were the most frequently mutated genes in PSCC specimens, while altered KRAS, HRAS, EGFR, ERBB2, and FLT3 may be correlated with nimotuzumab resistance. Furthermore, 3 patients that were human papillomavirus-positive each showed clinical response or stable disease. Conclusions EGFR mono-antibody may be a potential modality for locally advanced PSCC patients after chemotherapy failure. Further large-scale clinical studies are needed to elucidate the role of human papillomavirus status and critical gene mutations in the clinical response to EGFR-targeted therapy.

The weak decays of a spin-1 / 2 doubly charm baryon ( $$\mathcal{B}_{cc}$$ ) to a spin-1 / 2 singly charm baryon ( $$\mathcal{B}_c$$ ) and a light vector meson (V) are studied under a phenomenological scheme. The contributions are classified into different topological diagrams, among which the short distance ones are calculated under the factorization hypothesis, and the long distance contributions are modelled as final-state interactions (FSIs) which are estimated with the one-particle-exchange model. In calculation the topological contributions tend to fall in a hierarchy. The branching fractions or decay widths are estimated, and it indicates that $$\Xi _{cc}^+\rightarrow \Xi _c^+\pi ^+\pi ^-$$ and $$\Omega _{cc}^{+}\rightarrow \Xi _{c}^{+}K^-\pi ^+$$ can be used as candidate decays for searching $$\Xi _{cc}^+$$ and $$\Omega _{cc}^+$$ . Some decays that are mainly activated by the long distance effects are found, observation on which in future experiments can help to understand the role of FSIs in charm baryon decays.

Shortening of the 3′ untranslated regions (3′UTR) of mRNA is an important mechanism for oncogene activation. However, 3′UTR alteration events, their pathologic functions, and underlying mechanisms in human urothelial carcinoma of the bladder (UCB) are not clear. Here, we combine RNA sequencing, bioinformatics, and clinical studies in two independent cohorts of patients with UCB to identify a novel RAC1 shorter 3′UTR isoform that is frequently expressed in UCB and is critical in the tumorigenesis and acquisition of a poor prognostic phenotype in patients. Short 3′UTR isoform of RAC1 substantially upregulated RAC1 expression by escaping from miRNA-targeted repression and played an essential oncogenic role in UCB pathogenesis. An important cleavage/polyadenylation factor, cleavage stimulation factor 2 (CSTF2), induced 3′UTR shortening of RAC1 in UCB by mediating slow transcriptional elongation at RAC1. Cotranscriptional recruitment of CSTF2 on the GUAAU motif at proximal polyadenylation site of RAC1 attenuated the recruitment of two transcription factors AFF1 and AFF4, causing the defects in elongation. CSTF2 regulated the tumorigenic functions of the shorter RAC1 isoform in UCB cells, enhancing cell proliferation, migration, and invasion. The combination of high expression of CSTF2 and high usage of RAC1 short-3′UTR isoform may be used as a powerful biomarker to predict poor prognosis in UCB. Our findings also suggest a CSTF2-regulated RAC1-3′UTR shortening program as an exploitable therapeutic strategy for patients with UCB. Significance: These findings demonstrate that the short isoform of RAC1 is critical in UCB tumorigenesis and may have implications for developing new therapeutic strategies to treat this disease. Cancer Res; 78(20); 5848–62. ©2018 AACR.

An approach for the construction of furo[3,2

EDP-239, a novel hepatitis C virus (HCV) inhibitor targeting nonstructural protein 5A (NS5A), has been investigated in vitro and in vivo . EDP-239 is a potent, selective inhibitor with potency at picomolar to nanomolar concentrations against HCV genotypes 1 through 6. In the presence of human serum, the potency of EDP-239 was reduced by less than 4-fold. EDP-239 is additive to synergistic with other direct-acting antivirals (DAAs) or host-targeted antivirals (HTAs) in blocking HCV replication and suppresses the selection of resistance in vitro . Furthermore, EDP-239 retains potency against known DAA- or HTA-resistant variants, with half-maximal effective concentrations (EC 50 s) equivalent to those for the wild type. In a phase I, single-ascending-dose, placebo-controlled clinical trial, EDP-239 demonstrated excellent pharmacokinetic properties that supported once daily dosing. A single 100-mg dose of EDP-239 resulted in reductions in HCV genotype 1a viral RNA of >3 log 10 IU/ml within the first 48 h after dosing and reductions in genotype 1b viral RNA of >4-log 10 IU/ml within 96 h. (This study has been registered at ClinicalTrials.gov under identifier NCT01856426.)

Background Respiratory syncytial virus (RSV) infection presents a significant health challenge in young children, elderly and immunocompromised patients. To date, there are no effective treatments available. EDP-938 was designed to meet this unmet medical need and is currently in Phase 2 clinical trials. Herein we report its preclinical pharmacokinetic (PK) and pharmacodynamic (PD) properties. Methods The pharmacokinetics of EDP-938 following single intravenous and oral doses were determined in mice, rats, dogs, and monkeys. In vitro cellular permeability and metabolic stability were assayed using Caco-2 cells and human liver microsomes, respectively. In vivo pharmacodynamic efficacy of EDP-938 was conducted in the African green monkey model, in which animals experimentally challenged with RSV were orally dosed twice daily with 100 mg/kg EDP-938 for 6 days starting 24 hours prior to infection. Results EDP-938 was well absorbed in the preclinical species with oral bioavailability values ranging from 27.1% in dogs, 35.4% in mice, 35.7% in rats, and 39.5% in monkeys, after a single oral dose when formulated in 0.5% methylcellulose. EDP-938 showed a moderate in vitro permeability of 3.6 x 10–6 cm/sec in Caco-2 cells. Based on the outcome of these absorption studies, EDP-938 was projected to have good oral absorption in humans. EDP-938 had low intrinsic clearance of 5 mL/minute/mg in human liver microsomes. Moreover, EDP-938 demonstrated potent antiviral efficacy in an African green monkey model of RSV infection. In untreated monkeys the RSV RNA viral load in the bronchoalveolar lavage fluid peaked at 106 copies/mL on day 5 post-infection, by comparison in animals treated with EDP-938 the viral load was below the limit of detection by day 3 post-infection. The PK/PD modeling suggested that plasma trough concentrations ≥10 × EC90 led to >4-log viral load reduction in EDP-938 treated monkeys. Conclusion The favorable preclinical PK and PD properties of EDP-938 support its further clinical development as a novel treatment for RSV infection. Disclosures All authors: No reported disclosures.

Cytokine-induced killer (CIK) cells that are stimulated using mature dendritic cells (DCs), referred to as (DC-CIK cells) exhibit superior anti-tumor potency. Anti-programmed death-1 (PD-1) antibodies reinvigorate T cell-mediated antitumor immunity. This phase I study aimed to assess the safety and clinical activity of immunotherapy with PD-1 blockade (pembrolizumab)-activated autologous DC-CIK cells in patients with advanced solid tumors. Patients with selected types of advanced solid tumors received a single intravenous infusion of activated autologous DC-CIK cells weekly for the first month and every 2 weeks thereafter. The primary end points were safety and adverse event (AE) profiles. Antitumor responses, overall survival (OS), progression-free survival (PFS) and cytolytic activity were secondary end points. Treatment-related AEs occurred in 20/31 patients. Grade 3 or 4 toxicities, including fever and chills, were observed in two patients. All treatment-related AEs were reversible or controllable. The cytotoxicity of DC-CIK cells induced up-regulation of PD-L1 expression on autologous tumor cells. When activated using pembrolizumab ex vivo, DC-CIK cells exerted superior antitumor properties and elevated IFN-γ secretion. Objective responses (complete or partial responses) were observed in 7 of the 31patients.These responses were durable, with 6 of 7 responses lasting more than 5 months. The overall disease control rate in the patients was 64.5%. At the time of this report, the median OS and PFS were 270 and 162 days, respectively. In conclusions, treatment with pembrolizumab-activated autologous DC-CIK cells was safe and exerted encouraging antitumor activity in advanced solid tumors. A larger phase II trial is warranted.

Shortening of the 3' untranslated regions (3'UTR) of mRNA is an important mechanism for oncogene activation. However, 3'UTR alteration events, their pathologic functions, and underlying mechanisms in human urothelial carcinoma of the bladder (UCB) are not clear. Here, we combine RNA sequencing, bioinformatics, and clinical studies in two independent cohorts of patients with UCB to identify a novel

Glecaprevir (formerly ABT-493) is a novel hepatitis C virus (HCV) NS3/4A protease inhibitor (PI) with pangenotypic activity. It inhibited the enzymatic activity of purified NS3/4A proteases from HCV genotypes 1 to 6 in vitro (half-maximal [50%] inhibitory concentration = 3.5 to 11.3 nM) and the replication of stable HCV subgenomic replicons containing proteases from genotypes 1 to 6 (50% effective concentration [EC 50 ] = 0.21 to 4.6 nM). Glecaprevir had a median EC 50 of 0.30 nM (range, 0.05 to 3.8 nM) for HCV replicons containing proteases from 40 samples from patients infected with HCV genotypes 1 to 5. Importantly, glecaprevir was active against the protease from genotype 3, the most-difficult-to-treat HCV genotype, in both enzymatic and replicon assays demonstrating comparable activity against the other HCV genotypes. In drug-resistant colony selection studies, glecaprevir generally selected substitutions at NS3 amino acid position A156 in replicons containing proteases from genotypes 1a, 1b, 2a, 2b, 3a, and 4a and substitutions at position D/Q168 in replicons containing proteases from genotypes 3a, 5a, and 6a. Although the substitutions A156T and A156V in NS3 of genotype 1 reduced susceptibility to glecaprevir, replicons with these substitutions demonstrated a low replication efficiency in vitro . Glecaprevir is active against HCV with most of the common NS3 amino acid substitutions that are associated with reduced susceptibility to other currently approved HCV PIs, including those at positions 155 and 168. Combination of glecaprevir with HCV inhibitors with other mechanisms of action resulted in additive or synergistic antiviral activity. In summary, glecaprevir is a next-generation HCV PI with potent pangenotypic activity and a high barrier to the development of resistance.

We examined a novel farnesoid X receptor agonist, EDP-305, for its antifibrotic effect in bile duct ligation (BDL) and choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) models of hepatic injury. We used molecular magnetic resonance imaging with the type 1 collagen-binding probe EP-3533 and the oxidized collagen-specific probe gadolinium hydrazide to noninvasively measure treatment response. BDL rats (n = 8 for each group) were treated with either low or high doses of EDP-305 starting on day 4 after BDL and were imaged on day 18. CDAHFD mice (n = 8 for each group) were treated starting at 6 weeks after the diet and were imaged at 12 weeks. Liver tissue was subjected to pathologic and morphometric scoring of fibrosis, hydroxyproline quantitation, and determination of fibrogenic messenger RNA expression. High-dose EDP-305 (30 mg/kg) reduced liver fibrosis in both the BDL and CDAHFD models as measured by collagen proportional area, hydroxyproline analysis, and fibrogenic gene expression (all P < 0.05). Magnetic resonance signal intensity with both EP-3533 in the BDL model and gadolinium hydrazide in the CDAHFD model was reduced with EDP-305 30 mg/kg treatment (P < 0.01). Histologically, EDP-305 30 mg/kg halted fibrosis progression in the CDAHFD model. Conclusion: EDP-305 reduced fibrosis progression in rat BDL and mouse CDAHFD models. Molecular imaging of collagen and oxidized collagen is sensitive to changes in fibrosis and could be used to noninvasively measure treatment response in clinical trials. (Hepatology Communications 2018;2:821-835).

Li4Ti5O12/Ag composites were synthesized by a solid-state method. The effect of Ag modification on the physical and electrochemical properties is discussed by the characterizations of X-ray diffraction, scanning electron microscopy, cyclic voltammetry, electrochemical impedance spectroscopy, cycling and rate tests. The lattice parameter of Li4Ti5O12 with a low Ag content is almost not changed, but the lattice parameter becomes larger due to the high content of Ag. Li4Ti5O12/Ag material has a uniform particle size which is about 1 $\upmu$ m. Modification of appropriate Ag is beneficial to the reversible intercalation and deintercalation of Li + . Modification of Ag not only decreases the charge transfer resistance of Li4Ti5O12 material, but also improves the diffusion coefficient of lithium ion. Li4Ti5O12/Ag (3 mass%) material has the lowest charge transfer resistance, the highest diffusion coefficient of lithium ion and the best rate cycling performance.

Non-alcoholic steatohepatitis (NASH), characterized by hepatocyte injury, inflammation, and fibrosis, is the main cause of chronic liver disease in the Western world. There are currently no approved pharmacological therapies for NASH, underscoring the urgent need for effective treatments. The farnesoid X receptor (FXR) has emerged as an attractive target for the treatment of metabolic and chronic liver diseases. EDP-305 is an FXR agonist currently in phase 2 clinical trials for Primary Biliary Cholangitis (PBC) and NASH. Here, we demonstrate that EDP-305 is a selective and potent FXR agonist that regulates multiple pathways relevant to NASH progression. EDP-305 exhibits anti-fibrotic and anti-inflammatory gene signatures in human macrophage and stellate cell lines, as well as favorable effects on lipid metabolism in hepatocytes, including enhanced low density lipoprotein (LDL)-cholesterol uptake and decreased triglyceride accumulation. The therapeutic potential of EDP-305 was further evaluated in two murine models of NASH: a streptozotocin-high fat diet STAMTM model and a dietary induced NASH (DIN) model driven by high fat, cholesterol, and fructose feeding. In both NASH models, EDP-305 significantly decreased hepatocyte ballooning and lowered the non-alcoholic fatty liver disease (NAFLD) activity score. EDP-305 also significantly attenuated hepatic steatosis and dyslipidemia observed in the DIN mouse model. Conclusion: EDP-305 is a potent FXR agonist with a favorable gene expression profile for NASH treatment as evidenced by the hepato-protective and anti-steatotic effects observed in vivo. The preclinical characterization of EDP-305 presented here suggests that it holds promise for the treatment of NASH.

Microscale Li 4 Ti 5− x Zn x O 12 (0 ≤ x ≤ 0.2) particles with high phase purity were synthesized by a simple solid-state reaction. The effect of the zinc doping on the physicochemical properties of Li 4 Ti 5 O 12 (LTO) was extensively studied by TG-DSC, XRD, Raman spectroscopy, SEM, CV, EIS, and galvanostatic charge–discharge testing. The crystallization of lithium titanate oxide forms at about 750 °C. The lattice parameter of the Zn-doped LTO samples is slightly smaller than that for the pure LTO samples, and zinc doping does not change the basic Li 4 Ti 5 O 12 structure. Even though the material has a particle size of 1–2 μm, Zn-doped LTO shows very high excellent capacity retention between 0 and 2.5 V. Especially, in rate performance, the Li 4 Ti 4.8 Zn 0.2 O 12 sample maintains capacity of about 180 mAh g −1 until 5 C rates after 200 cycles, while the pure LTO sample shows severe capacity decline at corresponding rates. The reason for the high rate performance of Zn-doped LTO anode is ascertained to the diffusion coefficient ( D Li ) and reversible intercalation and deintercalation of lithium ion. The superior cycling performance and wide discharge voltage range, as well as simple synthesis route and low synthesis cost of the Zn-doped LTO are expected to show a potential commercial application.

A micro-sized particle Li 4 Ti 5− x La x O 12 (0 ≤ x ≤ 0.2) material has been synthesized by a simple solid-state method at air. The obtained Li 4 Ti 5− x La x O 12 materials are Li 3 x La 2/3− x TiO 3 (LLTO)–Li 4 Ti 5 O 12 (LTO) solid solution, and well crystallized with a particle size in the range of 1–2 μm. The electronic conductivity and lithium diffusion coefficient of La-modified LTO (Li 4 Ti 4.95 La 0.05 O 12 ) are improved because LLTO exhibits a high ionic conductivity during Li + extraction. Li 4 Ti 4.95 La 0.05 O 12 material shows discharge capacities of more than 206 and 197 mAh g −1 after 100 cycles at 1 C and 3 C charge–discharge rates, respectively. Especially, in rate performance, the Li 4 Ti 5− x La x O 12 ( x = 0.1, 0.2) samples maintain capacity of about 181 mAh g −1 until 5 C rates after 200 cycles, while the pure LTO sample shows a severe capacity decline at corresponding rate. These results suggest that La modifying is an effective way to improve the chemical stability of the electrode in contact with the electrolyte and improve their cyclability and rate capability during long term cycling. Since high rate performance is an important factor that needs to be considered in fabricating power batteries in industry, the La-modified LTO moves closer to real and large-scale applications.

To evaluate the efficacy of periprostatic nerve block anesthesia (PPNB) for pain relief in transrectal ultrasound-guided systematic prostate biopsy (PBx).We reviewed the data of patients undergoing initial PBx at our center from November, 2013 to January, 2015. Only the patients with 12-core systemic PBx were included and 111 patients were eligible for this study, among whom 52 patients received PPNB and 59 did not. PPNB was achieved by an injection of 5 mL of 1% lidocaine at the angle between the seminal vesicle and base of the prostate on each side before biopsy. The DRE pain score, probe insert pain score, and biopsy pain score were assessed by visual analogue scale (VAS) immediately after the biopsy. The complications were recorded and evaluated immediately after and at 7 days after the biopsy.The mean age, prostate volume, total prostate specific antigen (tPSA), free PSA (fPSA), and abnormal DRE were comparable between the 2 groups (P0.05). Immediately after the biopsy, no difference was found between the 2 groups in DRE pain score (1.40±0.98 vs 1.39±0.91, P=0.102) or probe insert pain score (2.07±0.96 vs 2.03±0.90, P=0.960), but the biopsy pain score was significantly lower in PPNB group than in no PPNB group (2.54±1.42 vs 3.07±1.43, P=0.033). The incidence of the procedure-related complications was similar between the 2 groups (P0.05).PPNB can significantly lower the biopsy pain score in PBx without increasing the incidence of complications.

EDP-239, a potent and selective hepatitis C virus (HCV) nonstructural protein 5A (NS5A) inhibitor developed for the treatment of HCV infection, has been investigated in vitro and in vivo . This study sought to characterize genotypic changes in the HCV NS5A sequence of genotype 1 (GT1) replicons and to compare those changes to GT1 viral RNA mutations isolated from clinical trial patients. Resistance selection experiments in vitro using a subgenomic replicon identified resistance-associated mutations (RAMs) at GT1a NS5A amino acid positions 24, 28, 30, 31, and 93 that confer various degrees of resistance to EDP-239. Key RAMs were similarly identified in GT1b NS5A at amino acid positions 31 and 93. Mutations F36L in GT1a and A92V in GT1b do not confer resistance to EDP-239 individually but were found to enhance the resistance of GT1a K24R and GT1b Y93H. RAMs were identified in GT1 patients at baseline or after dosing with EDP-239 that were similar to those detected in vitro . Baseline RAMs identified at NS5A position 93 in GT1, or positions 28 or 30 in GT1a only, correlated with a reduced treatment response. RAMs at additional positions were also detected and may have contributed to reduced EDP-239 efficacy. The most common GT1a and GT1b RAMs found to persist up to weeks 12, 24, or 48 were those at NS5A positions 28, 30, 31, 58 (GT1a only), and 93. Those RAMs persisting at the highest frequencies up to weeks 24 or 48 were L31M and Q30H/R for GT1a and L31M and Y93H for GT1b. (This study has been registered at ClinicalTrials.gov under identifier NCT01856426.)

RNF2 (ring finger protein 2) is frequently overexpressed in several types of human cancer, but the status of RNF2 amplification and expression in urothelial carcinoma of the bladder (UCB) and its clinical/prognostic significance is unclear. In this study, immunohistochemical analysis and fluorescence in situ hybridization (FISH) were used to examine the expression and amplification of RNF2 in 184 UCB patients after radical cystectomy. Overexpression of RNF2 was observed in 44.0% of UCBs and was found to significantly associate with shortened overall and cancer-specific survival (P P P RNF2 in 8/79 (10.1%) of informative UCB cases. Additionally, RNF2 overexpression was significantly associated with RNF2 gene amplification (P = 0.004) and cell proliferation (P = 0.003). These findings suggested that overexpression of RNF2, as examined by immunohistochemical analysis, might serve as a novel prognostic biomarker and potential therapeutic target for UCB patients who undergo radical cystectomy.

Trimethylation of lysine 27 on histone H3 (H3K27ME3) is a transcription-suppressive histone mark mediated by enhancer of zeste homolog 2 (EZH2). We have previously suggested that EZH2-mediated H3K27ME3 plays a critical oncogenic role in human hepatocellular carcinoma (HCC) aggressiveness. However, the direct downstream targets of EZH2-H3K27ME3 and the molecular mechanisms by which regulates HCC pathogenesis remain unclear. In this study, we used chromatin immunoprecipitation together with high-throughput sequencing (ChIP-seq) and gene expression profiling by microarray analysis to assess genome-wide chromatin occupancy of H3K27ME3 in HCC cells. We identified that claudin14 (CLDN14) is a potentially direct target for EZH2-mediated H3K27ME3 in HCC. In a large cohort of clinical HCC tissues, we found that low expression of CLDN14 was significantly associated with advanced tumor stage and determined to be an independent predictor of shortened survival of HCC patients. Next, functional experiment demonstrated that depletion of CLDN14 substantially restored EZH2-silenced HCC cells motility and invasive capacities and supported cell epithelial-mesenchymal transition (EMT). Furthermore, downregulation of CLDN14 dramatically re-enhanced the wnt/β-catenin signaling activity in EZH2-silenced HCC cells by increasing the levels of active β-catenin and promoting the nuclear localization of β-catenin. These results, collectively, uncover that CLDN14 is a novel direct target of EZH2-mediated H3K27ME3, and provide an explanation for the aggressive nature of HCC with downregulation of CLDN14 and the underling mechanism that links the tumor suppressor CLDN14 to the wnt/β-catenin signaling pathway.

Powders of spinel Li4Ti5O12 (LTO) were successfully synthesized at reducing conditions by solid-state method. The structure and physical properties of Li4Ti5O12 were examined by X-ray diffraction (XRD), Raman spectroscopy, scanning electronic microscopy (SEM), and differential capacitance, respectively. XRD shows that both samples are single-phase spinel compounds. LTO synthesized in Ar/H2(8% mol) has a larger lattice parameter than that in Ar. SEM indicates that all of the prepared powders have the uniform, nearly cubic structure morphology with narrow size distribution in the range of 200–300 nm. Raman spectra indicate that the Raman bands corresponding to the Ti–O vibration has a blue shift from 674 to 680 cm−1 due to the few H2 in the synthesized condition, indicating that there is very few oxygen vacancies in the Li4Ti5O12 synthesized under Ar/H2 (8% mol). The dQ/dV vs. voltage plots reveals the redox potentials for the synthesized Li4Ti5O12-negative electrode materials.

LiMn2O4 has been considered a promising cathode material for lithium-ion batteries in electric vehicles. However, there are still a number of problems of severe capacity fading before any materials modifications. Among all doped LiMn2O4, spinel LiNi0.5Mn1.5O4 material is seen as a potential cathode material for use in electric vehicles and energy storage systems in the future because of its high working potential (4.7 V), high energy density (the energy density of LiNi0.5Mn1.5O4 is 20% higher than that of LiCoO2), acceptable stability, and good cycling performance. In the presented paper, the structure and electrochemical performance of doped LiNi0.5Mn1.5O4 are reviewed. The rate capability, rate performance and cyclic life of various doped LiNi0.5Mn1.5O4 materials are described. This review also focuses on the present status of doped LiNi0.5Mn1.5O4, then on its near future developments.

LiFePO4-positive electrode material was successfully synthesized by a solid-state method, and the effect of storage temperatures on kinetics of lithium-ion insertion for LiFePO4-positive electrode material was investigated by electrochemical impedance spectroscopy. The charge-transfer resistance of LiFePO4 electrode decreases with increasing the storage temperatures. This suggests that it has a high electrochemical activity at high temperature. The diffusion coefficient of lithium ion is greatly increased with increasing the storage temperatures, indicating that the kinetics of Li+ and electron transfer into the electrodes were much fast at high storage temperature.

EDP-420 (EP-013420, S-013420) is a first-in-class bicyclolide (bridged bicyclic macrolide) currently in clinical development for the treatment of respiratory tract infections. It has good preclinical pharmacokinetic properties across multiple species and potent in vitro and in vivo activity against respiratory tract infection pathogens, including Haemophilus influenzae , atypical organisms (e.g., Chlamydia pneumoniae, Mycoplasma pneumoniae , and Legionella pneumophila ), and multidrug-resistant streptococci. The safety, tolerability, and pharmacokinetics of an orally administered EDP-420 suspension in 40 healthy adult subjects were assessed in a randomized, double-blind, placebo-controlled, ascending single-dose study. Eligible subjects were sequentially randomized into one of five study groups (i.e., 100-, 200-, 400-, 800-, or 1,200-mg dosing groups) consisting of eight subjects (six active and two placebo) each. EDP-420 was well tolerated. There were no serious adverse events reported, nor were there any dose-limiting clinical or laboratory adverse events reported. EDP-420 was rapidly absorbed after a single oral dose. The mean plasma terminal half-life ranged from 15.6 to 20.1 h with low clearance. At the 400-mg dose, the area under the curve was 14.4 μg·h/ml, which well exceeded the required area under the concentration-time curve to cover common respiratory tract infection pathogens based on preclinical pharmacokinetic/pharmacodynamic modeling. The long half-life and high systemic exposure of EDP-420 support once-daily dosing and may allow for shorter treatment durations compared to other macrolide antibiotics. Based on its human pharmacokinetic profiles, taken together with its in vitro/in vivo activity against common respiratory pathogens, EDP-420 warrants efficacy trials for the treatment of respiratory tract infections.

The origin of the nuclear tensor interaction in the covariant energy density functional (EDF) is presented in this work, associated with the Fock diagrams of Lorentz scalar and vector couplings. With this newly obtained relativistic formalism of the nuclear tensor interaction, more distinct tensor effects are found in the Fock diagrams of the Lorentz scalar and vector couplings, as compared to the Lorentz pseudo-vector and tensor channels. A unified and self-consistent treatment on both the nuclear tensor and spin-orbit interactions, which dominate the spin-dependent features of the nuclear force, is then achieved by the relativistic models. Moreover, careful analysis on the tensor strengths indicates the reliability of the nuclear tensor interaction in the covariant EDF for exploring the nuclear structure, excitation and decay modes., 5 pges, 3 figures

Since cellular zinc is not freely available as the inorganic ion, zinc proteins must acquire their metal from some other source. But how, when, and where they acquire it is unknown. Metallothionein can participate in the controlled delivery of zinc by binding it with high stability and by mobilizing it through a novel biochemical mechanism that critically depends on the redox activity of the zinc-sulfur bond. Thus, metallothionein activates zinc-depleted alcohol (sorbitol) dehydrogenases by glutathione-modulated zinc transfer. In addition to its catalytic, co-catalytic, and/or structural roles in a myriad of enzymes, zinc also inhibits some enzymes that are not necessarily zinc enzymes, e.g. glyceraldehyde and glycerol phosphate dehydrogenases, and aldehyde dehydrogenase. Zinc inhibits glycerol phosphate dehydrogenase with an IC(50) value of 100 nM. Zinc binding is slow at low pH, but instantaneous at high pH. Thionein, the apoprotein of metallothionein, re-activates the zinc-inhibited enzyme. Tight inhibition by zinc and activation of glycerol phosphate dehydrogenase by thionein, a biological chelating agent, provide further support that modulation of zinc binding by metallothionein and thionein is a physiological mechanism of enzyme regulation. Since glycerol phosphate dehydrogenase is a key enzyme in energy metabolism, the effect of zinc is expected to elicit significant physiological responses.

The release and transfer of zinc from metallothionein (MT) to zinc-depleted sorbitol dehydrogenase (EC 1.1.1.14 ) in vitro has been used to explore the role of MT in cellular zinc distribution. A 1:1 molar ratio of MT to sorbitol dehydrogenase is required for full reactivation, indicating that only one of the seven zinc atoms of MT is transferred in this process. Reduced glutathione (GSH) and glutathione disulfide (GSSG) are critical modulators of both the rate of zinc transfer and the ultimate number of zinc atoms transferred. GSSG increases the rate of zinc transfer 3-fold, and its concentration is the major determinant for efficient zinc transfer. GSH has a dual function. In the absence of GSSG, it inhibits zinc transfer from MT, indicating that MT is in a latent state under the relatively high cellular concentrations of GSH. In addition, it primes MT for the reaction with GSSG by enhancing the rate of zinc transfer 10-fold and by increasing the number of zinc atoms transferred to four. 65 Zn-labeling experiments confirm the release of one zinc from MT in the absence of glutathione and the more effective release of zinc in the presence of GSH and GSSG. In vivo , MT may keep the cellular concentrations of free zinc very low and, acting as a temporary cellular reservoir, release zinc in a process that is dynamically controlled by its interactions with both GSH and GSSG. These results suggest that a change of the redox state of the cell could serve as a driving force and signal for zinc distribution from MT.

Background Yes-associated protein 1 (YAP 1), the nuclear effector of the Hippo pathway, is a key regulator of organ size and a candidate human oncogene in multiple tumors. However, the expression dynamics of YAP 1 in urothelial carcinoma of the bladder (UCB) and its clinical/prognostic significance are unclear. Methods In this study, the methods of quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting and immunohistochemistry (IHC) were utilized to investigate mRNA/ protein expression of YAP 1 in UCBs. Spearman’s rank correlation, Kaplan-Meier plots and Cox proportional hazards regression model were used to analyze the data. Results Up-regulated expression of YAP 1 mRNA and protein was observed in the majority of UCBs by qRT-PCR and Western blotting, when compared with their paired normal bladder tissues. By IHC, positive expression of YAP 1 was examined in 113/213 (53.1%) of UCBs and in 6/86 (7.0%) of normal bladder specimens tissues. Positive expression of YAP 1 was correlated with poorer differentiation, higher T classification and higher N classification (P < 0.05). In univariate survival analysis, a significant association between positive expression of YAP 1 and shortened patients’ survival was found (P < 0.001). In different subsets of UCB patients, YAP 1 expression was also a prognostic indicator in patients with grade 2 (P = 0.005) or grade 3 (P = 0.046) UCB, and in patients in pT1 (P = 0.013), pT2-4 (P = 0.002), pN- (P < 0.001) or pT2-4/pN- (P = 0.004) stage. Importantly, YAP 1 expression (P = 0.003) together with pT and pN status (P< 0.05) provided significant independent prognostic parameters in multivariate analysis. Conclusions Our findings provide evidences that positive expression of YAP 1 in UCB may be important in the acquisition of an aggressive phenotype, and it is an independent biomarker for poor prognosis of patients with UCB.

We combined real-time RT-PCR and real-time PCR (R/P) assays using a hydrolysis probe to detect Mycobacterium tuberculosis complex (MTBC)-specific 16S rRNA and its rRNA gene (rDNA). The assay was applied to 28 nonrespiratory and 207 respiratory specimens from 218 patients. Total nucleic acids (including RNA and DNA) were extracted from samples, and results were considered positive if the repeat RT-PCR threshold cycle wasor =35 and the ratio of real-time RT-PCR and real-time PCR load wasor =1.51. The results were compared with those from existing methods, including smear, culture, and real-time PCR. Following resolution of the discrepant results between R/P assay and culture, the overall sensitivity, specificity, positive predictive values (PPV), and negative predictive values (NPV) of all samples (including nonrespiratory and respiratory specimens) were 98.2%, 97.2%, 91.7%, and 99.4%, respectively, for R/P assay, and 83.9%, 89.9%, 72.3%, and 94.7%, respectively, for real-time PCR. Furthermore, the R/P assay of four patient samples showed a higher ratio before treatment than after several days of treatment. We conclude that the R/P assay is a rapid and accurate method for direct detection of MTBC, which can distinguish viable and nonviable MTBC, and thus may guide patient therapy and public health decisions.

EDP-420 (also known as EP-013420, or S-013420) is a first-in-class bridged bicyclolide currently in clinical development for the treatment of respiratory tract infections (RTI) and has previously shown favorable pharmacokinetic (PK) and safety profiles after the administration of single oral doses of a suspension to healthy volunteers. Here we report its PK profile after the administration of multiple oral doses of a suspension to healthy adults. Bioequivalence between suspension and capsule formulations, as well as the effect of food, is also reported. The most important PK features of EDP-420 observed in these clinical studies are its long half-life of 17 to 18 h and its high systemic exposure, which support once-daily dosing and treatment durations potentially shorter than those of most other macrolide antibiotics. EDP-420 is readily absorbed following oral administration in both suspension and capsule formulations. In the multiple-oral-dose study, steady state was achieved on day 1 by using a loading dose of 400 mg/day, followed by 2 days of 200 mg/day. A high-fat meal had no effect on the bioavailability of EDP-420 administered in a capsule formulation. EDP-420 was well tolerated, with no serious or severe adverse events reported, and no subject was discontinued from the study due to an adverse event. Based on its human PK and safety profiles, together with its in vitro/in vivo activities against common respiratory pathogens, EDP-420 warrants further development, including trials for clinical efficacy in the treatment of RTI.