Bimetallic Nanozyme: A Credible Tag for In Situ-Catalyzed Reporter Deposition in the Lateral Flow Immunoassay for Ultrasensitive Cancer Diagnosis.

The lateral flow immunoassay (LFIA) is a sought-after point-of-care testing platform, yet the insufficient sensitivity of the LFIA limits its application in the detection of tumor biomarkers. Here, a colorimetric signal amplification method, bimetallic nanozyme-mediated in situ-catalyzed reporter deposition (BN-ISCRD), was designed for ultrasensitive cancer diagnosis. The bimetallic nanozyme used, palladium@iridium core-shell nanoparticles (Pd@Ir NPs), had ultrahigh enzyme-like activity, which was further explained by the electron transfer of Pd@Ir NPs and the change in the Gibbs free energy during catalysis through density functional theory calculations. With gastric cancer biomarkers pepsinogen I and pepsinogen II as model targets, this assay could achieve a cutoff value of 10 pg/mL, which was 200-fold lower than that without signal enhancement. The assay was applied to correctly identify 8 positive and 28 negative clinical samples. Overall, this BN-ISCRD-based LFIA showed great merits and potential in the application of ultrasensitive disease diagnosis.

Chemical Synthesis and Antigenicity Evaluation of an Aminoglycoside Trisaccharide Repeating Unit of Pseudomonas aeruginosa Serotype O5 O-Antigen Containing a Rare Dimeric-ManpN3NA.

Pseudomonas aeruginosa bacteria are becoming increasingly resistant against multiple antibiotics. Therefore, the development of vaccines to prevent infections with these bacteria is an urgent medical need. While the immunological activity of lipopolysaccharide O-antigens in P. aeruginosa is well-known, the specific protective epitopes remain unidentified. Herein, we present the first chemical synthesis of highly functionalized aminoglycoside trisaccharide 1 and its acetamido derivative 2 found in the P. aeruginosa serotype O5 O-antigen. The synthesis of the trisaccharide targets is based on balancing the reactivity of disaccharide acceptors and monosaccharide donors. Glycosylations were analyzed by quantifying the reactivity of the hydroxyl group of the disaccharide acceptor using the orbital-weighted Fukui function and dual descriptor. The stereoselective formation of 1,2-cis-α-fucosylamine linkages was achieved through a combination of remote acyl participation and reagent modulation. The simultaneous SN2 substitution of azide groups at C2' and C2″ enabled the efficient synthesis of 1,2-cis-ß-linkages for both 2,3-diamino-D-mannuronic acids. Through a strategic orthogonal modification, the five amino groups on target trisaccharide 1 were equipped with a rare acetamidino (Am) and four acetyl (Ac) groups. Glycan microarray analyses of sera from patients infected with P. aeruginosa indicated that trisaccharides 1 and 2 are key antigenic epitopes of the serotype O5 O-antigen. The acetamidino group is not an essential determinant of antibody binding. The ß-D-ManpNAc3NAcA residue is a key motif for the antigenicity of serotype O5 O-antigen. These findings serve as a foundation for the development of glycoconjugate vaccines targeting P. aeruginosa serotype O5.

TEAD4 antagonizes cellular senescence by remodeling chromatin accessibility at enhancer regions.

Dramatic alterations in epigenetic landscapes are known to impact genome accessibility and transcription. Extensive evidence demonstrates that senescent cells undergo significant changes in chromatin structure; however, the mechanisms underlying the crosstalk between epigenetic parameters and gene expression profiles have not been fully elucidated. In the present study, we delineate the genome-wide redistribution of accessible chromatin regions that lead to broad transcriptome effects during senescence. We report that distinct senescence-activated accessibility regions (SAAs) are always distributed in H3K27ac-occupied enhancer regions, where they are responsible for elevated flanking senescence-associated secretory phenotype (SASP) expression and aberrant cellular signaling relevant to SASP secretion. Mechanistically, a single transcription factor, TEAD4, moves away from H3K27ac-labled SAAs to allow for prominent chromatin accessibility reconstruction during senescence. The enhanced SAAs signal driven by TEAD4 suppression subsequently induces a robust increase in the expression of adjacent SASP genes and the secretion of downstream factors, which contribute to the progression of senescence. Our findings illustrate a dynamic landscape of chromatin accessibility following senescence entry, and further reveal an insightful function for TEAD4 in regulating the broad chromatin state that modulates the overall transcriptional program of SASP genes.

Effect of Side-Chain Functional Groups in the Immunogenicity of Bacterial Surface Glycans.

Glycans on the surface of bacteria have diverse and essential biological functions and have widely been employed for treating various bacterial infectious diseases. Furthermore, these glycans comprise various functional groups, such as O-, N-, and carboxyl-modified, which significantly increase the diversity of glycan structures. These functional groups are not only crucial for glycans' structural identity but are also essential for their biological functions. Therefore, a clear understanding of the biological functions of these modified groups in corresponding bacterial glycans is crucial for their medical applications. Thus far, the activities of functional groups in some biomedical active carbohydrates have been elucidated. It has been shown that some functional groups are key constituents of biologically active bacterial glycans, while others are actually not essential and may even mask the functions of the glycans. This paper reviews the structures of naturally occurring side-chain functional groups in glycans located on the bacterial surface and their roles in immunological responses.

Chemical Synthesis of the Highly Sterically Hindered Core Undecasaccharide of Helicobacter pylori Lipopolysaccharide for Antigenicity Evaluation with Human Serum.

Helicobacter pylori, listed as a human carcinogen by the Department of Health and Human Services, colonizes the gastric mucosa of more than half of the world's population. The individuals infected with H. pylori have a high risk to develop chronic gastritis, peptic ulcers, and even gastric cancer. The conserved core structure of H. pylori lipopolysaccharide (LPS) has been regarded as a promising candidate structure for development of a glycoconjugate vaccine targeting multiple serotypes. Here, we report a total synthesis of the core undecasaccharide of H. pylori LPS and its subunit antigens. The match and mismatch between the glycosyl donor and acceptor caused by the inert hydroxyl groups were addressed by a judicious choice of orthogonal protection strategies and glycosylation conditions. A combination of acyl remote participation and solvent effects has been applied for selective formation of the five 1,2-cis-glucosidic bonds. The high steric hindrance induced by the high carbon sugars and trinacriform architecture required that the core undecasaccharide was synthesized through a finely tuned linear assembly [2 + (1 + (3 + (1 + (1 + 3))))] rather than convergent strategies. An antigenicity evaluation using glycan microarrays showed that an α-(1 → 6)-glucan trisaccharide is recognized by IgG antibodies in sera of H. pylori-infected patients. The phosphate group of the inner core trisaccharide key epitope is very important for IgG recognition. These findings are an important step toward designing carbohydrate-based vaccines against H. pylori.

Granatripodins A-B, limonoids featuring a Tricyclo[3.3.1.02,8]nonane motif: Absolute configuration and agonistic effects on human pregnane-X-receptor.

Two unprecedented limonoids incorporating a sterically encumbered cyclopropane ring, named granatripodins A (1) and B (2), featuring the presence of a tricyclo[3.3.1.02,8]nonane motif, were obtained from seeds of the Thai Xylocarpus granatum. The planar structures and absolute configurations of these limonoids were unambiguously established by NMR investigations, TDDFT-ECD and DFT-NMR calculations, and single-crystal X-ray diffraction analysis (Cu Kα). Most notably, granatripodin A (1) exhibited agonistic effects on human pregnane-X-receptor at the concentration of 100.0 nM. The biosynthetic origins of these limonoids via a radical cascade reaction are proposed. This study exemplifies a universal approach for the stereochemical assignment of polycyclic compounds with a cyclopropane-embedded cage scaffold.

Discovery of Thai mangrove tetranortriterpenoids as agonists of human pregnane-X-receptor and inhibitors against human carboxylesterase 2.

Human pregnane-X-receptor (hPXR) is considered to be the key target for the treatment of cholestasis and liver injury. Agonists of hPXR are potential drug leads. Potent and selective inhibitors of human carboxylesterase 2 (hCES2) could be utilized to alleviate the toxicity induced by ester drugs. In this work, fifteen new tetranortriterpenoids with structure diversity, named thaigranatins F-T (1-15), including four limonoids containing a C1-O-C29 bridge (1-4), four mexicanolides (5-8), three phragmalins (9-11), two limonoids belonging to the small group of trichiliton A (12-13), and two apotirucallanes (14-15), were isolated from seeds of the Thai mangrove, Xylocarpus granatum. The structures of these compounds were established by high resolution-electrospray ionization mass spectroscopy, extensive NMR spectroscopic investigations, single-crystal X-ray diffraction analyses, and the comparison of experimental electronic circular dichroism spectra. Most notably, thaigranatins L (7) and P (11) exhibited agonistic effects on hPXR at the concentration of 10.0 µM and 10.0 nM, respectively, whereas thaigranatins J (5), M (8), and T (15) showed inhibitory activities against hCES2 with IC50 values of 6.63, 11.35, and 5.05 µM, respectively. The 8α,30α-epoxy moiety of mexicanolide and the Δ8,14 double bond of phragmalin are pivotal for agonistic effects of these limonoids on hPXR, whereas the 6-OAc group of mexicanolide is crucial for its inhibitory activity against hCES2. Additionally, the flexible C-17-side-chain with appropriate hydroxy groups is considered to be important for the inhibitory activity of apotirucallane against hCES2.

Inhibition of the glutathione biosynthetic pathway increases phytochemical toxicity to Spodoptera litura and Nilaparvata lugens.

Phytochemicals are toxic to insects, but their insecticidal efficiencies are usually low compared to synthetic insecticides. Understanding the mechanism of insect adaptation to phytochemicals will provide guidance for increasing their efficacy. Reduced glutathione (GSH) is a scavenger of reactive oxygen species (ROS) induced by phytochemicals. However, in insects, the pathway of GSH biosynthesis in response to phytochemicals is unclear. We found that exposure to 0.5% indole-3-methanol (I3C), xanthotoxin, and rotenone (ROT) significantly retarded the growth of Spodoptera litura larvae. The oxidative stress in S. litura larvae exposed to phytochemicals was increased. The up-regulation of glutamate cysteine ligase but not glutathione reductase revealed that the de novo synthesis pathway is responsible for GSH synthesis in phytochemical-treated larvae. Treatment with the inhibitor (BSO) of γ-glutamylcysteine synthetase (gclc), a subunit of glutamate cysteine ligase, resulted in decreases of GSH levels and GST activities, increases of ROS levels in I3C-treated larvae, which finally caused midgut necrosis and larval death. Treatment with BSO or I3C alone did not cause larval death. The addition of GSH could partly reduce the influence of I3C and BSO on S. litura growth. Nilaparvata lugens gclc RNAi confirmed the result of BSO treatment in S. litura. N. lugens gclc RNAi significantly increased the mortality of ROT-sprayed N. lugens, in which ROS levels were significantly increased. All data indicate that gclc is involved in insect response to phytochemical treatment. Treatment with dsgclc will increase the insecticidal efficacy of plant-derived compounds.

Chemical Synthesis Elucidates the Key Antigenic Epitope of the Autism-Related Bacterium Clostridium bolteae Capsular Octadecasaccharide.

The gut pathogen Clostridium bolteae has been associated with the onset of autism spectrum disorder (ASD). To create vaccines against C. bolteae, it is important to identify exact protective epitopes of the immunologically active capsular polysaccharide (CPS). Here, a series of C. bolteae CPS glycans, up to an octadecasaccharide, was prepared. Key to achieving the total syntheses is a [2+2] coupling strategy based on a ß-d-Rhap-(1→3)-α-d-Manp repeating unit that in turn was accessed by a stereoselective ß-d-rhamnosylation. The 4,6-O-benzylidene-induced conformational locking is a powerful strategy for forming a ß-d-mannose-type glycoside. An indirect strategy based on C2 epimerization of ß-d-quinovoside was efficiently achieved by Swern oxidation and borohydride reduction. Sequential glycosylation, and regioselective and global deprotection produced the disaccharide and tetrasaccharide, up to the octadecasaccharide. Glycan microarray analysis of sera from rabbits immunized with inactivated C. bolteae bacteria revealed a humoral immune response to the di- and tetrasaccharide, but none of the longer sequences. The tetrasaccharide may be a key motif for designing glycoconjugate vaccines against C. bolteae.

Chemical Synthesis and Immunological Evaluation of Helicobacter pylori Serotype O6 Tridecasaccharide O-Antigen Containing a dd-Heptoglycan.

The development of glycoconjugate vaccines against Helicobacter pylori is challenging. An exact epitope of the H. pylori lipo-polysaccharide (LPS) O-antigens that contain Lewis determinant oligosaccharides and unique dd-heptoglycans has not yet been identified. Reported here is the first total synthesis of H. pylori serotype O6 tridecasaccharide O-antigen containing a terminal Ley tetrasaccharide, a unique α-(1→3)-, α-(1→6)-, and α-(1→2)-linked heptoglycan, and a ß-d-galactose connector, by an [(2×1)+(3+8)] assembly sequence. Seven oligosaccharides covering different portions of the entire O-antigen were prepared for immunological investigations with a particular focus on elucidation of the roles of the dd-heptoglycan and Ley tetrasaccharide. Glycan microarray analysis of sera from rabbits immunized with isolated serotype O6 LPS revealed a humoral immune response to the α-(1→3)-linked heptoglycan, a key motif for designing glycoconjugate vaccines for H. pylori serotype O6.

Total Synthesis of a Densely Functionalized Plesiomonas shigelloides Serotype 51 Aminoglycoside Trisaccharide Antigen.

Plesiomonas shigelloides, a pathogen responsible for frequent outbreaks of severe travelers' diarrhea, causes grave extraintestinal infections. Sepsis and meningitis due to P. shigelloides are associated with a high mortality rate as antibiotic resistance increases and vaccines are not available. Carbohydrate antigens expressed by pathogens are often structurally unique and are targets for developing vaccines and diagnostics. Here, we report a total synthesis of the highly functionalized trisaccharide repeating unit 2 from P. shigelloides serotype 51 from three monosaccharides. A judicious choice of building blocks and reaction conditions allowed for the four amino groups adorning the sugar rings to be installed with two N-acetyl (Ac) groups, rare acetamidino (Am), and d-3-hydroxybutyryl (Hb) groups. The strategy for the differentiation of amino groups in trisaccharide 2 will serve well for the syntheses of other complex glycans.

Synergistic Glycosylation as Key to the Chemical Synthesis of an Outer Core Octasaccharide of Helicobacter pylori.

Helicobacter pylori, a widespread gastric bacterial pathogen that infects 90 % of the population in developing countries, causes chronic gastritis, peptic ulcers and gastric cancer. Battling H. pylori infection is a serious challenge due to the increased resistance to antibiotics and the lack of vaccines. The lipopolysaccharide covering the H. pylori cell-surface outer membrane is an attractive target for the development of a glycoconjugate vaccine. Here, we report a [3+5] convergent synthesis of an outer core octasaccharide of H. pylori employing just three orthogonally protected building blocks. A synergistic glycosylation strategy enables the creation of five pivotal 1,2-cis-α-glucosidic bonds consist of four types of linkages using just three monomers. This strategy can be expanded to many 1,2-cis-α-gluoside-containing oligosaccharides both in solution and solid phase.

Limonoids Containing a C1⁻O⁻C29 Moiety: Isolation, Structural Modification, and Antiviral Activity.

Five new limonoids named thaigranatins A⁻E (1⁻5), containing a C1⁻O⁻C29 moiety, were isolated from seeds of the Thai Xylocarpus granatum, collected at the mangrove swamp of Trang Province, together with the known limonoid, granatumin L (6). The structures of these compounds were established by HR-ESIMS and extensive NMR spectroscopic data. The absolute configuration of 1 was unequivocally determined by single-crystal X-ray diffraction analysis, conducted with Cu Kα radiation; whereas that of 2 or 6 was established to be the same as that of 1 by the similarity of their electronic circular dichroism (ECD) spectra. In view of the marked antiviral activity of 6, its structure was modified via hydrolysis with alkaline KOH, esterification with diazomethane and various organic acids, and oximization with hydroxyamine. Finally, 18 derivatives, viz. 7⁻10, 8a⁻8i, 9a⁻9b, and 10a⁻10c, were obtained. In vitro antiviral activities of these derivatives against human immunodeficiency virus 1 (HIV-1) and influenza A virus (IAV) were evaluated. Most notably, 8i exhibited marked inhibitory activity against HIV-1 with an IC50 value of 15.98 ± 6.87 µM and a CC50 value greater than 100.0 µM; whereas 10b showed significant inhibitory activity against IAV with an IC50 value of 14.02 ± 3.54 µM and a CC50 value greater than 100.0 µM.

Recent chemical synthesis and immunological evaluation of glycans related to bacterial lipopolysaccharides.

O-Antigens and core oligosaccharides from bacterial lipopolysaccharides (LPS) are often structurally unique and immunologically active, have become attractive targets in the development of antibacterial vaccines. Structurally well-defined and pure oligosaccharides can be used in identifying protective epitopes of the carbohydrate antigens, which is important for the design of an effective vaccine. Here, the recent progress on chemical synthesis and immunological evaluation of glycans related to O-antigens and core oligosaccharides from bacterial LPS are summarized.

Chemical Synthesis of Conjugation-Ready Trisaccharides Corresponding to Biological Repeating Units of Pseudomonas aeruginosa Serotype 10 and 19 O-Antigens.

Here we report the chemical synthesis of conjugation-ready trisaccharides, representing biological repeating units of Pseudomonas aeruginosa serotype 10 and 19 O-antigens. The α-d-QuiN3 glycosidic bond was stereoselectively synthesized through TMSI─Ph3P═O mediated 1,2-cis glycosylation. Selective oxidation of the C6-OH group at the disaccharide stage allowed for benzylidene-promoted construction of the α-l-GalN3 glycosidic bond and simplification of the postglycosylation process at the trisaccharide stage. The low reaction temperature and neighboring electron-donating effect facilitated the efficient synthesis of the trisaccharide.

Chemical Synthesis of the Trisaccharide Repeating Unit of the O-Antigen of Fusobacterium nucleatum ATCC 51191.

Herein, the trisaccharide repeating unit of Fusobacterium nucleatum ssp. animalis ATCC 51191, which is used to develop oncomicrobial vaccines, was efficiently synthesized for the first time. The synthetic approach featured the following: (i) construction of the 1,2-cis-glycosidic linkage using the large steric hindrance of a phthalimide group at C4 of fucosamine; (ii) synthesis of the trisaccharide via a linear [2 + 1] glycosylation strategy; and (iii) installation of l-alanine using hexafluorophosphate azabenzotriazole tetramethyl uronium as a promoter.

Chemical Synthesis of a Densely Functionalized Aminodisaccharide from Fusobacterium nucleatum ATCC 23726 O-Antigen.

Fusobacterium nucleatum, a colorectal-cancer-associated oncomicrobe, can trigger or accelerate numerous pathologies. We report the first synthesis of a conjugation-ready disaccharide containing six amino groups from F. nucleatum ATCC 23726 O-antigen. Rare 2,3-diamido-d-glucuronic acid amide and 2-acetamido-4-amino-d-fucose were synthesized from d-glucosamine through configuration inversion, nucleophilic substitution, C6 oxidation, and C6 deoxygenation. A judicious choice of protecting groups and reaction conditions enabled the selective installation of N-acetyl, N-propanoyl, N-formyl, and carboxamido groups.

Chemical Synthesis of an Orthogonally Protected 5,7-Diamino-3,5,7,9-tetradeoxy-d-glycero-l-gluco-2-nonulosonic Acid from N-Acetylneuraminic Acid.

Bacterial nonulosonic acids (NulOs), which feature a nine-carbon backbone, are associated with the biological functions of bacterial glycans. Here, an orthogonally protected 5-amino-7-azido-3,5,7,9-tetradeoxy-d-glycero-l-gluco-2-nonulosonic acid related to Fusobacterium nucleatum ATCC 23726 NulO was synthesized from N-acetylneuraminic acid with sequential performance of C5,7 azidation, C9 deoxygenation, C4 epimerization, and N5,7 differentiation. The C5 azido group in the obtained 5,7-diazido-NulO can be regioselectively reduced to differentiate the two amino groups.

Association of MTHFR gene polymorphisms with non-Hodgkin lymphoma risk: Evidence from 31 articles.

Background: Methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms, particularly C677T and A1298C, have been implicated in various cancers, including non-Hodgkin lymphoma (NHL); however, their association with NHL risk remains inconclusive. Methods: We conducted an updated meta-analysis to assess the relationship between MTHFR gene polymorphisms (C677T and A1298C) and NHL risk. Relevant studies were identified through systematic literature searches in multiple databases. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to evaluate the strength of the associations. Results: The meta-analysis included 32 studies (8222 cases vs. 12956 controls) for MTHFR C677T and 26 studies (6930 cases vs. 11611 controls) for the A1298C polymorphism. Our meta-analysis revealed no significant associations between MTHFR gene polymorphisms (C677T and A1298C) and NHL risk. However, subgroup analysis stratified by ethnicity and NHL subtype yielded interesting findings for the C677T polymorphism. Specifically, in the subgroup analysis of Caucasians, the C677T polymorphism was significantly associated with NHL risk (heterozygous: OR=1.16, 95% CI=1.02-1.32; allele comparison: OR=1.07, 95% CI=1.01-1.13). Furthermore, in the analysis stratified by NHL subtype, the C677T polymorphism was significantly associated with increased follicular lymphoma (FL) risk (homozygous: OR=1.25, 95% CI=1.02-1.53; recessive: OR=1.28, 95% CI=1.06-1.56). False-positive result possibility (FPRP) analysis verified that the association of the MTHFR C677T polymorphism with NHL risk for Caucasians and FL subtypes was a true positive and deserves attention. We also determined that the C677T polymorphism is an expression quantitative trait locus (eQTL) since it is associated with MTHFR gene expression. Conclusion: There was no overall association between MTHFR gene polymorphisms (C677T and A1298C) and NHL risk, but stratified analyses revealed significant associations in specific subgroups. While meta-analyses inherently build upon existing studies, our work distinguishes itself by incorporating recent data, applying rigorous analytical techniques, and providing more evidence of the MTHFR C677T polymorphism as an eQTL.

The TIAR-mediated Nrf2 response to oxidative stress is mediated through the Nrf2 noncoding 3'untranslated region in Spodoptera litura.

Nrf2 is a key regulator in the maintenance of cellular redox balance by regulating the expression of genes related to antioxidative responses and detoxification. Nrf2 protein levels are increased in response to oxidative stress. However, the regulation of the Nrf2 3'UTR on Nrf2 translation is unclear. Here, we report that the translational activity of the 3'UTR is required for Spodoptera litura Nrf2 protein expression. Experiments showed that the 3'UTR translation activity of S. litura Nrf2 was much higher than that of the 5'UTR. RNA interference (RNAi) of the expression of T cell internal antigen-related protein (TIAR), an RNA-binding protein that interacts with the 3'UTR of S. litura Nrf2, resulted in Nrf2 mRNA movement out of translationally active polysomes and a decrease in cellular Nrf2 protein levels. TIAR interacted with poly(A)-binding protein (PABP) and translation initiation factors eIF2-2 and eIF2-3 to enhance Nrf2 translation, indicating that the 3'UTR regulates Nrf2 translation. Diethyl maleate (DEM) treatment increased reactive oxygen species (ROS) in cells and enhanced Nrf2 levels, which had been reduced by cycloheximide (CHX), an inhibitor of de novo protein synthesis; Tiar RNAi increased ROS levels in DEM-treated cells, suggesting TIAR-mediated 3'UTR involvement in Nrf2 translation in response to DEM treatment. Thus, we reveal a posttranscriptional regulation mechanism of Nrf2, in which TIAR binds with the Nrf2 mRNA 3'UTR to enhance Nrf2 translation, facilitating the increase in Nrf2 protein levels in response to oxidative stress.