Discovery of highly potent covalent SARS-CoV-2 3CLpro inhibitors bearing 2-sulfoxyl-1,3,4-oxadiazole scaffold for combating COVID-19.

The coronavirus disease (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a major public health crisis, posing a significant threat to human well-being. Despite the availability of vaccines, COVID-19 continues to spread owing to the emergence of SARS-CoV-2 mutants. This highlights the urgent need for the discovery of more effective drugs to combat COVID-19. As an important target for COVID-19 treatment, 3C-like protease (3CLpro) plays a crucial role in the replication of SARS-CoV-2. In our previous research, we demonstrated the potent inhibitory activities of compound A1, which contains a 2-sulfonyl-1,3,4-oxadiazole scaffold, against SARS-CoV-2 3CLpro. Herein, we present a detailed investigation of structural optimization of A1 and conduct a study on the structure-activity relationship. Among the various compounds tested, sulfoxide D6 demonstrates a potent irreversible inhibitory activity (IC50 = 0.030 µM) against SARS-CoV-2 3CLpro, as well as a favorable selectivity towards host cysteine proteases such as cathepsin B and cathepsin L. Utilizing mass spectrometry-based peptide profiling, we found that D6 covalently binds to Cys145 of SARS-CoV-2 3CLpro. Some representative compounds, namely C11, D9 and D10 also demonstrates antiviral activity against SARS-CoV-2 in Vero E6 cells. Overall, the investigation of the 2-sulfoxyl-1,3,4-oxadiazole scaffold as a novel cysteine reactive warhead would provide valuable insights into the design of potent covalent 3CLpro inhibitors for COVID-19 treatment.

Description of two novel anaerobic members in the family Clostridiaceae, Anaeromonas gelatinilytica gen.nov., sp. nov., and Anaeromonas frigoriresistens sp. nov., isolated from saline lake sediment.

Cells of members of the family Clostridiaceae, phylum Firmicutes, are generally obligate anaerobic rods. Strains D2Q-14T and D2Q-11T were isolated from sediment of the saline lake Manisi in the Xinjiang Uygur Autonomous Region, PR China. In this study, we employed a polyphasic approach and whole genome analysis of the two isolates. Cells of both isolates were Gram-stain-positive rods that were motile by means of flagella and could utilize sulphate, thiosulphate, elemental sulphur and nitrate as electron acceptors. Phylogenetic analyses based on 16S rRNA gene and whole genome sequences indicated that strains D2Q-14T and D2Q-11T constituted a coherent cluster affiliated to the family Clostridiaceae. In addition, genome analysis revealed that strain D2Q-14Tharboured one nonribosomal peptide synthetase gene cluster, making up 1.4 % of the entire genome. The genome-based analysis, including average nucleotide identity, average amino acid identity and in silico DNA-DNA hybridization, biochemical, phenotypic and chemotaxonomic characterization, indicated that strains D2Q-14T and D2Q-11T represented two novel species of a novel genus in the family Clostridiaceae, for which we propose the names Anaeromonas gelatinilytica gen. nov., sp. nov. and Anaeromonas frigoriresistens sp. nov., with the type strains D2Q-14T (=KCTC 15986T=MCCC 1K04634T) and D2Q-11T (=KCTC 15985T=MCCC 1K04391T), respectively.

Identifying Circular RNA and Predicting Its Regulatory Interactions by Machine Learning.

Circular RNA (circRNA) is a closed long non-coding RNA (lncRNA) formed by covalently closed loops through back-splicing. Emerging evidence indicates that circRNA can influence cellular physiology through various molecular mechanisms. Thus, accurate circRNA identification and prediction of its regulatory information are critical for understanding its biogenesis. Although several computational tools based on machine learning have been proposed for circRNA identification, the prediction accuracy remains to be improved. Here, first we present circLGB, a machine learning-based framework to discriminate circRNA from other lncRNAs. circLGB integrates commonly used sequence-derived features and three new features containing adenosine to inosine (A-to-I) deamination, A-to-I density and the internal ribosome entry site. circLGB categorizes circRNAs by utilizing a LightGBM classifier with feature selection. Second, we introduce circMRT, an ensemble machine learning framework to systematically predict the regulatory information for circRNA, including their interactions with microRNA, the RNA binding protein, and transcriptional regulation. Feature sets including sequence-based features, graph features, genome context, and regulatory information features were modeled in circMRT. Experiments on public and our constructed datasets show that the proposed algorithms outperform the available state-of-the-art methods. circLGB is available at http://www.circlgb.com. Source codes are available at https://github.com/Peppags/circLGB-circMRT.

Anaerophilus nitritogenes gen. nov., sp. nov., isolated from salt lake sediment in Xinjiang Province, China.

An obligately anaerobic, nitrate-reducing bacterial strain (MJB2T) was isolated from sediments of saline in Xinjiang province of China. Cells were Gram-stain-positive rods and motile by means of flagella and formed endospores. The novel strain MJB2T was able to grow at 15-37 °C (optimum 28-30 °C), pH 5.8-9.4 (optimum 7.8) and with 1.0-7.0% NaCl (optimum 5.0-6.0%, w/v). Sulfate, sulfite, thiosulfate, elemental sulfur, nitrite and Fe(III) were not used as terminal electron acceptors. Oxidase and catalase reactions were positive. H2S was producted from L-cystine. Complex substrates such as beef extract, peptone and yeast extract can be used as sole energy sources. The DNA G+C content was 29.4 mol%. The major cellular fatty acids (> 10%) were C14:0, C16:1 cis 7 and C16:1 cis 9. The main polar lipids consisted of phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, three unidentified amino lipids, one unidentified amino glycolipid, two unidentified glycolipid, one unidentified aminophospholipid and two unidentified lipids. No respiratory quinones were detected. According to phylogenetic analysis based on 16S rRNA gene sequences, strain MJB2T was affiliated to the family Clostridiaceae (order Clostridiales) with highest 16S rRNA gene sequence similarity of 95.3% to Crassaminicella profunda Ra1766HT. Strain MJB2T exhibited 74.9% ANI values to C. profunda Ra1766HT. In silico DNA-DNA relatedness value between strain MJB2T and C. profunda Ra1766HT was 19.5%. The distinct biochemical, chemotaxonomic and phylogenetic differences from the previously described taxa supported that strain MJB2T represents a novel species of a new genus, for which the name Anaerophilus nitritogenes gen. nov., sp. nov. is proposed. The type strain is MJB2T (=KCTC 15800T=MCCC 1K03631T).

Mining the deep Red-Sea brine pool microbial community for anticancer therapeutics.

BACKGROUND: Microbial species in the brine pools of the Red Sea and the brine pool-seawater interfaces are exposed to high temperature, high salinity, low oxygen levels and high concentrations of heavy metals. As adaptations to these harsh conditions require a large suite of secondary metabolites, these microbes have a huge potential as a source of novel anticancer molecules. METHODS: A total of 60 ethyl-acetate extracts of newly isolated strains from extreme environments of the Red-Sea were isolated and tested against several human cancer cell lines for potential cytotoxic and apoptotic activities. RESULTS: Isolates from the Erba brine-pool accounted for 50% of active bacterial extracts capable of inducing 30% or greater inhibition of cell growth. Among the 60 extracts screened, seven showed selectivity towards triple negative BT20 cells compared to normal fibroblasts. CONCLUSION: In this study, we identified several extracts able to induce caspase-dependent apoptosis in various cancer cell lines. Further investigations and isolation of the active compounds of these Red Sea brine pool microbes may offer a chemotherapeutic potential for cancers with limited treatment options.

Meridianimarinicoccus roseus gen. nov., sp. nov., a novel genus of the family Rhodobacteraceae isolated from seawater.

A Gram-stain negative, pink-pigmented, strictly aerobic, non-motile and coccoid-shaped bacterial strain, designated TG-679T, was isolated from a deep-sea water sample collected from the South China Sea. Cells of strain TG-679T were catalase- and oxidase-positive, lacked bacteriochlorophyll a and carotenoid. Strain TG-679T was found to grow at 10-40 °C (optimum, 28 °C), pH 5.5-10.0 (optimum, pH 7.5) and with 0.5-2.0 % (w/v) NaCl (optimum, 1.5 %). Chemotaxonomic analysis of strain TG-679T indicated that the sole respiratory quinone was Q-10, the predominant cellular fatty acid was C18 : 1ω7c, and the major polar lipids consisted of phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylglycerol, phosphatidylcholine, an unidentified phospholipid and two unidentified glycolipids. The genomic DNA G+C content of strain TG-679T was 65.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain TG-679T constituted a separated branch in the family Rhodobacteraceae. Differential phenotypic properties, together with phylogenetic distinctiveness, demonstrated that strain TG-679T is clearly distinct from any validly published genus. Based on polyphasic taxonomic characterization, strain TG-679T is considered to represent a novel species of a novel genus, for which the name Meridianimarinicoccus roseus gen. nov., sp. nov. is proposed. The type strain of the species is TG-679T (=KCTC 62454T=MCCC 1K03496T).

Exogenous Amino Acid-Loaded Nanovehicles: Stepping across Endogenous Magnetic Resonance Spectroscopy.

Magnetic resonance spectroscopy (MRS) allows the assessment of metabolic contents and biochemical information in vivo. It provides essential compositional information in the diagnosis and monitoring of central nervous system (CNS) diseases, especially brain tumors. Conventional MRS is usually confined to endogenous metabolites that may lack specificity for certain disease such as differentiating glioma from other tumor and non-tumorous lesions. Therefore, exogenous MRS contrast agents (CAs) that may improve the sensitivity and specificity of MRS are highly desirable for its clinical use. In this work, a novel MRS CA, ß-alanine loaded hollow mesoporous silica nanospheres, with a high biosafety profile and characteristic MRS spectrum was synthesized and investigated. This new CA is further tested to realize the accurate functional MRS diagnosis of brain glioma with high sensitivity. The general approach of synthesizing disease specific CA for MRS may lead to a new era of molecular imaging.

Spiciferone analogs from an endophytic fungus Phoma betae collected from desert plants in West China.

Endophytic fungi from desert, arid, and grassland areas are an ecologically important but unique group with poor chemical investigation. During our ongoing study to mine bioactive secondary metabolites from unique fungal environments, a new shunt product spiciferone F (1) including two new analogs spiciferones G (2) and H (3) together with four known ones spiciferone A (4), spiciferol A (5), 6, and 7 were isolated from endophytic fungus Phoma betae inhabiting in plant Kalidium foliatum (Pall.) Moq from Ningxia Province of West China. The planar, relative, and absolute configurations of these new compounds were elucidated by nuclear magnetic resonance, high-resolution electrospray ionization mass spectrometry, and electronic circular dichroism experiments. According to the shunt products, intermediates and analogs isolated from this endophytic fungus, the possible biosynthetic pathway of spiciferones was reconstructed. Compounds 1-7 were evaluated cytotoxic activities against three cancer cell lines HCT 116, HeLa, and MCF7, and only did 1 display strong biological effect against MCF7 with a half-maximal inhibitory concentration value at 7.73 ± 0.11 µM compared with the cis-platinum (14.32 ± 1.01 µM).

Copper chloride dose-dependently alters spatial learning and memory, and glutamate levels, in the hippocampus of rats.

Copper is a trace element which exerts an important role in neuronal functions. Excessive Cu exposure is associated with central nervous system dysfunction, including memory loss. The present study examined the effects of CuCl2 exposure on the spatial learning and memory of rats, and on metabolites in the hippocampus. A total of 60 male Sprague­Dawley rats (10 rats/group) were intraperitoneally injected with various doses (0, 0.5, 1.0, 2.0, 4.0 and 6.0 mg/kg) of CuCl2 three times every other day for 6 days. Rats administered with 1.0 ml/kg sterile saline were used as controls. A total of 2 days subsequent to the final injection, the rats were subjected to the Morris water maze (MWM) test, followed by proton magnetic resonance spectroscopy (1H­MRS). The rats were subsequently sacrificed, and their hippocampal tissues were processed for high performance liquid chromatography (HPLC). The MWM test demonstrated that the high­dose groups exhibited worse spatial learning and memory compared with the controls; however, the rats that received a relatively low dose (2.0 mg/kg) exhibited a decreased impairment. The 1H­MRS results revealed increased Glu, N­acetyl­L­aspartate (NAA)+N­acetylaspartylglutamate, phosphocreatine (PCr) and Cr+PCr levels in the hippocampus of the 2.0 mg/kg group. HPLC analysis revealed increased γ­aminobutyric acid and glutamate (Glu) levels in the 2.0 mg/kg group, and decreased Glu levels in the 6.0 mg/kg group. The results of the present study demonstrated a beneficial effect of short­term exposure to a relatively low dose of CuCl2 on spatial learning and memory, and the association of this effect with increased NAA and Glu levels in the hippocampus.

Trichoderpyrone, a Unique Polyketide Hybrid with a Cyclopentenone-Pyrone Skeleton from the Plant Endophytic Fungus Trichoderma gamsii.

Trichoderpyrone (1), a unique polyketide with a cyclopentenone-pyrone hybrid skeleton, was isolated from the plant endophytic fungus Trichoderma gamsii. The structure of 1 was determined by detailed analysis of NMR data together with comparison of chemical shift values of similar fragments. The relative and absolute configurations were established by NOESY correlations and CD experiment. Trichoderpyrone (1) displayed weak cytotoxic activities against A549, HepG2, and HeLa cancer cell lines. 1 might originate from a hybrid biosynthetic pathway through two nonreduced (NR) polyketide megasynthetases.

Nuclear Overhauser Enhancement-Mediated Magnetization Transfer Imaging in Glioma with Different Progression at 7 T.

Glioma is a malignant neoplasm affecting the central nervous system. The conventional approaches to diagnosis, such as T1-weighted imaging (T1WI), T2-weighted imaging (T2WI), and contrast-enhanced T1WI, give an oversimplified representation of anatomic structures. Nuclear Overhauser enhancement (NOE) imaging is a special form of magnetization transfer (MT) that provides a new way to detect small solute pools through indirect measurement of attenuated water signals, and makes it possible to probe semisolid macromolecular protons. In this study, we investigated the correlation between the effect of NOE-mediated imaging and progression of glioma in a rat tumor model. We found that the NOE signal decreased in tumor region, and signal of tumor center and peritumoral normal tissue markedly decreased with growth of the glioma. At the same time, NOE signal in contralateral normal tissue dropped relatively late (at about day 16-20 after implanting the glioma cells). NOE imaging is a new contrast method that may provide helpful insights into the pathophysiology of glioma with regard to mobile proteins, lipids, and other metabolites. Further, NOE images differentiate normal brain tissue from glioma tissue at a molecular level. Our study indicates that NOE-mediated imaging is a new and promising approach for estimation of tumor progression.

Platycodin D isolated from the aerial parts of Platycodon grandiflorum protects alcohol-induced liver injury in mice.

Platycodin D (PD) is the main active saponin of Platycodon grandiflorum (PG) and is reported to exhibit multiple biological effects, including anti-tumor, anti-inflammation, and anti-obesity properties. Although recently there have been many research reports on the chemical constituents of the plant's roots, only few works have been reported on the aerial parts of PG. In the present study, we report the first isolation of PD from the aerial parts of PG and its protective effect against acute alcohol-induced liver oxidative injury and inflammatory response in mice. In brief, the protective effect was evaluated by tracking biochemical markers, enzymatic antioxidants and proinflammatory cytokines in serum and liver tissue. The results indicated that PD pretreatment significantly decreased the levels of triglyceride (TG), total cholesterol (TC), low density lipoprotein cholesterol (L-DLC) in serum and malondialdehyde (MDA) in liver. PD was also found to increase the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) in the liver (p < 0.05). In addition, PD markedly decreased the levels of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, and IL-6, which was caused by alcohol exposure (p < 0.05). In contrast, histopathological examinations revealed that PD pretreatment noticeably prevented alcohol-induced hepatocyte apoptosis and steatosis. Collectively, the present study clearly suggests that the protective effect exhibited by PD on alcohol-induced liver oxidative injury may occur via the alleviation of oxidative stress and inflammatory response.

Metabolic characterization of all-trans-retinoic acid (ATRA)-induced craniofacial development of murine embryos using in vivo proton magnetic resonance spectroscopy.

AIM: To characterize the abnormal metabolic profile of all-trans-retinoic acid (ATRA)-induced craniofacial development in mouse embryos using proton magnetic resonance spectroscopy (1H-MRS). METHODS: Timed-pregnant mice were treated by oral gavage on the morning of embryonic gestation day 11 (E11) with all-trans-retinoic acid (ATRA). Dosing solutions were adjusted by maternal body weight to provide 30, 70, or 100 mg/kg RA. The control group was given an equivalent volume of the carrier alone. Using an Agilent 7.0 T MR system and a combination of surface coil coils, a 3 mm×3 mm×3 mm 1H-MRS voxel was selected along the embryonic craniofacial tissue. 1H-MRS was performed with a single-voxel method using PRESS sequence and analyzed using LCModel software. Hematoxylin and eosin was used to detect and confirm cleft palate. RESULT: 1H-MRS revealed elevated choline levels in embryonic craniofacial tissue in the RA70 and RA100 groups compared to controls (P<0.05). Increased choline levels were also found in the RA70 and RA100 groups compared with the RA30 group (P<0.01). High intra-myocellular lipids at 1.30 ppm (IMCL13) in the RA100 group compared to the RA30 group were found (P<0.01). There were no significant changes in taurine, intra-myocellular lipids at 2.10 ppm (IMCL21), and extra-myocellular lipids at 2.30 ppm (EMCL23). Cleft palate formation was observed in all fetuses carried by mice administered 70 and 100 mg/kg RA. CONCLUSIONS: This novel study suggests that the elevated choline and lipid levels found by 1H-MRS may represent early biomarkers of craniofacial defects. Further studies will determine performance of this test and pathogenetic mechanisms of craniofacial malformation.

Synthesis and properties of the metallo-supramolecular polymer hydrogel poly[methyl vinyl ether-alt-mono-sodium maleate]·AgNO3: Ag+/Cu2+ ion exchange and effective antibacterial activity.

The commercial polymeric anhydride poly(methyl vinyl ether-alt-maleic anhydride) (PVM/MA) is converted by reaction with NaOH to give poly(methyl vinyl ether-alt-mono-sodium maleate) (PVM/Na-MA). By addition of AgNO3-solution, the formation of the silver(i) supramolecular polymer hydrogel poly[methyl vinyl ether-alt-mono-sodium maleate]·AgNO3 is reported. Freeze-dried samples of the hydrogel show a mesoporous network of polycarboxylate ligands that are crosslinked by silver(i) cations. In the intact hydrogel, ion-exchange studies are reported and it is shown that Ag+ ions can be exchanged by copper(ii) cations without disintegration of the hydrogel. The silver(i) hydrogel shows effective antibacterial activity and potential application as burn wound dressing.

Induction of apoptosis in cancer cell lines by the Red Sea brine pool bacterial extracts.

BACKGROUND: Marine microorganisms are considered to be an important source of bioactive molecules against various diseases and have great potential to increase the number of lead molecules in clinical trials. Progress in novel microbial culturing techniques as well as greater accessibility to unique oceanic habitats has placed the marine environment as a new frontier in the field of natural product drug discovery. METHODS: A total of 24 microbial extracts from deep-sea brine pools in the Red Sea have been evaluated for their anticancer potential against three human cancer cell lines. Downstream analysis of these six most potent extracts was done using various biological assays, such as Caspase-3/7 activity, mitochondrial membrane potential (MMP), PARP-1 cleavage and expression of γH2Ax, Caspase-8 and -9 using western blotting. RESULTS: In general, most of the microbial extracts were found to be cytotoxic against one or more cancer cell lines with cell line specific activities. Out of the 13 most active microbial extracts, six extracts were able to induce significantly higher apoptosis (>70%) in cancer cells. Mechanism level studies revealed that extracts from Chromohalobacter salexigens (P3-86A and P3-86B(2)) followed the sequence of events of apoptotic pathway involving MMP disruption, caspase-3/7 activity, caspase-8 cleavage, PARP-1 cleavage and Phosphatidylserine (PS) exposure, whereas another Chromohalobacter salexigens extract (K30) induced caspase-9 mediated apoptosis. The extracts from Halomonas meridiana (P3-37B), Chromohalobacter israelensis (K18) and Idiomarina loihiensis (P3-37C) were unable to induce any change in MMP in HeLa cancer cells, and thus suggested mitochondria-independent apoptosis induction. However, further detection of a PARP-1 cleavage product, and the observed changes in caspase-8 and -9 suggested the involvement of caspase-mediated apoptotic pathways. CONCLUSION: Altogether, the study offers novel findings regarding the anticancer potential of several halophilic bacterial species inhabiting the Red Sea (at the depth of 1500-2500 m), which constitute valuable candidates for further isolation and characterization of bioactive molecules.