Biocatalytic strategy for the construction of sp3-rich polycyclic compounds from directed evolution and computational modelling.

Catalysis with engineered enzymes has provided more efficient routes for the production of active pharmaceutical agents. However, the potential of biocatalysis to assist in early-stage drug discovery campaigns remains largely untapped. In this study, we have developed a biocatalytic strategy for the construction of sp3-rich polycyclic compounds via the intramolecular cyclopropanation of benzothiophenes and related heterocycles. Two carbene transferases with complementary regioisomer selectivity were evolved to catalyse the stereoselective cyclization of benzothiophene substrates bearing diazo ester groups at the C2 or C3 position of the heterocycle. The detailed mechanisms of these reactions were elucidated by a combination of crystallographic and computational analyses. Leveraging these insights, the substrate scope of one of the biocatalysts could be expanded to include previously unreactive substrates, highlighting the value of integrating evolutionary and rational strategies to develop enzymes for new-to-nature transformations. The molecular scaffolds accessed here feature a combination of three-dimensional and stereochemical complexity with 'rule-of-three' properties, which should make them highly valuable for fragment-based drug discovery campaigns.

Dibenzothiophene removal by environmental bacteria with differential accumulation of intracellular inorganic polyphosphate.

Dibenzothiophene (DBT), which belongs to the group of polycyclic aromatic heterocycles of sulfur, is a model substance to study the removal of sulfur compounds from oil due to its recalcitrance to traditional and specific removal processes. The aim of this work was to evaluate DBT bioremoval by environmental bacteria and its relationship with polyphosphate (polyP) accumulation, cell surface characteristics and bioemulsifying activity. Pseudomonas sp. P26 achieved the highest DBT removal percentage (48%) after 7 days of incubation. Moreover, positive correlations were estimated between DBT removal and bioemulsifying activity and biofilm formation. A strain-dependent relationship between the content of intracellular polyP and the presence of DBT in the culture medium was also demonstrated. The study of these bacterial characteristics, which could promote DBT transformation, is a first approach to select DBT-removing bacteria, in order to develop bioformulations that are able to contribute to desulfurization processes of petroleum-derived pollutants in the future.

Daily rhythms of REV-ERBα and its role as transcriptional repressor of clock genes in fish hepatic oscillator.

The REV-ERBα nuclear receptor is a key component of the molecular machinery of circadian oscillators in mammals. While the rhythmic expression of this receptor has been described in teleosts, several critical aspects of its regulation remain unknown, such as which synchronizers entrain its rhythm, and whether it can modulate the expression of other clock genes. The objective of this study was to gain deeper understanding of the role of REV-ERBα in the fish circadian system. To this end, we first investigated the cues that entrain the rhythm of rev-erbα expression in the goldfish (Carassius auratus) liver and hypothalamus. A 12-h shift in feeding time induced a parallel shift in the hepatic rhythm of rev-erbα expression, confirming that this gene is food-entrainable in the goldfish liver. In contrast, light seems the main driver of rev-erbα rhythmic expression in the hypothalamus. Next, we examined the effects of REV-ERBα activation on locomotor activity and hepatic expression of clock genes. Subchronic treatment with the REV-ERBα agonist SR9009 slightly decreased locomotor activity anticipating light onset and food arrival, and downregulated hepatic bmal1a, clock1a, cry1a, per1a and pparα expression. This generalized repressing action of REV-ERBα on the expression of hepatic clock genes was confirmed in vitro by using agonists (SR9009 and GSK4112) and antagonist (SR8278) of this receptor. Overall, the present work reveals that REV-ERBα modulates the daily expression of the main genes of the teleostean liver clock, reinforcing its role in the liver temporal homeostasis, which seems highly conserved in both fish and mammals.

Bio-catalytic degradation of dibenzothiophene (DBT) in petroleum distillate (diesel) by Pseudomonas spp.

Biodesulfurization (BDS) was employed in this study to degrade dibenzothiophene (DBT) which accounts for 70% of the sulfur compounds in diesel using a synthetic and typical South African diesel in the aqueous and biphasic medium. Two Pseudomonas sp. bacteria namely Pseudomonas aeruginosa and Pseudomonas putida were used as biocatalysts. The desulfurization pathways of DBT by the two bacteria were determined by gas chromatography (GC)/mass spectrometry (MS) and High-Performance Liquid Chromatography (HPLC). Both organisms were found to produce 2-hydroxy biphenyl, the desulfurized product of DBT. Results showed BDS performance of 67.53% and 50.02%, by Pseudomonas aeruginosa and Pseudomonas putida, respectively for 500 ppm initial DBT concentration. In order to study the desulfurization of diesel oils obtained from an oil refinery, resting cells studies by Pseudomonas aeruginosa were carried out which showed a decrease of about 30% and 70.54% DBT removal for 5200 ppm in hydrodesulfurization (HDS) feed diesel and 120 ppm in HDS outlet diesel, respectively. Pseudomonas aeruginosa and Pseudomonas putida selectively degraded DBT to form 2-HBP. Application of these bacteria for the desulfurization of diesel showed promising potential for decreasing the sulfur content of South African diesel oil.

Benzo[b]naphtho[d]thiophenes and naphthylbenzo[b]thiophenes: Their aryl hydrocarbon receptor-mediated activities and environmental presence.

Polycyclic aromatic sulfur heterocyclic compounds (PASHs) belong among ubiquitous environmental pollutants; however, their toxic effects remain poorly understood. Here, we studied the aryl hydrocarbon receptor (AhR)-mediated activity of dibenzothiophene, benzo[b]naphtho[d]thiophenes, and naphthylbenzo[b]thiophenes, as well as their presence in two types of environmental matrices: river sediments collected from both rural and urban areas, and in airborne particulate matter (PM2.5) sampled in cities with different levels and sources of pollution. Benzo[b]naphtho[2,1-d]thiophene, benzo[b]naphtho[2,3-d]thiophene, 2,2-naphthylbenzo[b]thiophene, and 2,1-naphthylbenzo[b]thiophene were newly identified as efficient AhR agonists in both rat and human AhR-based reporter gene assays, with 2,2-naphthylbenzo[b]thiophene being the most potent compound identified in both species. Benzo[b]naphtho[1,2-d]thiophene and 3,2-naphthylbenzo[b]thiophene elicited AhR-mediated activity only in the rat liver cell model, while dibenzothiophene and 3,1-naphthylbenzo[b]thiophene were inactive in either cell type. Independently of their ability to activate the AhR, benzo[b]naphtho[1,2-d]thiophene, 2,1-naphthylbenzo[b]thiophene, 3,1-naphthylbenzo[b]thiophene, and 3,2-naphthylbenzo[b]thiophene inhibited gap junctional intercellular communication in a model of rat liver epithelial cells. Benzo[b]naphtho[d]thiophenes were dominant PASHs present in both PM2.5 and sediment samples, with benzo[b]naphtho[2,1-d]thiophene being the most abundant one, followed by benzo[b]naphtho[2,3-d]thiophene. The levels of naphthylbenzo[b]thiophenes were mostly low or below detection limit. Benzo[b]naphtho[2,1-d]thiophene and benzo[b]naphtho[2,3-d]thiophene were identified as the most significant contributors to the AhR-mediated activity in the environmental samples evaluated in this study. Both induced nuclear translocation of the AhR, and they induced CYP1A1 expression in a time-dependent manner, suggesting that their AhR-mediated activity may depend on the rate of their intracellular metabolism. In conclusion, some PASHs could be significant contributors to the overall AhR-mediated toxicity of complex environmental samples suggesting that more attention should be paid to the potential health impacts of this group of environmental pollutants.

Assessment of the dibenzothiophene desulfurization potential of indigenously isolated bacterial consortium IQMJ-5: a different approach to safeguard the environment.

Biodesulfurization is emerging as a valuable technology for the desulfurization of dibenzothiophene (DBT) and its alkylated substitutes, which are otherwise regarded as refractory to other physical and chemical desulfurizing techniques. The inability of the currently identified pure cultures and artificial microbial consortia due to lower desulfurization rate and product inhibition issues has compelled the researcher to look for an alternative solution. Thus, in the present study, an indigenously isolated microbial consortium was employed to tackle the desulfurization issue. Herein, we isolated several kinds of DBT desulfurizing natural microbial consortia from hydrocarbon-contaminated soil samples by conventional enrichment technique. The most effective desulfurizing microbial consortium was sequenced through illumine sequencing technique. Finally, the effect of the products of the desulfurizing pathway (such as 2-hydroxybiphenyl (2-HBP) and sulfate (SO4-2) was evaluated on the growth and desulfurization capability of the isolated consortium. The outcomes of Gibb's assay analysis showed that six isolates followed the "4S" pathway and converted DBT to 2-HBP. Among the isolates, I5 showed maximum growth rate (1.078 g/L dry cell weight) and desulfurization activity (about 77% as indicated by HPLC analysis) and was considered for further in-depth experimentation. The analysis of 16S rRNA by high-throughput sequencing approach of the I5 isolate revealed five types of bacterial phyla including Proteobacteria, Bacteroidetes, Firmicutes, Patescibacteria, and Actinobacteria (in order of abundance). The isolate showed significant tolerance to the inhibitory effect of both 2-HBP and SO4-2 and maintained growth in the presence of even about 1.0 mM initial concentration of both products. This clearly suggests that the isolate can be an efficient candidate for future in-depth desulfurization studies of coal and other fossil fuels.

Evybactin is a DNA gyrase inhibitor that selectively kills Mycobacterium tuberculosis.

The antimicrobial resistance crisis requires the introduction of novel antibiotics. The use of conventional broad-spectrum compounds selects for resistance in off-target pathogens and harms the microbiome. This is especially true for Mycobacterium tuberculosis, where treatment requires a 6-month course of antibiotics. Here we show that a novel antimicrobial from Photorhabdus noenieputensis, which we named evybactin, is a potent and selective antibiotic acting against M. tuberculosis. Evybactin targets DNA gyrase and binds to a site overlapping with synthetic thiophene poisons. Given the conserved nature of DNA gyrase, the observed selectivity against M. tuberculosis is puzzling. We found that evybactin is smuggled into the cell by a promiscuous transporter of hydrophilic compounds, BacA. Evybactin is the first, but likely not the only, antimicrobial compound found to employ this unusual mechanism of selectivity.

Benzo[b]naphtho[1,2-d]thiophene Sulfoxides: Biomimetic Synthesis, Photophysical Properties, and Applications.

A practical synthesis of nonsymmetrical thiophene-fused aromatic systems has been developed that was inspired by the biodegradation of benzothiophene. For the first time, the photophysical properties of a series of π-conjugated benzo[b]naphtho[1,2-d]thiophene (BNT) sulfoxides were explored both in solution and in the solid state. The excellent fluorescence characteristics enable various applications of these compounds.

Hydroxyl containing benzo[b]thiophene analogs mitigates the acrylamide induced oxidative stress in the zebrafish larvae by stabilizing the glutathione redox cycle.

AIMS: This study aims to elucidate a systematic free-radical quenching ability of synthesized benzo[b]thiophene derivatives using in vitro assays and acrylamide induced oxidatively stressed model in zebrafish larvae. MATERIALS AND METHODS: Antioxidant activity of the compounds was evaluated using in vitro methods. The toxicity of the compounds was evaluated in Madin-Darby Canine Kidney (MDCK) cell line and zebrafish embryos. Oxidative stress was generated by acrylamide (1 mM) in zebrafish larvae and treated with compounds to evaluate the in vivo antioxidant ability. Specific fluorescence dyes were used to detect ROS generation, lipid peroxidation, and cell death followed by gene expression using RT PCR. Density functional theory (DFT) and in silico pharmacokinetics were also studied. KEY FINDINGS: Compound BP and EP have a greater in vitro free radical scavenging ability. The maximum tolerated concentration (MTC) of the compounds in zebrafish larvae is 80 µM. The antioxidant system in zebrafish larvae was dysregulated due to acrylamide exposure and improvement was found while treating acrylamide exposed larvae with compounds 1-(3-hydroxybenzo[b]thiophen-2-yl) ethanone (BP) and 1-(3-hydroxybenzo[b]thiophen-2-yl) propan-1-one hydrate (EP). Compound BP and EP enhanced the SOD and CAT activity, reduced the ROS and lipid peroxidation level, thus decreasing cell death in zebrafish larvae. Compound BP and EP also improved the glutathione redox cycle by stabilizing glutathione-related gene expressions. SIGNIFICANCE: Hydroxyl-containing compounds BP and EP are promising lead molecules for pathological conditions related to oxidative stress, which showed an attenuated effect on acrylamide-induced oxidative stress in zebrafish larvae by enhancing the glutathione redox cycle and enzymatic antioxidants.

Characterization of a dszEABC operon providing fast growth on dibenzothiophene and construction of broad-host-range biodesulfurization catalysts.

A new operon for biodesulfurization (BDS) of dibenzothiophene and derivatives has been isolated from a metagenomic library made from oil-contaminated soil, by selecting growth of E. coli on DBT as the sulfur source. This operon is similar to a dszEABC operon also isolated by metagenomic functional screening but exhibited substantial differences: (i) the new fosmid provides much faster growth on DBT; (ii) associated dszEABC genes can be expressed without the need of heterologous expression from the vector promoter; and (iii) monooxygenases encoded in the fosmid cannot oxidize indole to produce indigo. We show how expression of the new dszEABC operon is regulated by the sulfur source, being induced under sulfur-limiting conditions. Its transcription is activated by DszR, a type IV activator οf σN -dependent promoters. DszR is coded in a dszHR operon, whose transcription is in turn regulated by sulfur and presumably activated by the global regulator of sulfur metabolism CysB. Expression of dszH is essential for production of active DszR, although it is not involved in sulfur sensing or regulation. Two broad-host-range DBT biodesulfurization catalysts have been constructed and shown to provide DBT biodesulfurization capability to three Pseudomonas strains, displaying desirable characteristics for biocatalysts to be used in BDS processes.

Impacts of Sodium/Glucose Cotransporter-2 Inhibitors on Circulating Uric Acid Concentrations: A Systematic Review and Meta-Analysis.

BACKGROUND: Several trials have assessed the antihyperglycemic effects of sodium/glucose cotransporter-2 inhibitors (SGLT2i) in patients with type 2 diabetes mellitus (T2DM). We conducted a quantitative analysis to assess the impact of SGLT2is on serum uric acid (SUA) in patients with T2DM. METHODS: Placebo-controlled trials published before 13 August 2021 were identified by searching PubMed, Embase, Web of Science, and Scopus. The intervention group received SGLT2i as monotherapy or add-on treatment, and the control group received a placebo that was replaced with SGLT2i. Clinical trials providing changes in SUA were included. The mean change of SUA, glycated hemoglobin (HbA1c), fasting plasma glucose (FPG), and body weight were calculated (PROSPERO CRD42021287019). RESULTS: After screening of 1172 papers, 59 papers were included in the systematic review. A total of 55 trials (122 groups) of 7 types of SGLT2i on patients with T2DM were eligible for meta-analysis. All SGLT2is significantly decreased SUA levels compared with the placebo groups: empagliflozin mean difference (MD) = -40.98 µmol/L, 95% CI [-47.63, -34.32], dapagliflozin MD = -35.17 µmol/L, 95% CI [-39.68, -30.66], canagliflozin MD = -36.27 µmol/L, 95% CI [-41.62, -30.93], luseogliflozin MD = -24.269 µmol/L, 95% CI [-33.31, -15.22], tofogliflozin MD = -19.47 µmol/L, 95% CI [-27.40, -11.55], and ipragliflozin MD = -18.85 µmol/L, 95% CI [-27.20, -10.49]. SGLT2i also decreased FPG, body weight, and HbA1c levels. SUA reduction persisted during long-term treatment with SGLT2i (except for empagliflozin), while the SUA reduction was affected by the duration of diabetes. CONCLUSIONS: SGLT2i can be a valid therapeutic strategy for patients with T2DM and comorbid hyperuricemia. Besides reducing FPG, body weight, and HbA1c, SGLT2i can significantly decrease SUA levels compared to placebo (Total MD = -34.07 µmol/L, 95% CI [-37.00, -31.14]).

Synthesis and evaluation of RNase L-binding 2-aminothiophenes as anticancer agents.

Aminothiophene is a scaffold that is widely present in drugs and biologically active small molecules as chemical probes. In this study, 43 compounds sharing a 2-aminothiophenone-3-carboxylate (ATPC) scaffold, known to activate the ribonuclease L (RNase L), were synthesized and selected ATPCs showed enhancement of thermal stability of RNase L upon binding. Screening of antiproliferation activities against human cancer cell lines revealed that ATPCs represented by compounds 4l and 50 showed potent single-digit micromolar antiproliferation activity against human cancer cell lines. Compounds 4l and 50 exhibited time- and dose-dependent proliferation inhibition, induced cellular apoptosis measured by cleaved PARP and via flow cytometry, inhibited cell migration, and inhibited cell colony formation. Combining the results reported in this work, ATPCs were evaluated as potential anticancer agents mediated by RNase L-binding and apoptosis induction. The work contributes to the study on the polypharmacological properties of aminothiophene-containing small molecules.

Monooxygenase- and Dioxygenase-Catalyzed Oxidative Dearomatization of Thiophenes by Sulfoxidation, cis-Dihydroxylation and Epoxidation.

Enzymatic oxidations of thiophenes, including thiophene-containing drugs, are important for biodesulfurization of crude oil and drug metabolism of mono- and poly-cyclic thiophenes. Thiophene oxidative dearomatization pathways involve reactive metabolites, whose detection is important in the pharmaceutical industry, and are catalyzed by monooxygenase (sulfoxidation, epoxidation) and dioxygenase (sulfoxidation, dihydroxylation) enzymes. Sulfoxide and epoxide metabolites of thiophene substrates are often unstable, and, while cis-dihydrodiol metabolites are more stable, significant challenges are presented by both types of metabolite. Prediction of the structure, relative and absolute configuration, and enantiopurity of chiral metabolites obtained from thiophene enzymatic oxidation depends on the substrate, type of oxygenase selected, and molecular docking results. The racemization and dimerization of sulfoxides, cis/trans epimerization of dihydrodiol metabolites, and aromatization of epoxides are all factors associated with the mono- and di-oxygenase-catalyzed metabolism of thiophenes and thiophene-containing drugs and their applications in chemoenzymatic synthesis and medicine.

Species-dependent hepatic and intestinal metabolism of selective oestrogen receptor degrader LSZ102 by sulphation and glucuronidation.

LSZ102 is an orally bioavailable selective oestrogen receptor degrader in clinical development for the treatment of breast cancer. Preclinical studies showed efficacy in xenograft models on oral dosing. However, oral bioavailability was relatively low in several preclinical species (7-33%), and was associated with first-pass metabolism, particularly intestinal first-pass.To investigate metabolism and first-pass effects, metabolites were analysed in human plasma samples after oral dosing of LSZ102 to patients, rat plasma samples after oral dosing of [14C]LSZ102, and in vitro incubations of [14C]LSZ102 with human and rat hepatocytes and intestinal S9 fractions. The kinetics of human sulfotransferase (SULT) enzymes potentially involved in metabolism of LSZ102 was characterised.Sulphate metabolites were found to be the major components in human plasma, as well as in human hepatocytes and intestinal S9 fractions. Contrastingly, glucuronidation was predominant in rat plasma, hepatocytes and intestinal S9. LSZ102 was found to be metabolised by several human SULTs expressed in liver and intestine. The combined metabolism data in rat and human provide supporting evidence for an extensive intestinal first-pass metabolism effect via sulphation in human but glucuronidation in rat.As LSZ102 is metabolised by a number of different SULTs, drug-drug interactions resulting from the inhibition of one SULT are unlikely.Despite the observed species difference in metabolism, the major human metabolites of LSZ102, sulphate M5, glucuronide M4, and secondary glucuronide/sulphate metabolite M12, have no or weak pharmacological activity and are not considered a toxicity risk as they are phase II conjugative metabolites.

Synthesis and Evaluation of Structurally Diverse C-2-Substituted Thienopyrimidine-Based Inhibitors of the Human Geranylgeranyl Pyrophosphate Synthase.

Novel analogues of C-2-substituted thienopyrimidine-based bisphosphonates (C2-ThP-BPs) are described that are potent inhibitors of the human geranylgeranyl pyrophosphate synthase (hGGPPS). Members of this class of compounds induce target-selective apoptosis of multiple myeloma (MM) cells and exhibit antimyeloma activity in vivo. A key structural element of these inhibitors is a linker moiety that connects their (((2-phenylthieno[2,3-d]pyrimidin-4-yl)amino)methylene)bisphosphonic acid core to various side chains. The structural diversity of this linker moiety, as well as the side chains attached to it, was investigated and found to significantly impact the toxicity of these compounds in MM cells. The most potent inhibitor identified was evaluated in mouse and rat for liver toxicity and systemic exposure, respectively, providing further optimism for the potential value of such compounds as human therapeutics.

In vitro metabolism of the REV-ERB agonist SR-9009 and subsequent detection of metabolites in associated routine equine plasma and urine doping control samples.

SR-9009 is a synthetic compound widely available to purchase online as 'supplement' products due to its potential performance-enhancing effects, presenting a significant threat with regard to doping control in sport. In vitro metabolism with equine liver microsomes was performed to identify potential targets for detection of SR-9009. Six metabolites were identified, with the most abundant consisting of N-dealkylated metabolites (M1-M3). The addition of the identified metabolites to high-resolution accurate mass databases resulted in a positive finding for the N-dealkylated metabolite M1 of SR-9009 in an associated plasma and urine doping sample. Liquid chromatography-high-resolution mass spectrometry was used to verify the presence of the N-dealkylated metabolite (M1) in both matrices, with a low concentration of the parent compound and additional N-desalkyl metabolites (M2 and M3) detected in the plasma sample as supporting evidence of administration. To the best of the authors' knowledge, this is the first report of an adverse analytical finding in an equine sample for SR-9009 or its metabolites in equine doping control.

Metabolic drug survey highlights cancer cell dependencies and vulnerabilities.

Interrogation of cellular metabolism with high-throughput screening approaches can unravel contextual biology and identify cancer-specific metabolic vulnerabilities. To systematically study the consequences of distinct metabolic perturbations, we assemble a comprehensive metabolic drug library (CeMM Library of Metabolic Drugs; CLIMET) covering 243 compounds. We, next, characterize it phenotypically in a diverse panel of myeloid leukemia cell lines and primary patient cells. Analysis of the drug response profiles reveals that 77 drugs affect cell viability, with the top effective compounds targeting nucleic acid synthesis, oxidative stress, and the PI3K/mTOR pathway. Clustering of individual drug response profiles stratifies the cell lines into five functional groups, which link to specific molecular and metabolic features. Mechanistic characterization of selective responses to the PI3K inhibitor pictilisib, the fatty acid synthase inhibitor GSK2194069, and the SLC16A1 inhibitor AZD3965, bring forth biomarkers of drug response. Phenotypic screening using CLIMET represents a valuable tool to probe cellular metabolism and identify metabolic dependencies at large.

ROS activated prodrug for ALDH overexpressed cancer stem cells.

Aldehyde dehydrogenase (ALDH), a cancer stem cell biomarker, is related to drug resistance. Co-treatment of anti-cancer drug (CPT) and ALDH inhibitor (DEAB) can overcome the drug resistance of cancer stem cells (CSCs) and finally cure cancers without relapse. We herein introduce a prodrug (DE-CPT) - consisting of 1,3-oxathiolane as an ROS responsive scaffold, and an aldehyde protecting group of DEAB - to deliver the CPT and DEAB upon reaction with ROS. From tests of the sphere-forming ability and CSC marker subpopulation, we found that DE-CPT efficiently decreases the CSCs population and kills the cancer cells.

Effect of Erdosteine on Middle Ear Effusion in Rats by Mediating TLR4 Signaling Pathway.

The study aimed to investigate the effect of erdosteine on middle ear effusion in rats through mediating the Toll-like receptor 4 (TLR4) signaling pathway. Rats were injected with endotoxin to prepare the model of acute secretory otitis media (SOM). Then, they were divided into an acute SOM model group (model group, n = 15) and erdosteine treatment group (18 mg/kg, gavage, treatment group, n = 15). Besides, a normal group (n = 15) was set up. Two weeks later, routine biochemical indicators such as aspartate aminotransferase (AST) and alkaline phosphatase (ALP) were detected. The inflammatory effusion due to otitis media was scored. The content of myeloperoxidase (MPO), matrix metalloproteinase (MMP), and tumor necrosis factor-beta (TNF-ß) in middle ear lavage fluid was detected via enzyme-linked immunosorbent assay (ELISA). Additionally, histomorphological changes were observed with the help of hematoxylin-eosin (HE) staining, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blotting assays were carried out to measure the expression levels of TLR4 pathway genes and proteins as well as the messenger ribonucleic acid (mRNA) expression levels of key factors for otitis media (mucin 2 (MUC2) and MUC5A). In the model group, the levels of AST, ALP, and glutamic-pyruvic transaminase (GPT) were significantly increased (p < 0.05). Besides, the content of MPO, MMP, and TNF-ß was overtly raised in the model group (p < 0.05), while it was notably lowered in the treatment group (p < 0.05). In the treatment group, the cilia were slightly swollen, and inflammatory cells were fewer. The mRNA levels of MUC2, MUC5A, and pathway genes TLR4 and c-Jun N-terminal kinase (JNK) were elevated in the model group. In addition, the protein assay results revealed that the protein levels of TLR4 and JNK were evidently increased in the model group. Erdosteine can treat the middle ear effusion in rats by repressing the activation of the TLR4 signaling pathway.

Identification of a small molecule SR9009 that activates NRF2 to counteract cellular senescence.

The senescence-associated secretory phenotype (SASP) is a striking characteristic of senescence. Accumulation of SASP factors causes a pro-inflammatory response linked to chronic disease. Suppressing senescence and SASP represents a strategy to prevent or control senescence-associated diseases. Here, we identified a small molecule SR9009 as a potent SASP suppressor in therapy-induced senescence (TIS) and oncogene-induced senescence (OIS). The mechanism studies revealed that SR9009 inhibits the SASP and full DNA damage response (DDR) activation through the activation of the NRF2 pathway, thereby decreasing the ROS level by regulating the expression of antioxidant enzymes. We further identified that SR9009 effectively prevents cellular senescence and suppresses the SASP in the livers of both radiation-induced and oncogene-induced senescence mouse models, leading to alleviation of immune cell infiltration. Taken together, our findings suggested that SR9009 prevents cellular senescence via the NRF2 pathway in vitro and in vivo, and activation of NRF2 may be a novel therapeutic strategy for preventing cellular senescence.