Sequencing of N6-methyl-deoxyadenosine at single-base resolution across the mammalian genome.

Although DNA N6-methyl-deoxyadenosine (6mA) is abundant in bacteria and protists, its presence and function in mammalian genomes have been less clear. We present Direct-Read 6mA sequencing (DR-6mA-seq), an antibody-independent method, to measure 6mA at base resolution. DR-6mA-seq employs a unique mutation-based strategy to reveal 6mA sites as misincorporation signatures without any chemical or enzymatic modulation of 6mA. We validated DR-6mA-seq through the successful mapping of the well-characterized G(6mA)TC motif in the E. coli DNA. As expected, when applying DR-6mA-seq to mammalian systems, we found that genomic DNA (gDNA) 6mA abundance is generally low in most mammalian tissues and cells; however, we did observe distinct gDNA 6mA sites in mouse testis and glioblastoma cells. DR-6mA-seq provides an enabling tool to detect 6mA at single-base resolution for a comprehensive understanding of DNA 6mA in eukaryotes.

Trans-vaccenic acid reprograms CD8+ T cells and anti-tumour immunity.

Diet-derived nutrients are inextricably linked to human physiology by providing energy and biosynthetic building blocks and by functioning as regulatory molecules. However, the mechanisms by which circulating nutrients in the human body influence specific physiological processes remain largely unknown. Here we use a blood nutrient compound library-based screening approach to demonstrate that dietary trans-vaccenic acid (TVA) directly promotes effector CD8+ T cell function and anti-tumour immunity in vivo. TVA is the predominant form of trans-fatty acids enriched in human milk, but the human body cannot produce TVA endogenously1. Circulating TVA in humans is mainly from ruminant-derived foods including beef, lamb and dairy products such as milk and butter2,3, but only around 19% or 12% of dietary TVA is converted to rumenic acid by humans or mice, respectively4,5. Mechanistically, TVA inactivates the cell-surface receptor GPR43, an immunomodulatory G protein-coupled receptor activated by its short-chain fatty acid ligands6-8. TVA thus antagonizes the short-chain fatty acid agonists of GPR43, leading to activation of the cAMP-PKA-CREB axis for enhanced CD8+ T cell function. These findings reveal that diet-derived TVA represents a mechanism for host-extrinsic reprogramming of CD8+ T cells as opposed to the intrahost gut microbiota-derived short-chain fatty acids. TVA thus has translational potential for the treatment of tumours.

YTHDF2/m6 A/NF-κB axis controls anti-tumor immunity by regulating intratumoral Tregs.

N6 -methyladenosine (m6 A) in messenger RNA (mRNA) regulates immune cells in homeostasis and in response to infection and inflammation. The function of the m6 A reader YTHDF2 in the tumor microenvironment (TME) in these contexts has not been explored. We discovered that the loss of YTHDF2 in regulatory T (Treg) cells reduces tumor growth in mice. Deletion of Ythdf2 in Tregs does not affect peripheral immune homeostasis but leads to increased apoptosis and impaired suppressive function of Treg cells in the TME. Elevated tumor necrosis factor (TNF) signaling in the TME promotes YTHDF2 expression, which in turn regulates NF-κB signaling by accelerating the degradation of m6 A-modified transcripts that encode NF-κB-negative regulators. This TME-specific regulation of Treg by YTHDF2 points to YTHDF2 as a potential target for anti-cancer immunotherapy, where intratumoral Treg cells can be targeted to enhance anti-tumor immune response while avoiding Treg cells in the periphery to minimize undesired inflammations.

Well-Paired-Seq2: High-Throughput and High-Sensitivity Strategy for Characterizing Low RNA-Content Cell/Nucleus Transcriptomes.

Single-cell RNA sequencing (scRNA-seq) is a transformative technology that unravels the intricate cellular state heterogeneity. However, the Poisson-dependent cell capture and low sensitivity in scRNA-seq methods pose challenges for throughput and samples with a low RNA-content. Herein, to address these challenges, we present Well-Paired-Seq2 (WPS2), harnessing size-exclusion and quasi-static hydrodynamics for efficient cell capture. WPS2 exploits molecular crowding effect, tailing activity enhancement in reverse transcription, and homogeneous enzymatic reaction in the initial bead-based amplification to achieve 3116 genes and 8447 transcripts with an average of ∼20000 reads per cell. WPS2 detected 1420 more genes and 4864 more transcripts than our previous Well-Paired-Seq. It sensitively characterizes transcriptomes of low RNA-content single cells and nuclei, overcoming the Poisson limit for cell and barcoded bead capture. WPS2 also profiles transcriptomes from frozen clinical samples, revealing heterogeneous tumor copy number variations and intercellular crosstalk in clear cell renal cell carcinomas. Additionally, we provide the first single-cell-level characterization of rare metanephric adenoma (MA) and uncover potential specific markers. With the advantages of high sensitivity and high throughput, WPS2 holds promise for diverse basic and clinical research.

Integrated spatial metabolomics and network pharmacology to explore the pharmacodynamic substances and mechanism of Radix ginseng-Schisandra chinensis Herb Couple on Alzheimer's disease.

Radix ginseng and Schisandra chinensis have been extensively documented in traditional Chinese medicine (TCM) for their potential efficacy in treating dementia. However, the precise mechanism of their therapeutic effects remains to be fully elucidated. In this study, air flow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI) and network pharmacology are used to investigate the pharmacodynamics and mechanism underlying the herbal combination consisting of Radix ginseng-Schisandra chinensis (RS) in a rodent model for Alzheimer's disease (AD). Brain histopathological findings suggested that RS attenuates hippocampal damage in AD mice, making this combination a potential AD treatment. Twenty-eight biomarkers were identified by spatial metabolomics analysis, which are intricately linked to neuroinflammation, neurotransmitter imbalance, energy deficiency, oxidative stress, and aberrant fatty acid metabolism in AD. The total extract of RS (TE) affected 22 of these biomarkers, with the small molecule components of RS (SN) significantly influencing 19 and the large molecule components of RS (PR) impacting 14. Nine small molecule components are likely to dominate the pharmacodynamics of RS. We constructed a target interaction network based on the corresponding bioactivities that revealed relationships amongst 11 key biomarkers, 8 active ingredients and 12 critical targets. This research illustrates the immense potential of spatial metabolomics and network pharmacology in the study of TCM, revealing the targets and mechanisms underlying herbal formulas.

Narrowband TADF emitters with high utilization of triplet excitons: theoretical insights and molecular design.

Narrowband emitters with thermally activated delayed fluorescence (TADF) features, known as multi-resonant TADF (MR-TADF) emitters, are drawing increasing research interest owing to their properties of high efficiency and excellent color purity. However, MR-TADF-based devices often face serious efficiency roll-off at high luminance intensity, which could be attributed to undesired triplet-triplet annihilation (TTA) caused by the structural planarity and relatively small reverse intersystem crossing rate constants (krisc) of MR-TADF emitters. Herein, combining a sp3-C inserted strategy to suppress harmful bimolecular interactions and chalcogens to improve the krisc, a series of asymmetric narrowband emitters, namely, DMAC-O, DMAC-S, and DMAC-Se, have been theoretically designed to break the slow rate-limiting step of krisc of experimental BN-DMAC. For comparison, both O and Se atoms were doped into the MR skeleton to substitute two sp3-inserted units, yielding BN-O-Se. The combination of TD-DFT and the wavefunction-based STEOM-DLPNO-CCSD approach exhibits that those asymmetric molecules are promising for simultaneously exhibiting narrow emission spectral full-width at half-maximums (FWHMs) and high luminous efficiencies. The contributions of chalcogens to hole distributions result in red-shifted fluorescent peaks, and the asymmetric strategy also helps with twisted molecular configuration, which is beneficial for suppressing unfavorable TTA. Furthermore, the incorporation of chalcogens is sufficient to promote the intersystem crossing and reverse intersystem crossing channels of asymmetric emitters. More importantly, the doped heavy Se atom results in a significantly increased krisc of 2.32 × 106 s-1 for DMAC-Se, which is more than 200 times larger than 1.09 × 104 s-1 of pristine BN-DMAC. These results suggest that the combination of the heavy Se atom and an sp3-inserted unit is a feasible strategy for achieving poor planarity and significantly enhancing krisc, which will help in harvesting triplet excitons, thereby inhibiting efficiency roll-off in corresponding narrowband devices.

ROCK1 inhibition improves wound healing in diabetes via RIPK4/AMPK pathway.

Refractory wounds are a severe complication of diabetes mellitus that often leads to amputation because of the lack of effective treatments and therapeutic targets. The pathogenesis of refractory wounds is complex, involving many types of cells. Rho-associated protein kinase-1 (ROCK1) phosphorylates a series of substrates that trigger downstream signaling pathways, affecting multiple cellular processes, including cell migration, communication, and proliferation. The present study investigated the role of ROCK1 in diabetic wound healing and molecular mechanisms. Our results showed that ROCK1 expression significantly increased in wound granulation tissues in diabetic patients, streptozotocin (STZ)-induced diabetic mice, and db/db diabetic mice. Wound healing and blood perfusion were dose-dependently improved by the ROCK1 inhibitor fasudil in diabetic mice. In endothelial cells, fasudil and ROCK1 siRNA significantly elevated the phosphorylation of adenosine monophosphate-activated protein kinase at Thr172 (pThr172-AMPKα), the activity of endothelial nitric oxide synthase (eNOS), and suppressed the levels of mitochondrial reactive oxygen species (mtROS) and nitrotyrosine formation. Experiments using integrated bioinformatics analysis and coimmunoprecipitation established that ROCK1 inhibited pThr172-AMPKα by binding to receptor-interacting serine/threonine kinase 4 (RIPK4). These results suggest that fasudil accelerated wound repair and improved angiogenesis at least partially through the ROCK1/RIPK4/AMPK pathway. Fasudil may be a potential treatment for refractory wounds in diabetic patients.

Borrmann Type IV Gastric Cancer: Computed Tomography Features and Corresponding Pathological Findings.

OBJECTIVE: We aimed to analyze the association between computed tomography (CT) features and the corresponding pathological findings in Borrmann type IV (BT-4) gastric cancers and explore the pathological basis of the characteristic CT features. METHODS: This retrospective study included 84 patients with BT-4 gastric cancers who underwent contrast-enhanced CT and surgical resection. Preoperative CT features were evaluated, including the major location, range, circumferential invasion, perigastric fat infiltration, enlarged lymph nodes, layered enhancement, degree of enhancement, and peak enhanced phase. Postoperative pathological findings were also recorded. Differences in CT features according to different World Health Organization types, surgical margin, adjacent organ invasion, and peritoneal status were assessed using the χ 2 or Fisher exact test (n < 5). RESULTS: The most common World Health Organization type of BT-4 gastric cancer was poorly cohesive carcinoma (65.5%), which tended to show circumferential invasion, fewer enlarged lymph nodes, and layered enhancement. Although 82 patients with BT-4 gastric cancer (97.6%) had positive lymph nodes, only 26 (31.0%) had enlarged lymph nodes. Lesions originating from the gastroesophageal junction had a higher rate of positive margins ( P < 0.05). Adjacent organ invasion was more likely to occur in lesions with perigastric fat infiltration ( P < 0.05). Patients with circumferential invasion tended to show peritoneal metastasis ( P < 0.05). CONCLUSIONS: The characteristic CT features of BT-4 gastric cancer may be attributed to the corresponding pathological findings. Recognizing the association between CT features and pathological findings may help evaluate the aggressiveness of BT-4 gastric cancers.

Serum lipid metabolism characteristics and potential biomarkers in patients with unilateral sudden sensorineural hearing loss.

BACKGROUND: Glycerophospholipids (GPLs) are essential for cell membrane structure and function. Sphingomyelin and its metabolites regulate cell growth, apoptosis, and stress responses. This study aimed to investigate lipid metabolism in patients experiencing sudden sensorineural hearing loss across all frequencies (AF-SSNHL). METHODS: The study included 60 patients diagnosed with unilateral AF-SSNHL, among whom 30 patients had a level of hearing improvement ≥ 15 dB after 6 months of follow-up. A propensity score-matched (2:1) control group was used. Liquid chromatography‒mass spectrometry based untargeted lipidomics analysis combined with multivariate statistics was performed to investigate the lipids change. The "lipidome" R package and weighted gene co-expression network analysis (WGCNA) were utilised to assess the lipids' structural features and the association between lipids and hearing. RESULTS: Lipidomics successfully differentiated the AF-SSNHL group from the control group, identifying 17 risk factors, mainly including phosphatidylcholine (PC), phosphatidylethanolamine (PE), and related metabolites. The ratios of lysophosphatidylcholine/PC, lysophosphatidylethanolamine/PE, and lysodimethylphosphatidylethanolamine/PE were upregulated, while some glycerophospholipid (GPL)-plasmalogens were downregulated in the AF-SSNHL group, indicating abnormal metabolism of GPLs. Trihexosylceramide (d34:1), PE (18:1e_22:5), and sphingomyelin (d40:3) were significantly different between responders and nonresponders, and positively correlated with hearing improvement. Additionally, the results of the WGCNA also suggested that partial GPL-plasmalogens were positively associated with hearing improvement. CONCLUSION: AF-SSNHL patients exhibited abnormally high blood lipids and pronounced GPLs metabolic abnormalities. Sphingolipids and GPL-plasmalogens had an association with the level of hearing improvement. By understanding the lipid changes, clinicians may be able to predict the prognosis of hearing recovery and personalize treatment approaches.

Foodborne Disease Outbreaks Caused by Biotoxins in Yantai City: A 10-Year Spatiotemporal Monitoring Study.

Unsafe food causes 600 million cases of foodborne diseases and 420,000 deaths every year. Meanwhile, biological toxins such as poisonous mushrooms, saponins, and aflatoxin can cause significant damage to humans. Therefore, it is particularly important to study foodborne disease outbreaks caused by biotoxins (FDOB). We collected FDOB in Yantai City from 2013 to 2022 and further established a corresponding database. Statistical analysis was carried out according to time, place, pathogen, and contamination of pathogenic factors. There were 128 FDOB, resulting in 417 patients and 6 deaths. The third quarter was a high season for foodborne disease outbreaks, the number of events, patients and deaths accounted for 65.63% (84/128), 55.88% (233/417), and 100% (6/6) of the total number, respectively. The highest number of outbreaks per 10,000 persons was Qixia (0.41), followed by Zhifu (0.36) and Laiyang (0.33). The top three causes of outbreaks were poisonous mushroom toxin, saponins and hemagglutinin, and Lagenaria siceraria (Molina) Standl. Sixty-five (50.78%) outbreaks were attributed to poisonous mushroom toxin, 18 (14.06%) outbreaks to saponin and hemagglutinin, and 12 (9.38%) outbreaks to L. siceraria (Molina) Standl. The largest number of outbreaks, patients and deaths all occurred in families, accounting for 82.81% (106/128) outbreaks, 66.19% (276/417) patients, and 100% (6/6) deaths, respectively. Followed by catering service establishments, accounting for 14.84% (19/128), 30.22% (126/417), and 0% (0/6), respectively. The main poisoning link of outbreaks was ingestion and misuse, accounting for 72.66% (93/128), followed by improper processing, accounting for 20.31% (26/128). It is necessary to carry out targeted family publicity and education, strengthen the integration of medical and prevention, explore innovative monitoring and early warning mechanisms for foodborne diseases, and reduce the occurrence of underreporting of foodborne disease outbreaks.

Carex meyeriana Kunth Extract Is a Novel Natural Drug against Candida albicans.

As a widely distributed plant in Northeast China, Carex meyeriana Kunth (CMK) is generally considered to have antibacterial properties; however, there is a lack of scientific evidence for this. Therefore, we investigated the chemical composition of CMK extract and its effect against C. albicans. A total of 105 compounds were identified in the alcohol extracts of CMK by UPLC-Q-TOF-MS. Most were flavonoids, with Luteolin being the most represented. Among them, 19 compounds are found in the C. albicans lysates. After treatment with CMK ethanol extract, a significant reduction in the number of C. albicans colonies was observed in a vaginal douche solution from day 5 (p < 0.05). Furthermore, the CMK extract can reduce the number of C. albicans spores. The levels of IL-4, IL-6, IL-10, IL-1ß, and TNF-α in vaginal tissues all exhibited a significant decrease (p < 0.05) compared to those in the model group as determined by ELISA. The results of HE staining showed that CMK extract can eliminate vaginal mucosa inflammation. CMK adjusts the vaginal mucosa cells by targeting twenty-six different metabolites and five specific metabolic pathways in order to effectively eliminate inflammation. Simultaneously, the CMK regulates twenty-three types of metabolites and six metabolic pathways against C. albicans infection. So, CMK strongly inhibits the growth of C. albicans and significantly reduces vaginal inflammation, making it a promising candidate for treating C. albicans infection.

Establishment of Polydopamine-Modified HK-2 Cell Membrane Chromatography and Screening of Active Components from Plantago asiatica L.

Cell membrane chromatography (CMC) has been widely recognized as a highly efficient technique for in vitro screening of active compounds. Nevertheless, conventional CMC approaches suffer from a restricted repertoire of cell membrane proteins, making them susceptible to oversaturation. Moreover, the binding mechanism between silica gel and proteins primarily relies on intermolecular hydrogen bonding, which is inherently unstable and somewhat hampers the advancement of CMC. Consequently, this investigation aimed to establish a novel CMC column that could augment protein loading, enhance detection throughput, and bolster binding affinity through the introduction of covalent bonding with proteins. This study utilizes polydopamine (PDA)-coated silica gel, which is formed through the self-polymerization of dopamine (DA), as the carrier for the CMC column filler. The objective is to construct the HK-2/SiO2-PDA/CMC model to screen potential therapeutic drugs for gout. To compare the quantity and characteristics of Human Kidney-2 (HK-2) cell membrane proteins immobilized on SiO2-PDA and silica gel, the proteins were immobilized on both surfaces. The results indicate that SiO2-PDA has a notably greater affinity for membrane proteins compared to silica gel, resulting in a significant improvement in detection efficiency. Furthermore, a screening method utilizing HK-2/SiO2-PDA/CMC was utilized to identify seven potential anti-gout compounds derived from Plantago asiatica L. (PAL). The effectiveness of these compounds was further validated using an in vitro cell model of uric acid (UA) reabsorption. In conclusion, this study successfully developed and implemented a novel CMC filler, which has practical implications in the field.

Monitoring landfill volatile organic compounds emissions by an uncrewed aerial vehicle platform with infrared and visible-light cameras, remote monitoring, and sampling systems.

An uncrewed aerial vehicle (UAV) platform equipped with dual imaging cameras, a gas sampling system, and a remote synchronous monitoring system was developed to sample and analyze volatile organic compounds (VOCs) emitted from landfills. The remote synchronous monitoring system provided real-time video to administrators with specific permissions to assist in identifying sampling sites within extensive landfill areas. The sampling system included four kits capable of collecting samples from different locations during a single flight mission. Each kit comprised a 1 L Tedlar bag for measuring landfill VOC concentrations according to the TO-15 method prescribed by the US Environmental Protection Agency. The air sample was introduced into a Tedlar bag via pumping. A known volume of the sample was subsequently concentrated using a solid multisorbent concentrator. Following this, the sample underwent cold trap concentration and thermal desorption. The concentrated sample was then transferred to a chromatography-mass spectrometry system for separation and analysis. Since the anaerobic catabolism of organic waste is exothermic and emits VOCs, this study employed UAV thermal imaging to locate principal emission sources for sampling. Visible-light imaging helped identify newer or older landfill sections, aiding in the selection of appropriate sampling sites, particularly when surfaces were thermally disturbed by solar radiation. Field measurements were conducted under three meteorological conditions: sunny morning, cirrus morning, and thin cloud evening (2 h after sunset), identifying 119, 122, and 111 chemical species respectively. The sequence of total VOC concentrations measured correlated with the meteorological conditions as follows: cirrus morning > thin cloud evening > sunny morning. The results indicated that ambient temperature and global solar radiation significantly influenced daytime gas emissions from landfills. Evening thermal images, unaffected by solar heating, facilitated more accurate identification of major VOC emission points, resulting in higher VOC concentrations compared to those recorded in the sunny morning. VOCs from the landfill were categorized into nine organic groups: alkanes, alkenes, carbonyls, aromatics, alcohols, esters, ethers, organic oxides, and others. The classification was based on carbon-containing compounds (Cn, where the compound contains n carbon atoms). Alkanes were predominant in terms of Cn presence, followed by alcohols and carbonyls. Among the organic groups, organic oxides, particularly 2-heptyl-1,3-dioxolane, exhibited the highest concentrations, succeeded by alkenes. Sampling under cloudy conditions or in the evening is recommended to minimize the effects of surface temperature anomalies caused by solar radiation, which vary due to differences in land composition.

Promoting forest landscape dynamic prediction with an online collaborative strategy.

Modeling and predicting forest landscape dynamics are crucial for forest management and policy making, especially under the context of climate change and increased severities of disturbances. As forest landscapes change rapidly due to a variety of anthropogenic and natural factors, accurately and efficiently predicting forest dynamics requires the collaboration and synthesis of domain knowledge and experience from geographically dispersed experts. Owing to advanced web techniques, such collaboration can now be achieved to a certain extent, for example, discussion about modeling methods, consultation for model use, and surveying for stakeholders' feedback can be conducted on the web. However, a research gap remains in terms of how to facilitate online joint actions in the core task of forest landscape modeling by overcoming the challenges from decentralized and heterogeneous data, offline model computation modes, complex simulation scenarios, and exploratory modeling processes. Therefore, we propose an online collaborative strategy to enable collaborative forest landscape dynamic prediction with four core modules, namely data preparation, forest landscape model (FLM) computation, simulation scenario configuration, and process organization. These four modules are designed to support: (1) voluntary data collection and online processing, (2) online synchronous use of FLMs, (3) collaborative simulation scenario design, altering, and execution, and (4) participatory modeling process customization and coordination. We used the LANDIS-II model as a representative FLM to demonstrate the online collaborative strategy for predicting the dynamics of forest aboveground biomass. The results showed that the online collaboration strategy effectively promoted forest landscape dynamic prediction in data preparation, scenario configuration, and task arrangement, thus supporting forest-related decision making.

Safety and antitumour activity of cadonilimab, an anti-PD-1/CTLA-4 bispecific antibody, for patients with advanced solid tumours (COMPASSION-03): a multicentre, open-label, phase 1b/2 trial.

BACKGROUND: Immune checkpoint inhibitors targeting PD-1 or CTLA-4 individually have shown substantial clinical benefits in the treatment of malignancies. We aimed to assess the safety and antitumour activity of cadonilimab monotherapy, a bispecific PD-1/CTLA-4 antibody, in patients with advanced solid tumours. METHODS: This multicentre, open-label, phase 1b/2 trial was conducted across 30 hospitals in China. Patients aged 18 years or older with histologically or cytologically confirmed, unresectable advanced solid tumours, unsuccessful completion of at least one previous systemic therapy, and an Eastern Cooperative Oncology Group performance status of 0 or 1 were eligible for inclusion. Patients who had previously received anti-PD-1, anti-PD-L1, or anti-CTLA-4 treatment were not eligible for inclusion. In the dose escalation phase of phase 1b, patients received intravenous cadonilimab at 6 mg/kg and 10 mg/kg every 2 weeks. In the dose expansion phase of phase 1b, cadonilimab at 6 mg/kg and a fixed dose of 450 mg were given intravenously every 2 weeks. In phase 2, cadonilimab at 6 mg/kg was administered intravenously every 2 weeks in three cohorts: patients with cervical cancer, oesophageal squamous cell carcinoma, and hepatocellular carcinoma. The primary endpoints were the safety of cadonilimab in phase 1b and objective response rate in phase 2, based on the Response Evaluation Criteria in Solid Tumors (RECIST), version 1.1. The safety analysis was done in all patients who received at least one dose of cadonilimab. Antitumour activity was assessed in the full analysis set for the cervical cancer cohort, and in all patients with measurable disease at baseline and who received at least one dose of cadonilimab in the oesophageal squamous cell carcinoma and hepatocellular carcinoma cohorts. The study is registered on ClinicalTrial.gov, NCT03852251, and closed to new participants; follow-up has been completed. FINDINGS: Between Jan 18, 2019, and Jan 8, 2021, 240 patients (83 [43 male and 40 female] in phase 1b and 157 in phase 2) were enrolled. Phase 2 enrolled 111 female patients with cervical cancer, 22 patients with oesophageal squamous cell carcinoma (15 male and seven female), and 24 patients with hepatocellular carcinoma (17 male and seven female). During dose escalation, no dose-limiting toxicities occurred. Grade 3-4 treatment-related adverse events occurred in 67 (28%) of 240 patients; the most frequent grade 3 or worse treatment-related adverse events were anaemia (seven [3%]), increased lipase (four [2%]), decreased bodyweight (three [1%]), decreased appetite (four [2%]), decreased neutrophil count (three [1%]), and infusion-related reaction (two [1%]). 17 (7%) patients discontinued treatment due to treatment-related adverse events. 54 (23%) of 240 patients reported serious treatment-related adverse events, including five patients who died (one due to myocardial infarction; cause unknown for four). In phase 2, in the cervical cancer cohort, with a median follow-up of 14·6 months (IQR 13·1-17·5), the objective response rate was 32·3% (32 of 99; 95% CI 23·3-42·5). In the oesophageal squamous cell carcinoma cohort, with a median follow-up of 17·9 months (IQR 4·0-15·1), the objective response rate was 18·2% (four of 22; 95% CI 5·2-40·3). In the hepatocellular carcinoma cohort, with a median follow-up of 19·6 months (IQR 8·7-19·8), the objective response rate was 16·7% (four of 24; 95% CI 4·7-37·4). INTERPRETATION: Cadonilimab showed an encouraging tumour response rate, with a manageable safety profile, suggesting the potential of cadonilimab for the treatment of advanced solid tumours. FUNDING: Akeso Biopharma. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

The METTL5-TRMT112 N6-methyladenosine methyltransferase complex regulates mRNA translation via 18S rRNA methylation.

Ribosomal RNAs (rRNAs) have long been known to carry chemical modifications, including 2'O-methylation, pseudouridylation, N6-methyladenosine (m6A), and N6,6-dimethyladenosine. While the functions of many of these modifications are unclear, some are highly conserved and occur in regions of the ribosome critical for mRNA decoding. Both 28S rRNA and 18S rRNA carry single m6A sites, and while the methyltransferase ZCCHC4 has been identified as the enzyme responsible for the 28S rRNA m6A modification, the methyltransferase responsible for the 18S rRNA m6A modification has remained unclear. Here, we show that the METTL5-TRMT112 methyltransferase complex installs the m6A modification at position 1832 of human 18S rRNA. Our work supports findings that TRMT112 is required for METTL5 stability and reveals that human METTL5 mutations associated with microcephaly and intellectual disability disrupt this interaction. We show that loss of METTL5 in human cancer cell lines and in mice regulates gene expression at the translational level; additionally, Mettl5 knockout mice display reduced body size and evidence of metabolic defects. While recent work has focused heavily on m6A modifications in mRNA and their roles in mRNA processing and translation, we demonstrate here that deorphanizing putative methyltransferase enzymes can reveal previously unappreciated regulatory roles for m6A in noncoding RNAs.

Asymmetric Total Synthesis of (+)-Phainanoid A and Biological Evaluation of the Natural Product and Its Synthetic Analogues.

Here, we report our detailed efforts toward the synthesis of phainanoids, a novel class of dammarane-type triterpenoids with potent immunosuppressive activities and unique structural features. Systematic model studies have been carried out, and efficient approaches have been established to construct the benzofuranone-based 4,5-spirocycle, the D/E/F tricyclic core, the [4.3.1] propellane, and the 5,5-oxaspirolactone moieties. The asymmetric synthesis of (+)-phainanoid A has been achieved through kinetic resolution of the tricyclic core followed by diastereoselective installation of the A/B/C and G/H rings and fragment coupling with the enantioenriched I/J rings. In addition, novel estrone-derived phainanoid analogues have been prepared. The immunosuppressive and cell survival assays revealed that (+)-phainanoid A and some of its synthetic analogues can specifically inhibit stimulation-induced lymphocyte proliferation but not cell survival at their effective concentrations. Preliminary structure-activity relationship information has been obtained, which could inspire future design of immunosuppressive phainanoid analogues.

HMGB1 accumulation in cytoplasm mediates noise-induced cochlear damage.

Damage-associated molecular pattern molecules (DAMPs) play a critical role in mediating cochlear cell death, which leads to noise-induced hearing loss (NIHL). High-mobility group box 1 (HMGB1), a prototypical DAMP released from cells, has been extensively studied in the context of various diseases. However, whether extracellular HMGB1 contributes to cochlear pathogenesis in NIHL and the potential signals initiating HMGB1 release from cochlear cells are not well understood. Here, through the transfection of the adeno-associated virus with HMGB1-HA-tag, we first investigated early cytoplasmic accumulation of HMGB1 in cochlear hair cells after noise exposure. We found that the cochlear administration of HMGB1-neutralizing antibody immediately after noise exposure significantly alleviated hearing loss and outer hair cells (OHCs) death induced by noise exposure. In addition, activation of signal transducer and activators of transcription 1 (STAT1) and cellular hyperacetylation were verified as potential canonical initiators of HMGB1 cytoplasmic accumulation. These findings reveal the adverse effects of extracellular HMGB1 on the cochlea and the potential signaling events mediating HMGB1 release in hair cells, indicating multiple potential pharmacotherapeutic targets for NIHL.

Shikonin inhibits neuronal apoptosis via regulating endoplasmic reticulum stress in the rat model of double-level chronic cervical cord compression.

Cervical spondylotic myelopathy (CSM) is a clinically symptomatic entity arising from the spinal cord compression by degenerative diseases. Although endoplasmic reticulum (ER) stress has been commonly observed in several neurodegenerative diseases, the relationship between ER stress and CSM remains unknown. Shikonin is known to protect PC12 by inhibiting apoptosis in vitro. This study hypothesised that ER stress was vital in neuronal apoptosis in CSM. Shikonin might inhibit such responses by regulating ER stress through the protein kinase-like ER kinase-eukaryotic translation initiation factor 2 α-subunit-C/EBP homologous protein (PERK-eIF2α-CHOP) signalling pathway. Thus, the aim of this study was evaluating the neuroprotective effect of shikonin in rats with double-level chronic cervical cord compression, as well as primary rat cortical neurons with glutamate-induced neurotoxicity. The result showed that ER stress-related upregulation of PERK-eIF2α-CHOP resulted in rat neuronal apoptosis after chronic cervical cord compression; then, shikonin promoted motor recovery and inhibited neuronal apoptosis by attenuating PERK-eIF2α-CHOP and prevented Bax translocation from cytoplasm to mitochondrion induced by CHOP of neurons in rats with chronic compression. Also, it was found that shikonin could protect rat primary cortical neuron against glutamate toxicity by regulating ER stress through the PERK-eIF2α-CHOP pathway in vitro. In conclusion, shikonin might inhibit neuronal apoptosis by regulating ER stress through attenuating the activation of PERK-eIF2α-CHOP.

Ligand-targeted fishing of α-glucosidase inhibitors from Tribulus terrestris L. based on chitosan-functionalized multi-walled carbon nanotubes with immobilized α-glucosidase.

α-Glucosidase inhibitors in natural products are one of the promising drugs for the treatment of type 2 diabetes. However, due to the complexity of the matrix, it is challenging to comprehensibly clarify the specific pharmacodynamic substances. In this study, a novel high-throughput inhibitor screening strategy was established based on covalent binding of α-glucosidase on chitosan-functionalized multi-walled carbon nanotubes coupled with high-resolution mass spectrometry. The synthesized MWCNTs@CS@GA@α-Glu was characterized by TEM, SEM, FTIR, Raman, and TG. Performance studies showed that the microreactor exhibited stronger thermostability and pH tolerance than that of the free one while maintaining its inherent catalytic activity. Feasibility study applying a model mixture of known α-glucosidase ligand and non-ligands indicated the selectivity and specificity of the system. By integrating ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-QTOF-MS) with ion mobility mass spectrometry (IMS), 15 ligands were obtained and tentatively identified from Tribulus terrestris L., including 8 steroidal saponins, 4 flavonoids, and 3 alkaloids. These inhibitors were further validated by in vivo experiments and molecular docking simulation.