Design and Synthesis of Cyclic Dinucleotide Analogues Containing Triazolyl C-Nucleosides.

Natural cyclic dinucleotide (CDN) is the secondary messenger involved in bacterial hemostasis, human innate immunity, and bacterial antiphage immunity. Synthetic CDN and its analogues are key molecular probes and potential immunotherapeutic agents. Several CDN analogues are under clinical research for antitumor immunotherapy. A myriad of synthetic methods have been developed and reported for the preparation of CDN and its analogues. However, most of the protocols require multiple steps, and only one CDN or its analogue is prepared at a time. In this study, a strategy based on a macrocyclic ribose phosphate skeleton containing a 1'-alkynyl group was designed and developed to prepare CDN analogues containing triazolyl C-nucleosides by click chemistry. Combinatorial application of click chemistry and the sulfenylation cascade to the macrocyclic skeleton expanded the diversity of the CDN analogues. This macrocyclic skeleton strategy rapidly and efficiently provides CDN analogues to facilitate research on microbiology, immunology, and immunotherapy.

Unraveling the Binding Mode of Cyclic Adenosine-Inosine Monophosphate (cAIMP) to STING through Molecular Dynamics Simulations.

The stimulator of interferon genes (STING) plays a significant role in immune defense and protection against tumor proliferation. Many cyclic dinucleotide (CDN) analogues have been reported to regulate its activity, but the dynamic process involved when the ligands activate STING remains unclear. In this work, all-atom molecular dynamics simulations were performed to explore the binding mode between human STING (hSTING) and four cyclic adenosine-inosine monophosphate analogs (cAIMPs), as well as 2',3'-cGMP-AMP (2',3'-cGAMP). The results indicate that these cAIMPs adopt a U-shaped configuration within the binding pocket, forming extensive non-covalent interaction networks with hSTING. These interactions play a significant role in augmenting the binding, particularly in interactions with Tyr167, Arg238, Thr263, and Thr267. Additionally, the presence of hydrophobic interactions between the ligand and the receptor further contributes to the overall stability of the binding. In this work, the conformational changes in hSTING upon binding these cAIMPs were also studied and a significant tendency for hSTING to shift from open to closed state was observed after binding some of the cAIMP ligands.

Co-, N-doped carbon dot nanozymes based on an untriggered ROS generation approach for anti-biofilm activities and in vivo anti-bacterial treatment.

Bacterial infections originating from food, water, and soil are widely recognized as significant global public health concerns. Biofilms are implicated in approximately two-thirds of bacterial infections. In recent times, nanomaterials have emerged as potential agents for combating biofilms and bacteria, with many of them being activated by light and H2O2 to generate reactive oxygen species (ROS). However, this energy-consuming and extrinsic substrate pattern poses many challenges for practical application. Consequently, there is a pressing need to develop methods for the untriggered generation of ROS to effectively address biofilm and bacterial infections. In this study, we investigated the oxidase-like activity of the Co,N-doped carbon dot (CoNCD) nanozyme, which facilitated the oxidation of ambient O2 to generate 1O2 in the absence of light and H2O2 supplementation; this resulted in effective biofilm cleavage and enhanced bactericidal effects. CoNCDs could become a potential candidate for wound healing and treatment of acute peritonitis in vivo, which can be primarily attributed to the spontaneous production of ROS. This study presents a convenient ROS generator that does not necessitate any specific triggering conditions. The nanozyme properties of CoNCDs exhibit significant promise as a potential remedy for diseases, specifically as an anti-biofilm and anti-bacterial agent.

A novel self-assembled nucleobase-nanofiber platform of CDN to activate the STING pathway for synergistic cancer immunotherapy.

2', 3'-cGAMP (CDN) as cGAS-STING pathway agonist is extensively used in tumor treatment. However, due to its negatively charged nature (containing two phosphate groups) and high hydrophilicity, CDN faces challenges in crossing cell membranes, resulting in reduced efficiency of its use. Additionally, CDN is susceptible to inactivation through phosphodiesterase hydrolysis. Therefore, the development of a new drug delivery system for CDN is necessary to prevent hydrolysis and enhance targeted accumulation in tumors, as well as improve cellular uptake for STING activation. In this study, we have developed peptide-polymer nanofibers (PEG-Q11) that incorporate thymine (T) and arginine (R) residues to facilitate complexation with CDN through the principles of Watson-Crick base pairing with thymine and favorable electrostatic interactions and bidentate hydrogen bonding with arginine side chains. The entrapment efficiency (EE) of PEG-Q11T3R4@CDN was found to be 51% higher than that of PEG-Q11@CDN. Due to its favorable biocompatibility, PEG-Q11T3R4@CDN was employed for immunotherapy in mouse CT26 tumors. In local tumor treatment, the administration of PEG-Q11T3R4@CDN at a low dose and through a single injection exhibited inhibitory effects. Furthermore, the local injection of PEG-Q11T3R4@CDN resulted in systemic therapeutic responses, effectively suppressing tumor metastasis by activating CD8 + T cells to target distant tumors. This research not only underscores the potential of PEG-Q11T3R4@CDN as an efficient therapeutic agent but also highlights its ability to achieve long-lasting systemic therapeutic outcomes following local treatment. Consequently, PEG-Q11T3R4@CDN represents a promising strategy for immunization.

Remodeling of Tumor Microenvironment by Nanozyme Combined cGAS-STING Signaling Pathway Agonist for Enhancing Cancer Immunotherapy.

Nanozymes and cyclic GMP-AMP synthase (cGAS) the stimulator of interferon genes (STING) signaling pathway, as powerful organons, can remodel the tumor microenvironment (TME) to increase efficacy and overcome drug resistance in cancer immunotherapy. Nanozymes have the potential to manipulate the TME by producing reactive oxygen species (ROS), which lead to positive oxidative stress in tumor cells. Cyclic dinucleotide (2',3'-cGAMP), as a second messenger, exists in the TME and can regulate it to achieve antitumor activity. In this work, Co,N-doped carbon dots (CoNCDs) were used as a model nanozyme to evaluate the properties of the anti-tumor mechanism, and effective inhibition of S180 tumor was achieved. Based on CoNCDs' good biocompatibility and therapeutic effect on the tumor, we then introduced the cGAS-STING agonist, and the combination of the CoNCDs and STING agonist significantly inhibited tumor growth, and no significant systemic toxicity was observed. The combined system achieved the enhanced tumor synergistic immunotherapy through TME reprogramming via the peroxidase-like activity of the CoNCDs and cGAS-STING signaling pathway agonist synergistically. Our work provides not only a new effective way to reprogram TME in vivo, but also a promising synergic antitumor therapy strategy.

Synthetic K+ Channels Constructed by Rebuilding the Core Modules of Natural K+ Channels in an Artificial System.

Different types of natural K+ channels share similar core modules and cation permeability characteristics. In this study, we have developed novel artificial K+ channels by rebuilding the core modules of natural K+ channels in artificial systems. All the channels displayed high selectivity for K+ over Na+ and exhibited a selectivity sequence of K+ ≈Rb+ during the transport process, which is highly consistent with the cation permeability characteristics of natural K+ channels. More importantly, these artificial channels could be efficiently inserted into cell membranes and mediate the transmembrane transport of K+ , disrupting the cellular K+ homeostasis and eventually triggering the apoptosis of cells. These findings demonstrate that, by rebuilding the core modules of natural K+ channels in artificial systems, the structures, transport behaviors, and physiological functions of natural K+ channels can be mimicked in synthetic channels.

Fluoro-functionalized ionic covalent organic frameworks (F-iCOFs) for highly selective enrichment and sensitive determination of perfluorinated sulfonates by MALDI-MS.

An easily prepared fluoro-functionalized ionic covalent organic framework (F-iCOF) has been implemented into MALDI-TOF MS, enabling the highly selective enrichment and sensitive determination of perfluorinated sulfonate (PFS) contaminants in a rapid and convenient manner. The good thermal stability and excellent optical absorption properties of F-iCOF makes it a brilliant matrix with no background noise. Moreover, benefitting from the large surface area, appropriate pore size, good water dispersibility, and abundant fluorine atom and cationic characteristic of F-iCOF, it exhibited superior adsorption capacity and enrichment selectivity towards PFSs. Good signal responses for PFSs were obtained in the presence of various interfering compounds such as BSA, HA, or even more than 100-fold excess of glutamic acid and similar in structure sodium alkyl sulfonates, highlighting the specific selectivity of F-iCOF. Calibration curves for potassium perflurobutane sulfonate (PFBSK) in tap water and whole blood were established with good linear correlation in the range 1-500 pg mL-1. The limits of detection and quantification for PFBSK were as low as 0.04 pg mL-1 and 0.05 pg mL-1, respectively, which are comparable or better than the existing methods for the determination of PFSs.

High-throughput profiling volatiles in edible oils by cooling assisted solid-phase microextraction technique for sensitive discrimination of edible oils adulteration.

In this study, a cooling assisted solid-phase microextraction technique (CA-SPME) was proposed and used for identifying volatile and semi-volatile compounds in edible oil innovatively coupled to gas chromatography-mass spectrometry. Compared with regular SPME technique, CA-SPME presented significantly higher extraction efficiencies for analytes in edible oil due to its synergistic effect of heating and cooling. After optimization of the extraction conditions including heating temperature, cooling temperature, extraction time, and added amount of edible oil, thirty-eight, thirty-six, twenty-nine, and thirty-three kinds of compounds in peanut oil, olive oil, canola oil, and soybean oil were successfully identified, respectively, using DVB/CAR/PDMS coating with extraction time of 30 min and edible oil amounts of 20 µL. Principal component analysis, partial least squares discriminant analysis, and hierarchical clustering analysis (HCA) were performed to evaluate the potential of proposed method in discriminating edible oils adulteration (peanut oil adulterated with canola oil, peanut oil adulterated with soybean oil, olive oil adulterated with canola oil) subsequently. Results demonstrated that the method was useful in successful discrimination of pure and adulterated edible oils with adulteration percentages ranging from 0.5 to 10%. Furthermore, volatiles contributing to classifications between pure and adulterated edible oils were also illustrated based on variable importance for the projection analysis and distributions of volatiles in HCA heatmaps. The proposed method provided a novel strategy for sensitive detection of edible oil adulteration without any other sample pretreatment.

Lotus-like Ni@NiO nanoparticles embedded porous carbon derived from MOF-74/cellulose nanocrystal hybrids as solid phase microextraction coating for ultrasensitive determination of chlorobenzenes from water.

Lotus-like Ni@NiO embedded porous carbons (Ni@NiO/PCs) were fabricated by pyrolysis of MOF-74/cellulose nanocrystal hybrids, and used as a solid phase microextraction (SPME) coating for ultrasensitive determination of chlorobenzenes (CBs) from water combined with gas chromatography-mass spectrometry. Owing to its abundant chemical groups, high porosity, and excellent thermal stability, the as-prepared Ni@NiO/PCs presented superior extraction performance compared to commercial SPME coatings. Notably, Ni@NiO/PCs derived from MOF-74/CNC hybrids presented higher extraction efficiencies towards CBs than that derived from pristine CNC and MOF-74 due to the formation of micro/mesopores and more abundant oxygen-containing groups. Under the optimum extraction conditions, the proposed analytical method presented wide linearity range (0.5-1500 ng L-1), ultra-low detection of limit (0.005-0.049 ng L-1), and excellent precision with relative standard deviations of 4.7-9.2% for a single fiber and 8.8-10.9% for 5 fibers, and long lifetime (≥160 times). The proposed analytical method was finally applied for determination of CBs from real water samples, and the recoveries were in the range of 93.2-116.8% towards eight CBs. This study delivered a novel and efficient sorbent as SPME coating to extraction and determination of CBs from water.

Carbon Dot-Decorated Graphite Carbon Nitride Composites for Enhanced Solid-Phase Microextraction of Chlorobenzenes from Water.

In this work, carbon dot-decorated graphite carbon nitride composites (CDs/g-C3N4) were synthesized and innovatively used as a SPME coating for the sensitive determination of chlorobenzenes (CBs) from water samples, coupled with gas chromatography-mass spectrometry. The CDs/g-C3N4 coating presented superior extraction performance in comparison to pristine g-C3N4, owing to the enhancement of active groups by CDs. The extraction capacities of as-prepared SPME coatings are higher than those of commercial coatings due to the functions of nitrogen-containing and oxygen-containing group binding, π-π stacking, and hydrophobic interactions. Under optimized conditions, the proposed method exhibits a wide linearity range (0.25-2500 ng L-1), extremely low detection of limits (0.002-0.086 ng L-1), and excellent precision, with relative standard deviations of 5.3-9.7% for a single fiber and 7.5-12.6% for five fibers. Finally, the proposed method was successfully applied for the analysis of CBs from real river water samples, with spiked recoveries ranging from 73.4 to 109.1%. This study developed a novel and efficient SPME coating material for extracting organic pollutants from environmental samples.

Core-shell structured Fe2O3/CeO2@MnO2 microspheres with abundant surface oxygen for sensitive solid-phase microextraction of polycyclic aromatic hydrocarbons from water.

Core-shell structured Fe2O3/CeO2@MnO2 microspheres were fabricated and used as solid-phase microextraction coating for determination of polycyclic aromatic hydrocarbons (PAHs) in water samples. XPS spectra demonstrated the generation of abundant surface oxygen on Fe2O3/CeO2@MnO2 microspheres, which provided binding sites for enhancement of analyte extraction. Under optimized conditions, the proposed method presented good linearity in the concentration range 0.04-100 ng mL-1, with low limits of detection varying from 0.38 to 3.57 ng L-1 for eight PAHs. Relative standard deviations for a single fiber and five batches of fibers were in the ranges of 4.1-8.2% and 7.1-11.4%, respectively. The proposed method was successfully used for determination of PAHs in real river water samples with recoveries ranging from 87.1 to 115.9%. The proposed method using as-prepared Fe2O3/CeO2@MnO2 microspheres as SPME coating exhibit significant potential for real sample analysis due to its excellent reproducibility, high sensitivity, and good linearity.

Fullerenol as a water-soluble MALDI-MS matrix for rapid analysis of small molecules and efficient quantification of saccharin sodium in foods.

Due to the strong background interferences in the low-mass region and poor reproducibility of conventional organic matrices, it is of great importance to develop a novel matrix for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to qualitatively and quantitatively analyze small molecules. In this work, water-soluble fullerenol C60(OH)24-26 was selected as a MALDI matrix for the analysis of low-molecular-weight compounds in consideration of optical absorption property, water solubility and stability. Compared with the traditional matrices, fullerenol demonstrated lower background interference and stronger peak intensity. In addition, the hydrophilic fullerenol could avoid the heterogeneous crystallization in sample preparation, increase the reproducibility and sensitivity of MALDI-MS, and ameliorate quantitative analysis of small molecules. With saccharin as model analyte, quantitative analysis was carried out using fullerenol as matrix. The results demonstrated satisfying reproducibility and good tolerance to salt. The limit-of-detection of the quantitative analysis was as low as 4 pmol, and the linear range is 1-100 µg mL-1 with R2 greater than 0.99. The analytical results also showed excellent precision and accuracy, low matrix effect and good recovery rate. Fullerenol as a potential matrix was further validated in the quantification of saccharin sodium in different real food samples, such as nuts and drinks. This work not only confirms the potential of fullerenol for the quantitative analysis in food field, but also provides a new technique for rapid analysis of small molecules.

Effect of charge status on the ion transport and antimicrobial activity of synthetic channels.

A class of unimolecular channels formed by pillararene-gramicidin hybrid molecules are presented. The charge status of the peptide domain in these channels has a significant impact on their ion transport and antimicrobial activity. These channels exhibited different membrane-association abilities between microbial cells and mammalian cells. One of the channels displayed a higher antimicrobial activity towards S. aureus (IC50 = 0.55 µM) and negligible hemolytic toxicity, showing potential to serve as a systemic antibiotic.

Three-dimensional pompon-like Au/ZnO porous microspheres as solid phase microextraction coating for determination of volatile fatty acids from foot odor.

In this study, facile fabrication of three-dimensional (3D) pompon-like gold/zinc oxide (Au/ZnO) porous microspheres by hydrothermal procedure was investigated. These microspheres were utilized as solid phase microextraction (SPME) coating for determination of volatile fatty acids (VFAs) from foot odor coupling with gas chromatography-mass spectrometry (GC-MS). SEM and TEM characterizations showed that as-prepared material was composed of 1D porous nanowires and presented a uniform coating on stainless-steel wire. The extraction of VFAs including propanoic acid, butyric acid, isobutanoic acid, isovaleric acid, hexanoic acid, and heptylic acid was carried out by headspace model after sampling from human foot using cotton wool strips. Following optimization of extraction parameters including extraction temperature and time and desorption temperature and time, the as-prepared SPME coating presented better extraction efficiency than commercial DVB/CAR/PDMS fiber towards all the VFAs due to its excellent properties. Under the optimized conditions, the method exhibited good linearity (0.5-200 ng) with regression coefficients (R2) ranging from 0.9836 to 0.9981 for all the analytes. The limits of detection ranged from 0.017 to 0.098 ng. Single fiber repeatability varied from 6.5% to 11.2% and the fiber-to-fiber reproducibility ranged from 8.6% to 12.3%. The proposed method was successfully applied for extraction and determination of VFAs from foot odor after sampling using cotton wool strips.

Light-activated "cycle-reversible intramolecular charge transfer" fluorescent probe: monitoring of pHi trace change induced by UV light in programmed cell death.

Under the synergistic effects of protonation and deprotonation, a light-activated fluorescent probe (UV-SP) exhibited "cycle-reversible intramolecular charge transfer (ICT)" for different pH after activation by UV light, resulting in emission of multiple ratio fluorescent signals (FI563/FI595 and FI664/FI595). Based on these kinds of response signals, UV-SP can specifically monitor the cycle-reversible trace change of intracellular pH caused by UV radiation. More importantly, according to the stable and invariant multiple ratio fluorescent signals, UV-SP can sort cells entering programmed death.

Distinct Dynamic and Conformational Features of Human STING in Response to 2'3'-cGAMP and c-di-GMP.

The human stimulator of interferon genes protein (hSTING) can bind cyclic dinucleotides (CDNs) to activate the production of type I interferons and inflammatory cytokines. These CDNs can be either bacterial secondary messengers, 3'3'-CDNs, or endogenous 2'3'-cGAMP. cGAMP, with a unique 2'-5' bond, is the most potent activator of hSTING among all CDNs. However, current understanding of the molecular principles underlying the unique ability of 2'3'-cGAMP to potently activate hSTINGs other than 3'3'-CDNs remains incomplete. In this work, molecular dynamics simulations were used to provide an atomistic picture of the binding of 2'3'-cGAMP and one 3'3'-CDN (c-di-GMP) to hSTING. The results suggest that hSTING binds more strongly to 2'3'-cGAMP than to c-di-GMP, which prefers to bind with a more open and flexible state of hSTING. Finally, a potential "dock-lock-anchor" mechanism is proposed for the activation of hSTING upon the binding of a potent ligand. It is believed that deep insights into understanding the binding of hSTING with 3'3'-CDNs and the endogenous 2'3'-cGAMP would help to establish the principles underlying powerful 2'3'-cGAMP signaling and the nature of hSTING activation, as well as related drug design.

Artificial K+ Channels Formed by Pillararene-Cyclodextrin Hybrid Molecules: Tuning Cation Selectivity and Generating Membrane Potential.

A class of artificial K+ channels formed by pillararene-cyclodextrin hybrid molecules have been designed and synthesized. These channels efficiently inserted into lipid bilayers and displayed high selectivity for K+ over Na+ in fluorescence and electrophysiological experiments. The cation transport selectivity of the artificial channels is tunable by varying the length of the linkers between pillararene and cyclodexrin. The shortest channel showed specific transmembrane transport preference for K+ over all alkali metal ions (selective sequence: K+ > Cs+ > Rb+ > Na+ > Li+ ), and is rarely observed for artificial K+ channels. The high selectivity of this artificial channel for K+ over Na+ ensures specific transmembrane translocation of K+ , and generated stable membrane potential across lipid bilayers.

A unimolecular channel formed by dual helical peptide modified pillar[5]arene: correlating transmembrane transport properties with antimicrobial activity and haemolytic toxicity.

Five unimolecular channels with different lengths are presented. The varying length of these channels has significant impact on their transmembrane transport properties, which are directly correlated with their antimicrobial activity and inversely correlated with their haemolytic toxicity. By further structural optimization, these new channels could reach high antimicrobial activity and very low haemolytic toxicity, with the potential to serve as systemic antibiotics.

Covalent Attachment of Daptomycin to Ti6Al4V Alloy Surfaces by a Thioether Linkage to Inhibit Colonization by Staphylococcus aureus.

Infections are a devastating complication of titanium alloy orthopedic implants. Current therapies include antibiotic-impregnated bone cement and antibiotic-containing coatings. Daptomycin (DAP) (1) is a novel peptide antibiotic that penetrates the cell membranes of Gram-positive bacteria. Few DAP-resistant strains have appeared so far. We hypothesized that when DAP covalently bonded via a flexible, hydrophilic spacer it could prevent bacterial colonization of titanium alloy surfaces. We designed and synthesized a series of DAP conjugates for bonding to the surface of Ti6Al4V foils through tetra(ethylene glycol) spacers via thioether linkages. The stability and antimicrobial activity of the attached conjugates were evaluated using Staphylococcus aureus ATCC 25923. Colonization of the Ti6Al4V foils was inhibited by 72% at 8 h and 54% at 24 h. The strategy described in this report provides a new, more facile way to prepare bactericidal Ti6Al4V implants.

One-pot formation of hydrazide macrocycles with modified cavities: an example of pH-sensitive unimolecular cation channels.

A series of pH-sensitive, cation-selective hydrazide macrocyclic channels have been synthesized. The macrocyclic channels bear multiple carboxyls in the inner cavity, which have a significant impact on their membrane-incorporation ability and NH4+ transport activity. Moreover, the K+/Cl- selectivities of the macrocyclic channels can be tuned by the pH value of the electrolyte.