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Multi-target therapies have been considered one of the viable options to overcome the challenges to eradicate intrinsic and acquired drug-resistant cancer cells. While to increase the efficacy of therapeutics, the use of a single drug against multiple structurally similar sites, which noncommittedly modulate several vital cellular pathways proposed as a potential alternative to a 'single drug single target'. Besides, it reduces the usage of a number of drugs and their side effects. Topoisomerase II enzyme plays a very significant role in DNA replication and thus served as an important target for numerous anti-cancer agents. However, in spite of promising clinical results, in several cases, it was found that cancer cells have developed resistance against the anti-cancer agents targeting this enzyme. Therefore, multi-target therapies have been proposed as an alternative to overcome different drug resistance mechanisms while topoisomerases II are a primary target site. In this review, we have tried to discuss the characteristics of the binding cavity available for interactions of drugs, and potent inhibitors concurrently modulate the functions of topoisomerases II as well as other structurally related target sites. Additionally, the mechanism of drug resistance by considering molecular and cellular insights by including various types of cancers.

Extrahepatic arginases are postulated to be involved in cardiovascular-related pathologies by competing with nitric oxide synthase (NOS) for the common substrate l-arginine, subsequently decreasing nitric oxide production. However, previous models used to study arginase and NOS competition did not account for steady state level of l-arginine pool, which is dependent on conditions of l-arginine supply and utilization pathways. This work aimed at revisiting the concept of NOS and arginase competition while considering different conditions of l-arginine supply and l-arginine utilization pathways.Mouse macrophage-like RAW cells and human vascular endothelial cells co-expressing NOS and arginase were used to reevaluate the concept of substrate competition between arginase and NOS under conditions of l-arginine supply that mimicked either a continuous (similar to in vivo conditions) or a limited supply (similar to previous in vitro models). Enzyme kinetics simulation models were used to gain mechanistic insight and to evaluate the tenability of a substrate competition between the two enzymes. In addition to arginase and NOS, other l-arginine pathways such as transporters and utilization towards protein synthesis were considered to understand the intricacies of l-arginine metabolism. Our results indicate that when there is a continuous supply of l-arginine, as is the case for most cells in vivo, arginase does not affect NOS activity by a substrate competition. Furthermore, we demonstrate that l-arginine pathways such as transporters and protein synthesis are more likely to affect NOS activity than arginase.Arginase does not outcompete NOS for the common substrate l-arginine. Findings from this study should be considered to better understand the role of arginase in certain pathologies and for the interpretation of in vivo studies with arginase inhibitors.

Endoplasmic reticulum (ER) stress is enhanced in non-alcoholic steatohepatitis (NASH). Among three signalling pathways, the IRE1α/XBP1 signalling pathway is strongly implicated in the pathogenesis of NASH but its significance is still largely uncharacterised. In this report, we constructed a hepatocyte-specific XBP1-Luciferase knock-in mouse model that allows in vivo monitoring of the IRE1α/XBP1 activity in hepatocytes. Using this mouse model, we found that IRE1α/XBP1 was activated within hepatocytes during the pathogenesis of NASH. Significantly, a specific IRE1α kinase-inhibiting RNase attenuator, KIRA8, attenuated NASH in mice. In conclusion, our hepatocyte-specific XBP1 splicing reporter mouse represents a valid model for research and drug development of NASH, which showed that the IRE1α-induced XBP splicing is potentiated in hepatocytes during pathogenesis of NASH. Furthermore, we carried out the proof-of-concept study to demonstrate that the allosteric IRE1α RNase inhibitor serves as a promising therapeutic agent for the treatment of NASH.

Pancreatic lipase (PL) is the main digestive enzyme that is responsible for breaking triglycerides into smaller components for absorption. Inhibition of PL can effectively reduce triglyceride absorption, helping to prevent the development of obesity. The objective of this study was to investigate the PL inhibitory activity of peptides derived from sesame (Sesamum indicum L.) in silico and in vitro. In silico proteolysis of sesame proteins with pepsin, trypsin and chymotrypsin was performed with ExPASy PeptideCutter. Six peptides (TF, EW, QWM, NIF, AGY and PIF) were screened out by PeptideRanker, SwissADME and AutoDock. Molecular docking analysis showed that these six peptides could interact directly with Phe77, His151, Ser152, Phe215 and His263 at the key sites of PL. The six peptides were further synthesized to verify their PL-inhibitory effects in vitro, and TF, EW, QWM, NIF and AGY exhibited inhibitory activity on PL with IC

Pecan has been widely recognized for its high phenolic content and related health benefits. Previous studies indicated that pecan consumption might be beneficial in treating type 2 diabetes mellitus (T2DM). The objective of this study was to investigate the enzyme inhibitory activities of pecan phenolic compounds (PPC) and the effect in starch hydrolysis by in vitro simulation digestion. PPC was extracted with a solvent mixture from pecan powder and freeze-dried. PPC was tested for the inhibitory effects on α-amylase and α-glucosidase via enzyme kinetics study. Static in vitro digestion trials were conducted to evaluate the effect of intake of PPC and pecan powder on starch digestion. PPC displayed a potent inhibition effect against α-amylase and α-glucosidase with IC

Select protease inhibitors (PI) have been found to be effective in decreasing human papillomavirus oncoprotein expression. This study evaluated whether the topical PI, Saquinavir (SQV), promotes viral clearance in an infectious mouse model with Mus musculus papillomavirus 1 (MmuPV1). NOD scid gamma (NSG) mice were anally infected with ∼4 × 10sup8/supviral genome equivalents of MmuPV1 and 120 days post-infection (when majority have high-grade anal dysplasia), began topical treatments: control (mock), 7,12-dimethylbenz(a)anthracene (DMBA) only, once weekly to promote carcinogenesis, 1% SQV only, daily (Monday - Friday), and SQV + DMBA. Viral MmuPV1 load was analyzed from anal lavages pre and post-treatment. Anal tissue was harvested, processed, and evaluated for drug absorption, grade of anal disease, and anal viral RNA. Results suggest that topical SQV promotes decreased viral shedding in female mice treated with SQV.

Mutant isocitrate dehydrogenase (IDH) in chondrosarcoma produces the oncometabolite 2-hydroxyglutarate (2-HG) and contributes to malignant progression, and is therefore a potential therapeutic target for chondrosarcoma. Robust historical control data are important in clinical trials of rare cancers such as chondrosarcoma in order to show a clear benefit of new drugs. However, it remains controversial whether IDH mutation status is associated with the clinical outcome of chondrosarcoma, and this hinders the development of mutant IDH inhibitors in clinical trials.background METHODS: We investigated the relationship between IDH gene status and clinicopathological data in 38 chondrosarcoma patients from whom frozen tumor samples were obtained at the time of biopsy or surgery. Targeted next-generation sequencing was also performed to compare genetic alterations between patients with and without IDH mutations.The results revealed 15 cases (40%) of heterozygous IDH1 mutations and five cases (13%) of IDH2 mutations. IDH-mutant chondrosarcoma was associated with worse overall survival than IDH-wild-type chondrosarcoma (IDH1/2 Mut vs. IDH Wt, P = 0.006; IDH1 Mut vs. IDH Wt, P = 0.030; IDH2 Mut vs. IDH Wt, P 0.0001). IDH mutation was also a significant poor prognostic factor both in univariate (P = 0.026) and multivariate (P = 0.048) analyses. Targeted next-generation sequencing revealed that characteristic mutations in chondrosarcoma, including TP53 and COL2A1, were more common in the IDH-mutant group than in the IDH-wild-type group.results CONCLUSION: This study is the first to report in detail the characteristics and clinical courses of IDH-mutant chondrosarcoma patients in Japan. Our data suggested that IDH-mutant chondrosarcomas might have a worse prognosis than that of IDH-wild-type chondrosarcoma, possibly through the more aggressive characters after metastasis. This information will be useful for designing clinical trials of mutant IDH inhibitors for treatment of advanced chondrosarcoma.

Inhibition of the bromodomain of the CREB (cyclic-AMP response element-binding protein) binding protein (CBP) is a particularly promising new therapeutic approach for cancer. Benzimidazole derivatives CCS1477 and its analogues (8 and 9) are highly potent and selective CBP bromodomain inhibitors, with K

The "astrocyte-to-neuron lactate shuttle" (ANLS) mechanism is part of the central inhibitory pathway to modulate sodium intake. An interaction between the GABAergic neurons and nitric oxide (NO) in the subfornical organ (SFO) in salt-appetite inhibition has been suggested. In addition, NO is a key molecule involved in astrocytic energy metabolism and lactate production. In the present study, we hypothesized there is an interaction between astrocytic lactate and central NO to negatively modulate water and sodium intake through the ANLS mechanism. The results showed that central Nω-nitro-L-arginine methyl ester (L-NAME, NO-synthase inhibition) induced an increase in water and sodium intake. These responses were attenuated by previous central microinjection of fluorocitrate (FCt, a reversible glial inhibitor). Interestingly, L-NAME-induced water and sodium intake were also decreased by previous microinjection of lactate but did not change after inhibition of the ANLS mechanism by α-cyano 4-hydroxycinnamic acid (α-CHCA), an inhibitor of the MCT lactate transporter. Our results suggest a central interaction between NO, glial cells, and lactate to modulate water and sodium intake.

Smilax china L. showed various biological activities mainly due to its phenolic components; however, the mechanism of isolated phenolic fraction against xanthine oxidase (XO) has not been investigated. Quercetin-3-O-rhamnoside (QORh) and chlorogenic acid (CGA) extracted from Smilax china L. ethyl acetate fraction was analyzed for its XO inhibitory kinetics and mechanism using multispectroscopic methods and molecular docking techniques. QORh and CGA reversibly inhibited XO activity in competitive and non-competitive modes, respectively. The bioactive compounds bound with XO were dominated mainly by hydrogen bonds and van der Waals forces to form QORh-XO, and CGA-XO complexes with one affinity binding site. The synchronous fluorescence, circular dichroism, three-dimensional (3D) fluorescence, and Fourier transform infrared spectra exhibited that XO binding with QORh or CGA leads to the secondary and tertiary structural variation of the protein. Additionally, molecular docking further revealed that QORh binds to the active site of XO and forms hydrogen coupling with amino acid residues. The results showed that QORh and CGA had inhibitory activity on XO, which might be further used to modify the bioactive compounds and improve their efficacy to treat gout.

Dysregulation of EZH2 has a crucial role in lymphomagenesis. We did a first-in-human study to assess the safety, pharmacokinetics, pharmacodynamics, and preliminary clinical activity of SHR2554, an oral EZH2 inhibitor, in patients with relapsed or refractory mature lymphoid neoplasms, including B-cell lymphomas, T-cell lymphomas, and classical Hodgkin lymphoma.This was a multicentre, dose-escalation, dose-expansion, and clinical expansion phase 1 study done at 13 hospitals in China. Eligible patients had histologically or cytologically confirmed mature lymphoid neoplasms that had relapsed or were refractory to standard systemic therapies or had no standard-of-care. The study included a dose-escalation phase, at doses of SHR2554 from 50 mg to 800 mg twice daily; a dose-expansion phase, at two selected doses; and a subsequent clinical expansion phase at the recommended phase 2 dose in selected tumours. Primary endpoints were the safety, maximum tolerated dose, and recommended phase 2 dose. Objective response rate was a secondary endpoint. Safety and activity were assessed in all patients who received at least one dose of SHR2554 and had at least one post-baseline evaluation. This study is registered with ClinicalTrials.gov, NCT03603951, and follow-up is ongoing.Between Aug 14, 2018, and July 13, 2021, 113 patients received SHR2554. At data cutoff (Sept 10, 2021), the median follow-up duration was 7·0 months (IQR 3·7-12·0). 71 (63%) patients were men and 42 (37%) were women, 110 (97%) were of Han ethnicity and 3 (3%) of other ethnicities, and 53 (47%) had received three or more lines of previous anticancer therapies. Dose-limiting toxicities occurred in two (67%) of three patients who received 400 mg SHR2554 twice daily and one (17%) of six patients who received 350 mg SHR2554 twice daily. The maximum tolerated dose and recommended phase 2 dose was determined to be 350 mg twice daily. The most common grade 3 or 4 treatment-related adverse events in all 113 patients were decreased platelet count (20 [18%]), decreased neutrophil count (ten [9%]), decreased white blood cell count (nine [8%]), and anaemia (seven [6%]). 18 (16%) patients had serious treatment-related adverse events. Two patients (2%) died due to treatment-related adverse events: one (1%) due to skin infection and toxic epidermal necrolysis and one (1%) due to respiratory failure. 107 (95%) of the 113 enrolled patients had post-baseline assessments for tumour response and were included in the activity analysis. 46 (43%; 95% CI 33-53) of these 107 patients had an overall response.SHR2554 showed an acceptable safety profile and promising antitumour activity in patients with relapsed or refractory lymphomas, providing evidence for future investigations.Jiangsu Hengrui Pharmaceuticals.For the Chinese translation of the abstract see Supplementary Materials section.

Prolyl endopeptidase or prolyl oligopeptidase (PEP or POP) is highly expressed in brain, and associated with autism spectrum disorders, dementia, aging and various psychological disorders, such as schizophrenia, mania, and neurodegeneration. To design highly potent and novel POP inhibitors, structure-based virtual screening was carried out using pharmacophore modeling and molecular docking studies. The docking based active compounds [incensole (1), incensole acetate (2), incensone (3), incensfuran (4), and epi-incensole acetate (5)] were selected and their dynamic behavior was studied through molecular dynamic simulation. Later, the top-ranked [predicted active, (1-5)] and lower-ranked [predicted in-active, (6-10)] compounds were tested by in-vitro assay. The in-vitro results showed that all top-ranked compounds (1-5) found significantly active against POP enzyme with IC

Treatment options for malignant pleural mesothelioma are scarce. Tazemetostat, a selective oral enhancer of zeste homolog 2 (EZH2) inhibitor, has shown antitumour activity in several haematological cancers and solid tumours. We aimed to evaluate the anti-tumour activity and safety of tazemetostat in patients with measurable relapsed or refractory malignant pleural mesothelioma.We conducted an open-label, single-arm phase 2 study at 16 hospitals in France, the UK, and the USA. Eligible patients were aged 18 years or older with malignant pleural mesothelioma of any histology that was relapsed or refractory after treatment with at least one pemetrexed-containing regimen, an Eastern Cooperative Oncology Group performance status of 0 or 1, and a life expectancy of greater than 3 months. In part 1 of the study, participants received oral tazemetostat 800 mg once on day 1 and then twice daily from day 2 onwards. In part 2, participants received oral tazemetostat 800 mg twice daily starting on day 1 of cycle 1, using a two-stage Green-Dahlberg design. Tazemetostat was administered in 21-day cycles for approximately 17 cycles. The primary endpoint of part 1 was the pharmacokinetics of tazemetostat and its metabolite at day 15 after administration of 800 mg tazemetostat, as measured by maximum serum concentration (CBetween July 29, 2016, and June 2, 2017, 74 patients were enrolled (13 in part 1 and 61 in part 2) and received tazemetostat, 73 (99%) of whom had BAP1-inactivated tumours. In part 1, following repeat dosing of tazemetostat at steady state, on day 15 of cycle 1, the mean CFurther refinement of biomarkers for tazemetostat activity in malignant pleural mesothelioma beyond BAP1 inactivation could help identify a subset of tumours that are most likely to derive prolonged benefit or shrinkage from this therapy.Epizyme.

Flavivirus, such as Dengue Virus (DENV) and Zika virus (ZIKV), infects millions of people and cause the death of thousands of people every year. Despite many efforts, there is no approved anti-flaviviral treatment available. In particular, some antiflavivirus compounds were investigated the cellular activities of DENV and ZIKV, but lacking the exploration of specific target enzyme, thereby resulting in the hindrance of structure-based drug design. One example is Montlukast, which was found to inhibit the replicon replication in DENV and ZIKV infected cells, with EC

Lysine specific demethylase 1 (LSD1), a transcriptional corepressor or coactivator that serves as a demethylase of histone 3 lysine 4 and 9, has become a potential therapeutic target for cancer therapy. LSD1 mediates many cellular signaling pathways and regulates cancer cell proliferation, invasion, migration, and differentiation. Recent research has focused on the exploration of its pharmacological inhibitors. Natural products are a major source of compounds with abundant scaffold diversity and structural complexity, which have made a major contribution to drug discovery, particularly anticancer agents. In this review, we briefly highlight recent advances in natural LSD1 inhibitors over the past decade. We present a comprehensive review on their discovery and identification process, natural plant sources, chemical structures, anticancer effects, and structure-activity relationships, and finally provide our perspective on the development of novel natural LSD1 inhibitors for cancer therapy.

The progression of chronic kidney disease (CKD) increases the risks of cardiovascular morbidity and end-stage kidney disease. Indoxyl sulfate (IS), which is derived from dietary l-tryptophan by the action of bacterial l-tryptophan indole-lyase (TIL) in the gut, serves as a uremic toxin that exacerbates CKD-related kidney disorder. A mouse model previously showed that inhibition of TIL by 2-aza-l-tyrosine effectively reduced the plasma IS level, causing the recovery of renal damage. In this study, we found that (+)-sesamin and related lignans, which occur abundantly in sesame seeds, inhibit intestinal bacteria TILs. Kinetic studies revealed that (+)-sesamin and sesamol competitively inhibited Escherichia coli TIL (EcTIL) with K

Pyruvate dehydrogenase kinase 1 (PDK1) is a Ser/Thr kinase that inactivates mitochondrial pyruvate dehydrogenase (PDH), leading to switch of glucose metabolism from mitochondrial oxidation to aerobic glycolysis. We previously reported that PDK1 inhibition is a potent therapeutic strategy in multiple myeloma (MM). However, availability of PDK1 inhibitors, which are effective at low concentrations, are limited at present, making PDK1 inhibition difficult to apply in the clinic. In the present study, we examined the efficacy and mechanism of action of JX06, a novel PDK1 inhibitor, against MM cells. We confirmed that PDK1 is highly expressed in normal plasma cells and MM cells using publicly available gene expression datasets. JX06 suppressed cell growth and induced apoptosis against MM cells from approximately 0.5 μM JX06 treatment reduced PDH phosphorylation, suggesting that JX06 is indeed inhibiting PDK1. Intracellular metabolite analysis revealed that JX06 treatment reduced metabolites associated with glucose metabolism of MM cells. Additionally, JX06 in combination with a well-known proteasome inhibitor, bortezomib, significantly increased MM cell death, which raises the possibility of combination use of JX06 with proteasome inhibitors in the clinic. These findings demonstrate that PDK1 can be potentially targeted by JX06 in MM through glycolysis inhibition, leading to a novel therapeutic strategy in MM.

In Brazil, the major vector of arboviruses is Aedes aegypti, which can transmit several alpha and flaviviruses. In this work, a pacifastin protease inhibitor library was constructed and used to select mutants for Ae. aegypti larvae digestive enzymes. The library contained a total of 3.25 × 10

Protein tyrosine phosphatase 1B (PTP1B) is a member of the phosphotyrosine phosphatase family and plays an important role in the signal transduction of diabetes. Inhibition of PTP1B activity can increase insulin sensitivity and reduce blood sugar levels. Therefore, it is urgent to find compounds with novel structures that can inhibit PTP1B. This study designed imidazolidine-2,4-dione derivatives through the computer-aided drug design (CADD) strategy, and the Comp#10 showed outstanding inhibitory ability. (IC50 = 2.07 μM) and selectivity. The inhibitory mechanism at molecular level of Comp#10 on PTP1B was studied by molecular dynamics simulation. The results show that the catalytic region of PTP1B protein is more stable, which makes the catalytic sites unsuitable for exposure. Interestingly, the most obvious changes in the interaction between residues in the P-loop region (such as: His214, Cys215, and Ser216). In short, this study reported for the first time that imidazolidine-2,4-dione derivatives as novel PTP1B inhibitors had good inhibitory activity and selectivity, providing new ideas for the development of small molecule PTP1B inhibitors.

The structural evolution of lotus starch (LS)-chlorogenic acid (CA) complexes was investigated after microwave-heating treatment, to reveal the relationship between the interactions of lotus starch and chlorogenic acid molecules, and the digestive properties of the starch, after microwave gelatinization. During the early stage of microwave gelatinization (65, 70 °C), CA was mainly participating in the rearrangement of starch molecules in a weakly-bound form, and at that stage, the LS-CA complex acted as an inhibitor of digestion, under small intestine conditions, mainly through the release of CA, which inhibited amylase. However, during the late stage of microwave gelatinization (85 °C), many chlorogenic acid molecules entered the hydrophobic helical cavity of the starch, promoting formation of the V-type starch helical structure in the LS-CA complex, which made a major contribution to inhibiting digestion under oral digestion conditions.

Introduction Carpolobia lutea root extract (CLRE) has been reported to enhance penile erection. However, the mechanism involved is poorly understood. We investigated in vitro mechanisms of CLRE action on contractile activity of rabbit corpus cavernosum (CC). Methods Corpus cavernosum strips from four healthy male New Zealand rabbits (2.5–3.0 kg) were mounted on an organ chamber and contracted with phenylephrine (PE) (10−9 to 10−5 M) and Potassium Chloride (KCl) (10–50 mM) before treatment with various concentrations of CLRE (0.1–1.2 mg/ml). Interactions between CLRE and a Nitric Oxide Synthase (NOS) inhibitor (N-nitro- l -arginine methyl ester – l -NAME 10−4 M); guanylyl cyclase inhibitors (Oxalodiazolo 4,3-a quinoxalin-1-one – ODQ 10 μM, 20 μM, 30 μM), and (methylene blue 10–30 μM); a cyclooxygenase inhibitor (10−4 M indomethacin); potassium-channel inhibitors (100 μM tetraethyl ammonium TEA), (100 ηM apamin) and (glibenclamide 10 μM and 20 μM); and a calcium-channel inhibitor (−10−4 M nifedipine) were investigated. Results Maximal contractions of KCl and PE contracted CC strips were significantly reduced in a concentration-dependent manner (40.8 ± 3.6% and 38.6 ± 4.0% from 64.6 ± 2.9% and 98.1 ± 4.2% respectively). Relaxant effect of CLRE was significantly reduced by ODQ (38.6 ± 4.0% to 6.4 ± 1.3% and 38.6 ± 4.0% to 7.2 ± 1.2%), nifedipine (38.6 ± 4.0% to 21.1 ± 2.7%) and glibenclamide (40.8 ± 3.6% to 31.5 ± 3.3%). However l -NAME, indomethacin, methylene blue, TEA and apamin did not inhibit relaxation by CLRE. Conclusion Concentration-dependent relaxant effect of CLRE in rabbit CC involves the soluble guanylate cyclase/cyclase Guanosine Monophosphate system, and activation of ATP-dependent K+ channels.

Thymidylate synthase (TS) is a key enzyme in nucleotide biosynthesis. A study performed by our group on human monocyte-derived macrophages (MDMs) infected with HIV-1 showed that many enzymes related to the folate cycle pathway, such as TS, are upregulated in productively infected cells. Here, we suggest that TS is essential for an effective HIV-1 infection in MDMs. Indeed, a TS specific small interfering RNA (siRNA) as well as the TS specific inhibitor Raltitrexed (RTX) caused a reduction in productively infected cells. Quantitative PCR analysis showed that this treatment decreased the efficacy of the early steps of the viral cycle. The RTX inhibitory effect was counteracted by dNTP addition. These results suggest that TS is essential for the early stages of HIV-1 infection by providing optimal dNTP concentrations in MDMs. TS and its related pathway may thus be considered as a potential therapeutic target for HIV-1 treatment.

To evaluate changes in health-related and vision-related quality of life (VRQoL) among patients with noninfectious uveitis who were treated with antimetabolites.Secondary analysis of a randomized controlled trial.Patients with noninfectious uveitis from India, the United States, Australia, Saudi Arabia, and Mexico.From 2013 through 2017, 216 participants were randomized to receive 25 mg weekly oral methotrexate or 1.5 g twice daily oral mycophenolate mofetil. Median changes in quality of life (QoL) were measured using Wilcoxon signed-rank tests, and differences between treatment groups were measured using linear mixed models, adjusting for baseline QoL score, age, gender, and site. Among Indian patients, VRQoL scores from a general scale (the National Eye Institute Visual Function Questionnaire [NEI-VFQ]) and a culturally specific scale (the Indian Visual Function Questionnaire [IND-VFQ]) were compared using Pearson correlation tests.Vision-related QoL (NEI-VFQ and IND-VFQ) and health-related QoL (HRQoL; physical component score [PCS] and mental component score [MCS] of the Medical Outcomes Study 36-Item Short Form Survey [SF-36v2]) were measured at baseline, the primary end point (6 months or treatment failure before 6 months), and the secondary end point (12 months or treatment failure between 6 and 12 months).Among 193 participants who reached the primary end point, VRQoL increased from baseline by a median of 12.0 points (interquartile range [IQR], 1.0-26.1, NEI-VFQ scale), physical HRQoL increased by a median of 3.6 points (IQR, -1.4 to 14.9, PCS SF-36v2), and mental HRQoL increased by a median of 3.0 points (IQR, -3.7 to 11.9, MCS SF-36v2). These improvements in NEI-VFQ, SF-36v2 PCS, and SF-36v2 MCS scores all were significant (P0.01). The linear mixed models showed that QoL did not differ between treatment groups for each QoL assessment (NEI-VFQ, IND-VFQ, PCS SF-36v2, and MCS SF-36v2; P0.05 for all). The NEI-VFQ and IND-VFQ scores for Indian participants were correlated highly at baseline and the primary and secondary end points (correlation coefficients, 0.87, 0.80, and 0.90, respectively).Among patients treated with methotrexate or mycophenolate mofetil for uveitis, VRQoL and HRQoL improved significantly over the course of 1 year and did not differ by treatment allocation. These findings suggest that antimetabolites could improve overall patient well-being and daily functioning.

Epigenetic enzyme-targeted therapy is a promising new development in the field of drug discovery. To date, histone deacetylases and DNA methyltransferases have been investigated as druggable epigenetic enzyme targets in cancer therapeutics. Histone methyltransferases and lysine demethylase inhibitors are the latest groups of epi-drugs being actively studied in clinical trials. KDM4s are JmjC domain-containing histone H3 lysine 9/36 demethylase enzymes, belonging to the 2-OG-dependent oxygenases, which are upregulated in multiple malignancies. In the recent years, these enzymes have captured much attention as a novel target in cancer therapy. Herein, we traverse the discovery path and current challenges in designing potent KDM4 inhibitors as potential anticancer agents. We discuss the considerable efforts and proposed future strategies to develop selective small molecule inhibitors of KDM4s, highlighting scaffold candidates and cyclic skeletons for which activity data, selectivity profiles and structure-activity relationships (SARs) have been studied.

In this study, the inhibitory activities of eight caffeoylquinic acids (CQAs) against xanthine oxidase (XOD) in vitro were investigated, and the interaction mechanisms between each compound and XOD were studied. HPLC and fluorescence spectra showed that the inhibitory activities of dicaffeoylquinic acids (diCQAs) were higher than that of monocaffeoylquinic acids (monoCQAs), due to the main roles of hydrophobic interaction and hydrogen bond between XOD and diCQAs. Both the binding constant and the lowest binding energy data indicated that the affinities of diCQAs to XOD were stronger than that of monoCQAs. Circular dichroism showed that the structure of XOD was compacted with the increased of α-helix content, resulting in decreased enzyme catalytic activity. Molecular docking revealed that CQAs preferentially bind to the flavin adenine dinucleotide region in XOD. These results provided the mechanisms of CQAs on inhibiting XOD and the further utilization of CQAs as XOD inhibitors to prevent hyperuricemia.

Enhancement of fibrinolysis constitutes a promising approach to treat thrombotic diseases. Venous thrombosis and thromboembolism risks are associated with increased plasma levels of TAFI (Thrombin Activatable Fibrinolysis Inhibitor) as well as its active form TAFIa. A new TAFIa inhibitor, namely S62798 has been identified. Its ability to enhance fibrinolysis was investigated both in vitro and in vivo in a mouse model of pulmonary thromboembolism, as well as its effect on bleeding. S62798 is a highly selective human, mouse and rat TAFIa inhibitor (IC50 = 11; 270; 178 nmol/L, respectively). It accelerates lysis of a human clot in vitro, evaluated by thromboelastometry (EC50 = 27 nmol/L). In a rat tail bleeding model, no effect of S62798 treatment was observed up to 20 mg/kg. Enhancement of endogenous fibrinolysis by S62798 was investigated in a mouse model of Tissue Factor-induced pulmonary thromboembolism. Intravenous administration of S62798 decreased pulmonary fibrin clots with a minimal effective dose of 0.03 mg/kg. Finally, effect of S62798 in combination with heparin was evaluated. When treatment of heparin was done in a curative setting, no effect was observed whereas a significantly decreased pulmonary fibrin deposition was observed in response to S62798 alone or in combination with heparin. This study demonstrates that S62798 is a potent TAFIa inhibitor with minimal risk of bleeding. In vivo, curative S62798 intravenous treatment, alone or associated with heparin, accelerated clot lysis by potentiating endogenous fibrinolysis and thus decreased pulmonary fibrin clots. S62798 is expected to be a therapeutic option for pulmonary embolism patients on top of anticoagulants.

BAFF and APRIL regulate B cell homeostasis by binding to their three receptors BAFFR, BCMA and TACI. The complexity of this system is further increased by shedding of these three receptors; this reduces signaling due to the display of less surface receptors. Further, soluble forms, sBCMA and sTACI, were detected in body fluids and serve as biomarker in malignancies, autoimmune diseases and immunodeficiencies. sBCMA and sTACI function as decoys blocking BAFF and APRIL. BCMA is a promising therapeutic target in multiple myeloma, but sBCMA may reduce therapeutic activity of CAR T cells, bispecific antibodies, and antibody-drug conjugates. Insights into the biochemical mechanism of shedding of BCMA can be harnessed to improve BCMA-directed therapy by blocking its shedding with a γ-secretase inhibitor.

Phosphorylation of RNA polymerase II (RNAP II) coordinates the temporal progression of eukaryotic transcription. The development and application of chemical genetic methods have enhanced our ability to investigate the intricate and intertwined pathways regulated by the kinases and phosphatases targeting RNAP II to ensure transcription accuracy and efficiency. Although identifying small molecules that modulate these enzymes has been challenging due to their highly conserved structures, powerful new chemical biology strategies such as targeted covalent inhibitors and small molecule degraders have significantly improved chemical probe specificity. The recent success in discovering phosphatase holoenzyme activators and inhibitors, which demonstrates the feasibility of selective targeting of individual phosphatase complexes, opens up new avenues into the study of transcription. Herein, we summarize how chemical biology is used to delineate kinases' identities involved in RNAP II regulation and new concepts in inhibitor/activator design implemented for kinases/phosphatases involved in modulating RNAP II-mediated transcription.

Nucleosome remodeling provides access to genomic DNA for recruitment of the transcriptional machinery to mediate gene expression. The aberrant function of nucleosome remodeling complexes has been correlated to human cancer, making them emerging therapeutic targets. The bromodomain PHD finger transcription factor, BPTF, is the largest member of the human nucleosome remodeling factor NURF. Over the last five years, BPTF has become increasingly identified as a protumorigenic factor, prompting investigations into the molecular mechanisms associated with BPTF function. Despite a druggable bromodomain, small molecule discovery is at an early stage. Here we highlight recent investigations into the biology being discovered for BPTF, chemical biology approaches used to study its function, and small molecule inhibitors being designed as future chemical probes and therapeutics.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) represents a significant threat to human health. Despite its similarity to related coronaviruses, there are currently no specific treatments for COVID-19 infection, and therefore there is an urgent need to develop therapies for this and future coronavirus outbreaks. Formation of the cap at the 5′ end of viral RNA has been shown to help coronaviruses evade host defenses. Nonstructural protein 14 (nsp14) is responsible for N7-methylation of the cap guanosine in coronaviruses. This enzyme is highly conserved among coronaviruses and is a bifunctional protein with both N7-methyltransferase and 3′-5′ exonuclease activities that distinguish nsp14 from its human equivalent. Mutational analysis of SARS-CoV nsp14 highlighted its role in viral replication and translation efficiency of the viral genome. In this paper, we describe the characterization and development of a high-throughput assay for nsp14 utilizing RapidFire technology. The assay has been used to screen a library of 1771 Food and Drug Administration (FDA)-approved drugs. From this, we have validated nitazoxanide as a selective inhibitor of the methyltransferase activity of nsp14. Although modestly active, this compound could serve as a starting point for further optimization.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus responsible for the global COVID-19 pandemic. Nonstructural protein 14 (NSP14), which features exonuclease (ExoN) and guanine N7 methyltransferase activity, is a critical player in SARS-CoV-2 replication and fidelity and represents an attractive antiviral target. Initiating drug discovery efforts for nucleases such as NSP14 remains a challenge due to a lack of suitable high-throughput assay methodologies. This report describes the combination of self-assembled monolayers and matrix-assisted laser desorption ionization mass spectrometry to enable the first label-free and high-throughput assay for NSP14 ExoN activity. The assay was used to measure NSP14 activity and gain insight into substrate specificity and the reaction mechanism. Next, the assay was optimized for kinetically balanced conditions and miniaturized, while achieving a robust assay (Z factor > 0.8) and a significant assay window (signal-to-background ratio > 200). Screening 10,240 small molecules from a diverse library revealed candidate inhibitors, which were counterscreened for NSP14 selectivity and RNA intercalation. The assay methodology described here will enable, for the first time, a label-free and high-throughput assay for NSP14 ExoN activity to accelerate drug discovery efforts and, due to the assay flexibility, can be more broadly applicable for measuring other enzyme activities from other viruses or implicated in various pathologies.

Human fatty acid synthase (FASN) is the sole cytosolic enzyme responsible for de novo lipid synthesis. FASN is essential for cancer cell survival and contributes to drug and radiation resistance by up-regulating DNA damage repair but not required for most non-lipogenic tissues. Thus, FASN is an attractive target for drug discovery. However, despite decades of effort in targeting FASN, no FASN inhibitors have been approved due to poor pharmacokinetics or toxicities. Here, we show that the FDA-approved proton pump inhibitors (PPIs) effectively inhibit FASN and suppress breast cancer cell survival. PPI inhibition of FASN leads to suppression of non-homologous end joining repair of DNA damages by reducing FASN-mediated PARP1 expression, resulting in apoptosis from oxidative DNA damages and sensitization of cellular resistance to doxorubicin and ionizing radiation. Mining electronic medical records of 6754 breast cancer patients showed that PPI usage significantly increased overall survival and reduced disease recurrence of these patients. Hence, PPIs may be repurposed as anticancer drugs for breast cancer treatments by targeting FASN to overcome drug and radiation resistance.

Proteolysis-targeting chimeras (PROTACs) that degrade disease-causing proteins by hijacking the endogenous ubiquitin-proteasome system have emerged as an exciting and transformative technology in both chemical biology and drug discovery. Currently, the majority of PROTACs use reversible non-covalent ligands for both the target protein of interest (POI) and E3 ligase. In this review, we explore the burgeoning role of reversible and irreversible covalent chemistry in targeted protein degradation. We highlight the key advantages of targeted covalent inhibitors, whether as the target POI or E3 ligase ligand, such as their ability to enhance the selectivity of PROTACs, enable access to more of the "undruggable" proteome and expand the repertoire of recruited E3 ligases.

We have previously demonstrated that hydrogen sulfide (H2S), the third endogenous gasotransmitter, ameliorates the depression- and anxiety-like behaviors in diabetic rats, but the underlying mechanism remains unclear. The present was aimed to investigate whether the hippocampal phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway mediates H2S-ameliorated depression- and anxiety-like behaviors in diabetic rats by improving the hippocampal neurogenesis. The depression-like behaviors were examined by Tail suspension test (TST), the anxiety-like behaviors were examined by Elevated plus maze test (EPM), and the locomotor activity was detected by Open Field Test (OFT). The expressions of doublecortin (DCX), neuron-specific nuclear protein (NeuN), glial fibrillary acidic protein (GFAP), p-AKT, and AKT in the hippocampus were determined by Western blot analysis. Results showed that NaHS, a donor of exogenous H2S, not only activated the hippocampal PI3K/AKT pathway, as evidenced by the increase of phosphorylated AKT, but also favorably reversed streptozotocin (STZ)-disturbed hippocampal neurogenesis, as evidenced by the increases in the expressions of DCX and NeuN as well as the decrease in the expression of GFAP in the hippocampus of STZ-induced diabetic rats. Furthermore, inhibited PI3K/AKT pathway by LY294002 significantly abolished H2S-exerted the improvement of hippocampal neurogenesis and the antidepressant- and anxiolytic-like effects in the STZ-induced diabetic rats. Taken together, these results uncover that the activation of hippocampal PI3K/AKT pathway plays an important role to restore hippocampal neurogenesis and subsequently to mediate the antidepressant- and anxiolytic-like roles of H2S in STZ-induced diabetic rats and enhance our understanding of the robustness of H2S as a therapeutic strategy for treatment of depression in diabetes mellitus.

Deubiquitinating enzymes (DUBs) are a largely understudied and untapped resource in the toolkit of protein degradation functionalities. They comprise a large repertoire of enzymes that remove ubiquitin from substrates in a variety of cellular and pathophysiological contexts, and have enormous potential for research and clinical use. It is only within the last 5 years that potent, selective, and well-characterized small-molecule inhibitors of DUBs have been described. These compounds are now being used to study the biological roles of DUBs. Here, we describe downstream applications of small-molecule inhibitors for studying DUBs and provide a framework for future studies. We highlight recent examples of using these inhibitors to confirm and explore the role of these enzymes in both normal and pathological contexts. These studies represent the first steps in the burgeoning field of pharmacological and chemoproteomic studies of DUBs, which will be critical for the continued advancement of DUB field.

For centuries, dietary ginger has been known for its antioxidant, anticancer, and antibacterial properties. In the current study, we examined the link between antibacterial properties of 7 dietary ginger phenolics (DGPs)-gingerenone A, 6-gingerol, 8-gingerol, 10-gingerol, paradol, 6-shogaol, and zingerone-and inhibition of bacterial ATP synthase. DGPs caused complete (100%) inhibition of wild-type Escherichia coli membrane-bound F1Fo ATP synthase, but partial and variable (0%-87%) inhibition of phytochemical binding site mutant enzymes αR283D, αE284R, βV265Q, and γT273A. The mutant enzyme ATPase activity was 16-fold to 100-fold lower than that of the wild-type enzyme. The growth of wild-type, null, and mutant strains in the presence of the 7 DGPs were abrogated to variable degrees on limiting glucose and succinate media. DGPs-caused variable inhibitory profiles of wild-type and mutant ATP synthase confirm that residues of α-, β-, and γ-subunits are involved in the formation of phytochemical binding site. The variable degree of growth in the presence of DGPs also indicates the possibility of molecular targets other than ATP synthase. Our results establish that antibacterial properties of DGPs can be linked to the binding and inhibition of bacterial ATP synthase. Therefore, bacterial ATP synthase is a valuable molecular target for DGPs.

Induced protein degradation accomplishes elimination, rather than inhibition, of pathological proteins. Key to the success of this novel therapeutic modality is the modification of proteins with ubiquitin chains, which is brought about by molecular glues or bivalent compounds that induce proximity between the target protein and an E3 ligase. The human genome encodes ∼600 E3 ligases that differ widely in their structures, catalytic mechanisms, modes of regulation, and physiological roles. While many of these enzymes hold great promise for drug discovery, few have been successfully engaged by small-molecule degraders. Here, we review E3 ligases that are being used for induced protein degradation. Based on these prior successes and our growing understanding of the biology and biochemistry of E3 ligases, we propose new ubiquitylation enzymes that can be harnessed for drug discovery to firmly establish induced protein degradation as a specific and efficient therapeutic approach.

Since the EU approval of nitisinone in 2005, prognosis for patients with hereditary tyrosinaemia type 1 has changed dramatically, with patients living with the disease now reaching adulthood for the first time in history. This study aimed to assess the long-term safety and outcomes of nitisinone treatment in patients with hereditary tyrosinaemia type 1.We did a non-interventional, non-comparative, multicentre study in 77 sites across 17 countries in Europe and collected retrospective and prospective longitudinal data in patients with hereditary tyrosinaemia type 1 who were treated with oral nitisinone during the study period (Feb 21, 2005, to Sept 30, 2019). There were no specific exclusion criteria. Patients were followed-up with an investigator at least annually for as long as they were treated, or until the end of the study. The primary endpoints, occurrence of adverse events related to hepatic, renal, ophthalmic, haematological, or cognitive or developmental function, were assessed in the complete set (all patients already receiving treatment at the index date [Feb 21, 2005] or starting treatment thereafter) and the index set (the subset of patients who had their first dose on the index date or later only).315 patients were enrolled during the study period (complete set). Additionally, data from 24 patients who had liver transplantation or died during the post-marketing surveillance programme were retrieved (extended analysis set; 339 patients). Median treatment duration was 11·2 years (range 0·7-28·4); cumulative nitisinone exposure was 3172·7 patient-years. Patients who were diagnosed by neonatal screening started nitisinone treatment at median age 0·8 months versus 8·5 months in those who presented clinically. Incidences of hepatic, renal, ophthalmic, haematological, or cognitive or developmental adverse events were low. Occurrence of liver transplantation or death was more frequent the later that treatment was initiated (none of 70 patients who started treatment at age28 days vs 35 [13%] of 268 patients who started treatment at age ≥28 days). 279 (89%) of 315 patients were assessed as having either very good or good nitisinone treatment compliance. Treatment and diet compliance declined as patients aged. Suboptimal plasma phenylalanine and tyrosine levels were observed. The majority of patients were reported to have good overall clinical condition throughout treatment; 176 (87%) of 203 during the entire study, 98% following 1 year of treatment.Long-term nitisinone treatment was well tolerated and no new safety signals were revealed. Life-limiting hepatic disease appears to have been prevented by early treatment start. Neonatal screening was the most effective way of ensuring early treatment. Standardised monitoring of blood tyrosine, phenylalanine, and nitisinone levels has potential to guide individualised therapy.Swedish Orphan Biovitrum (Sobi).

Urease is potential target for various human's health complications, such as peptic ulcer, gastric cancer and kidney stone formation. The present study was based on synthesis of new hybrid pharmacophore N-substituted hydrazine-carbothioamides as potential urease inhibitors. Presented method gave excellent yield in range of 85–95% for hydrazine-carbothioamides derivatives (3a-s) after reaction of mono- and disubstituted hydrazides (1a-k) and substituted isothiocyanates (2a-d). All newly derivatives were characterized by advanced spectroscopic techniques (FTIR, 1HNMR, 13CNMR, EMS) and were assessed for their urease inhibition potential. All analogs except for 3k, 3l and 3m demonstrated strong inhibitory potential for urease with IC50 values of 8.45 ± 0.14 to 25.72 ± 0.23 μM as compared to standard thiourea (IC50 21.26 ± 0.35 μM). The structure-activity relationship and mode of interaction was established by molecular docking studies. It was revealed that the N-substituted hydrazine-carbothioamides interacted with nickel atoms present in the active site of urease and supported the correlations with the experimental findings. Therefore, the afforded hydrazine-carbothioamides derivatives are interesting hits for urease inhibition studies with future prospects of modification and optimization.

A large alkaptonuria (AKU) cohort was studied to better characterise the poorly understood phenotype of aortic stenosis of rare disease AKU.Eighty-one patients attended the National Alkaptonuria Centre (NAC) between 2007 and 2020. Nine only attended once. Fifty-one attended more than once and received nitisinone 2 mg daily. Twenty-one attended at least twice without receiving nitisinone. Assessments included questionnaire analysis, standard transthoracic echocardiography, as well as photographs of ochronotic pigment in eyes and ears at baseline when 2 mg nitisinone was commenced, and yearly thereafter. Blood and urine samples were collected for chemical measurement. The prevalence of aortic stenosis and aortic valve replacement were 22.2 and 6.2% in the current group. Aortic maximum velocity (Vmax) was directly related to varying degrees to age (R = 0.58, p 0.001), systolic blood pressure (R = 0.32, p 0.05), serum homogentisic acid (sHGA) (R = 0.28, p 0.05), ochronosis scores (R = 0.72, p 0.001), and alkaptonuria severity score index (AKUSSI) (R = 0.58, p 0.001) on linear regression analysis. Age and ochronosis scores were significantly related to Vmax on multiple regression analysis (p 0.005). Nitisinone decreased sHGA, 24-h urine HGA (uHGAAortic valve disease is highly prevalent in this NAC cohort, and strongly associated with ochronosis and disease severity. Nitisinone decreases ochronosis and had a similar significant effect on Vmax.

Background To investigate the efficacy of a novel experimental model for exploring visual function using a contrast-optomotor response (C-OMR) assay made by applying the contrast sensitivity test to the OMR assay in zebrafish. Methods Zebrafish larvae were treated with 0 (control), 5, 10, or 15 μM gentamicin and digoxin for 24 h at four days post-fertilization (dpf). Zebrafish larvae were assessed using the C-OMR assay with graded contrast gray-white stripes at 5 dpf, and the results were expressed as the percentage of larvae that finished swimming for 30 s (n = 20 per each group). The same C-OMR assay was repeated four times using different larvae. Results The percentage of larvae that finished swimming within 30 s was significantly reduced in larvae treated with 5, 10, and 15 μM gentamicin and 10 and 15 μM digoxin as compared to the Control groups. The C-OMR assay could distinguish that the decrease in visual function was different depending on the concentration of gentamicin and digoxin (5, 10, and 15 μM), whereas the OMR test with one contrast gray-white stripe could not. Conclusions The method of analyzing zebrafish OMR using graded contrast gray-white stripes is more sensitive than the OMR assay alone and may be more useful for assessing the drug toxicity and eye-related diseases to improve the understanding of drug-induced ocular side effects in the clinic.

Long-term potentiation (LTP) and long-term depression (LTD) are key forms of synaptic plasticity in the hippocampus. LTP and LTD are believed to underlie the processes occurring during learning and memory. Search of mechanisms responsible for switching from LTP to LTD and vice versa is an important fundamental task. Protein synthesis blockers (PSB) are widely used in models of memory impairment and LTP suppression. Here, we found that blockade of serine/threonine phosphatases 1 (PP1) and 2A (PP2A) with the specific blockers, calyculin A (CalyA) or okadaic acid (OA), and simultaneous blockade of the protein translation by anisomycin or cycloheximide leads to a switch from PSB-impaired LTP to LTD. PP1/PP2A-dependent LTD was extremely sensitive to the intensity of the test stimuli, whose increase restored the field excitatory postsynaptic potentials (fEPSP) to the values corresponding to control LTP in the non-treated slices. PP1/PP2A blockade affected the basal synaptic transmission, increasing the paired-pulse facilitation (PPF) ratio, and restored the PSB-impaired PPF 3 h after tetanus. Prolonged exposure to anisomycin led to the NO synthesis increase (measured using fluorescent dye) both in the dendrites and somata of CA1, CA3, dentate gyrus (DG) hippocampal layers. OA partially prevented the NO production in the CA1 dendrites, as well in the CA3 and DG somas. Direct measurements of changes in serine/threonine phosphatase (STPP) activity revealed importance of the PP1/PP2A-dependent component in the late LTP phase (L-LTP) in anisomycin-treated slices. Thus, serine/threonine phosphatases PP1/PP2A influence both basal synaptic transmission and stimulation-induced synaptic plasticity.

The mitogen-activated protein kinase (MAPK) pathway plays an important role in the colorectal cancer (CRC) progression, being supposed to be activated by the gene mutations, such as BRAF or KRAS. Although the inhibitors of extracellular signal-regulated kinase (ERK) have demonstrated efficacy in the cells with the BRAF or KRAS mutations, a clinical response is not always associated with the molecular signature. The patient-derived organoids (PDO) have emerged as a powerful in vitro model system to study cancer, and it has been widely applied for the drug screening. The present study aims to analyze the association between the molecular characteristics which analyzed by next-generation sequencing (NGS) and sensitivity to the ERK inhibitor (i.e., SCH772984) in PDO derived from CRC specimens. A drug sensitivity test for the SCH772984 was conducted using 14 CRC cell lines, and the results demonstrated that the sensitivity was in agreement with the BRAF mutation, but was not completely consistent with the KRAS status. In the drug sensitivity test for PDO, 6 out of 7 cases with either BRAF or KRAS mutations showed sensitivity to the SCH772984, while 5 out of 6 cases of both BRAF and KRAS wild-types were resistant. The results of this study suggested that the molecular status of the clinical specimens are likely to represent the sensitivity in the PDOs but is not necessarily absolutely overlapping. PDO might be able to complement the limitations of the gene panel and have the potential to provide a novel precision medicine.

A recent paper published in Nature Medicine by Calle et al. reported anti-nonalcoholic steatohepatitis (NASH) efficiencies by acetyl-CoA carboxylase (ACC) 1/2 inhibitors alone or by co-administration with a ACC1/2 inhibitor and a diacylglycerol acyltransferase 2 (DGAT2) inhibitor. Whereas the monotherapy achieved remarkable reductions in liver steatosis but induced hyperlipidemia, DGAT2 inhibitor co-administration mitigated the increase in serum triglycerides (TGs).

Fully understanding the regulatory network under the pluripotency of embryonic stem cells (ESC) is a prerequisite for their safe application. Here, we addressed the characteristics of metastasis-associated (MTA) family members in human ESCs and found that knockdown of the expression of MTA2 and MTA3, but not MTA1, would induce differentiation. High-throughput sequence and quantitative real-time PCR showed that the decreased MTA2 or MTA3 gene transcript mainly led to the emergence of mesendoderm associated markers. Finally, based on the chemical small molecule library screening, we observed that addition of ID8, a specific inhibitor of the dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs), was able to impair the differentiation phenotype induced by MTA2 and MTA3 reduction. Functional assay showed that ID8 could mediate differentiation caused by MTA2 or MTA3 knockdown mainly through inhibition of DYRK4 activity. Therefore, our finding provides the evidence that the functions of MTA family genes in human ESCs are different. Revealing the function of MTA in ESCs with different pluripotency states will help us better understand and apply stem cells.

Alpha-1 acid glycoprotein (AGP) is a major acute-phase protein that is involved in drug/ligand binding and regulation of immune response. In response to inflammation, AGP secretion from the liver increases, resulting in elevated concentration of plasma AGP. AGP exhibits multiple N-glycosylation sites, and thus, is highly glycosylated. Although AGP glycosylation is considered to affect its functions, the significance of AGP glycosylation for its secretion is unclear. In this study, we investigated the effects of AGP glycosylation using glycosylation-deficient mouse AGP mutants lacking one, four, or all five N-glycosylation sites. Furthermore, we examined the effects of endoplasmic reticulum (ER) stress-inducing reagents, including tunicamycin and thapsigargin, which induce ER stress in an N-glycosylation-dependent and -independent manner, respectively. Here, we found that glycosylation deficiency and ER stress induce a little or no effect on AGP secretion. Conversely, thapsigargin significantly suppressed AGP secretion in glycosylation-independent manner. These findings indicate that AGP secretion is regulated via thapsigargin-sensitive pathway that might be further controlled by the intracellular calcium concentrations.

Novel therapeutic strategies for Alzheimer's disease (AD) are of the greatest priority given the consistent failure of recent clinical trials focused on Aβ or pTau. Earlier, we demonstrated that mild mitochondrial complex I inhibitor CP2 blocks neurodegeneration and cognitive decline in multiple mouse models of AD. To evaluate the safety of CP2 in humans, we performed a genome-wide association study (GWAS) using 196 lymphoblastoid cell lines and identified 11 SNP loci and 64 mRNA expression probe sets that potentially associate with CP2 susceptibility. Using primary mouse neurons and pharmacokinetic study, we show that CP2 is generally safe at a therapeutic dose.

The indispensable role of Beta-site amyloid precursor protein cleaving enzyme-1 (BACE1) in Amyloid beta (Aβ) plaques generation and Aβ-mediated synaptic dysfunctions makes it a crucial target for therapeutic intervention in Alzheimer's disease (AD). In order to find out the potential inhibitors of BACE1, the present study focused on five phytochemicals from Pueraria tuberosa, namely, daidzin, genistin, mangiferin, puerarin, and tuberosin. A molecular docking study showed that all five phytochemicals presented the strongest BACE1 inhibition. Integrated molecular dynamics simulations and free energy calculations demonstrated that all five natural compounds have stable and favorable energies causing strong binding with the pocket site of BACE1 on 50 ns. All these molecules also passed Lipinski's rule of five. To validate the molecular modeling based findings, we primarily targeted the cognitive decline associated with BACE1 expression in AD flies with P. tuberosa. Significant improvement in cognitive decline was observed in AD flies in different behavioral assays such as Larval crawling assay (16.38%), Larval light preference assay (26.39%), Climbing assay (32.97%), Cold sensitivity assay (43.6%), and Thermal sensitivity assay (44.42%). The present findings suggest that P. tuberosa may be considered as a promising dietary supplement that can significantly ameliorate cognitive decline caused by BACE1 in Alzheimer's disease (AD).

The burden of obesity is increasing all over the world. Except for Orlistat, no effective anti-obesity drug is currently available. Therefore, a search for the new anti-obesity compound is need of time. This study demonstrates macromolecular interaction and inhibitory effect of pentacyclic triterpenoids (PTT) on pancreatic lipase (PL). In the present study PTTs from endophytic Colletotrichum gigasporum were found to show significant inhibitory activity against PL with IC50 of 16.62 ± 1.43 μg/mL. The PTT isolated through bioassay-guided isolation showed a dose-dependent (R2 = 0.915) inhibition against porcine PL and the results were comparable with the standard (Orlistat). Based on inhibition kinetic data, the gradual increase in Km (app) with increasing PTT concentration indicated that the mode of interaction of PTT with PL was a competitive type, and it directly competed with the substrate (pNPB) for the active site of PL. In vivo studies in Wistar rats at the oral dose (100 mg/kg body weight) of PTT significantly decreased (p