Genetically encoded bioorthogonal tryptophan decaging in living cells.

Tryptophan (Trp) plays a critical role in the regulation of protein structure, interactions and functions through its π system and indole N-H group. A generalizable method for blocking and rescuing Trp interactions would enable the gain-of-function manipulation of various Trp-containing proteins in vivo, but generating such a platform remains challenging. Here we develop a genetically encoded N1-vinyl-caged Trp capable of rapid and bioorthogonal decaging through an optimized inverse electron-demand Diels-Alder reaction, allowing site-specific activation of Trp on a protein of interest in living cells. This chemical activation of a genetically encoded caged-tryptophan (Trp-CAGE) strategy enables precise activation of the Trp of interest underlying diverse important molecular interactions. We demonstrate the utility of Trp-CAGE across various protein families, such as catalase-peroxidases and kinases, as translation initiators and posttranslational modification readers, allowing the modulation of epigenetic signalling in a temporally controlled manner. Coupled with computer-aided prediction, our strategy paves the way for bioorthogonal Trp activation on more than 28,000 candidate proteins within their native cellular settings.

Cancer-associated fibroblasts reprogram cysteine metabolism to increase tumor resistance to ferroptosis in pancreatic cancer.

Background: Pancreatic ductal adenocarcinoma (PDAC) is an insidious, rapidly progressing malignancy of the gastrointestinal tract. Due to its dense fibrous stroma and complex tumor microenvironment, neither of which is sensitive to radiotherapy, pancreatic adenocarcinoma is one of the malignancies with the poorest prognosis. Therefore, detailed elucidation of the inhibitory microenvironment of PDAC is essential for the development of novel therapeutic strategies. Methods: We analyzed the association between cancer-associated fibroblasts (CAFs) and resistance to ferroptosis in PDAC using conditioned CAF medium and co-culture of pancreatic cancer cells. Abnormal cysteine metabolism was observed in CAFs using non-targeted metabolomics analysis with liquid chromatography-tandem mass spectrometry (LC-MS/MS). The regulatory effects of cysteine were investigated in PDAC cells through measurement of cell cloning, cell death, cell function, and EdU assays. The effects of exogenous cysteine intake were examined in a mouse xenograft model and the effects of the cysteine pathway on ferroptosis in PDAC were investigated by western blotting, measurement of glutathione and reactive oxygen species levels, among others. Results: It was found that CAFs played a critical role in PDAC metabolism by secreting cysteine, which could increase tumor resistance to ferroptosis. A previously unrecognized function of the sulfur transfer pathway in CAFs was identified, which increased the extracellular supply of cysteine to support glutathione synthesis and thus inducing ferroptosis resistance. Cysteine secretion by CAFs was found to be mediated by the TGF-ß/SMAD3/ATF4 signaling axis. Conclusion: Taken together, the findings demonstrate a novel metabolic relationship between CAFs and cancer cells, in which cysteine generated by CAFs acts as a substrate in the prevention of oxidative damage in PDAC and thus suggests new therapeutic targets for PDAC.

Targeted Drug Delivery System Based on Copper Sulfide for Synergistic Near-Infrared Photothermal Therapy/Photodynamic Therapy/Chemotherapy of Triple Negative Breast Cancer.

Multi-modal synergistic therapy, especially the integration of near-infrared laser phototherapies and chemotherapy, is often sought after owing to its minimal invasiveness, low side effects, and improved anticancer therapeutic efficacy. Herein, CuS nanoparticles were first coated with zinc phthalocyanine derivant (Pc)-functionalized mesoporous silica (mSiO2-Pc) to achieve a drug delivery system (CuS@mSiO2-Pc) with photothermal/photodynamic therapy. Chemical drug DOX was subsequently loaded for chemotherapy, and hyaluronic acid (HA) was employed as a covering material with cancer targeting. The as-obtained CuS@mSiO2-Pc(DOX)@HA nanoparticles were nano-sized with good biocompatibility, effective DOX loading, and controllable DOX releasing. Expectedly, this multifunctional nanoplatform exhibits effective generation of reactive oxygen species and hyperthermia upon the near-infrared laser irradiation. Most importantly, the nanoparticles were targeted into 4T1 cells and showed significantly remarkable cytotoxicity under near-infrared laser irradiation, proving their synergistic therapeutic efficacy. Therefore, this targeted drug system based on CuS with synergistic photothermal therapy/photodynamic therapy/chemotherapy has great application prospects in clinical anticancer treatment for triple negative breast cancer.

The multi-functional system of electrochemical desalination, RhB degradation and Cr (VI) removal.

SYNOPSIS: : The single function of salt removal limits the further development of the CDI system. A multi-function CDI device is proposed to achieve electrochemical desalination, organics degradation and dichromate ion removal.

Physics-informed deep-learning parameterization of ocean vertical mixing improves climate simulations.

Uncertainties in ocean-mixing parameterizations are primary sources for ocean and climate modeling biases. Due to lack of process understanding, traditional physics-driven parameterizations perform unsatisfactorily in the tropics. Recent advances in the deep-learning method and the new availability of long-term turbulence measurements provide an opportunity to explore data-driven approaches to parameterizing oceanic vertical-mixing processes. Here, we describe a novel parameterization based on an artificial neural network trained using a decadal-long time record of hydrographic and turbulence observations in the tropical Pacific. This data-driven parameterization achieves higher accuracy than current parameterizations, demonstrating good generalization ability under physical constraints. When integrated into an ocean model, our parameterization facilitates improved simulations in both ocean-only and coupled modeling. As a novel application of machine learning to the geophysical fluid, these results show the feasibility of using limited observations and well-understood physical constraints to construct a physics-informed deep-learning parameterization for improved climate simulations.

hsa_circ_0115355 promotes pancreatic ß-cell function in patients with type 2 diabetes through the miR-145/SIRT1 axis.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is a complex metabolic disease closely related to obesity, a growing global health problem. T2DM is characterized by decreased islet beta-cell mass and impaired insulin release from these cells, and this dysfunction is exacerbated by hyperglycemia (glucolipotoxicity). Circular RNAs (circRNAs) are abnormally expressed and play a regulatory role in T2DM. OBJECTIVE: This study aimed to evaluate the function and molecular mechanism of hsa_circ_0115355 in the progression of T2DM. METHODS: The regulatory effect of hsa_circ_0115355 on INS-1 cell function was assessed under glucolipotoxicity by MTT, flow cytometry analysis, and insulin secretion assay. Dual-luciferase experiments revealed a direct interaction of hsa_circ_0115355 with miR-145 and miR-145 with SIRT1. Furthermore, the regulatory role of the hsa_circ_0115355/miR-145/SIRT1 axis was verified by examining the function of INS-1. RESULTS: In this study, hsa_circ_0115355 was significantly underexpressed in both patients with T2DM and INS-1 cell lines. This study thus showed that hsa_circ_0115355 inhibits the occurrence and development of T2DM by regulating the expression of SIRT1 by adsorbing miR-145. CONCLUSION: The underexpression hsa_circ_0115355 is also a potential novel diagnostic marker and therapeutic target for T2DM.

IGF1 receptor-targeted black TiO2 nanoprobes for MRI-guided synergetic photothermal-chemotherapy in drug resistant pancreatic tumor.

Pancreatic ductal adenocarcinoma (PDAC) is among the deadliest malignant tumors with features of matrix barrier caused poor drug permeability, and susceptibility to drug resistance. Herein, a PDAC and its stromal cell dual-targeted photothermal-chemotherapy strategy is explored to loosen the matrix and reverse drug resistance. To achieve this goal, black TiO2-Gd nanocomposites were conjugated with insulin like growth factor 1 (IGF1), and loaded with gemcitabine (GEM) to construct bTiO2-Gd-IGF1-GEM nanoprobes. In vitro results show that under 808 nm near-infrared irradiation, killing effect of the nanoprobes on drug-resistant MIA PaCa-2 cell is 3.3 times than that of GEM alone. In vivo experiments indicate the synergetic photothermal-chemotherapy not only loosens fibrous matrix of pancreatic tumor model, but also dramatically inhibits tumor growth, and almost completely eradicates the tumor after 12 days of treatment. In addition, relaxation rate of the nanoprobes is 8.2 times than commercial contrast agent Magnevist, therefore boosts the signal of magnetic resonance imaging in pancreatic tumor. In conclusion, our results reinforce that the prepared nanoprobes are promising to break matrix barrier and overcome drug resistance in PDAC.

Photo-Assisted Rechargeable Battery Desalination.

Herein, we propose a novel design of photo-assisted battery desalination, which provides the tri-function within a single device including the photo-assisted charge (electrical energy saving), energy storage, and desalination (salt removal). The photoelectrode (N719/TiO2) is directly integrated into the zinc-iodide (Zn-I) battery with the desalination stream in the middle portion of the device. This architecture can provide a reduced energy consumption up to 50%, an energy output of 42 W h mol-1NaCl, and a desalination rate of 13 µg/cm2 min-1. This work is significant for the inter-discipline study of the redox flow energy storage and energy-saving desalination.

Exosomal circular RNA hsa_circ_0006220, and hsa_circ_0001666 as biomarkers in the diagnosis of pancreatic cancer.

BACKGROUND: Pancreatic cancer is a highly malignant tumor of the digestive system. OBJECTIVE: Exosomal circular RNA can be used as a biomarker for the early diagnosis of pancreatic cancer. METHODS: The expression of various differentially expressed circRNAs in pancreatic cancer tissues was analyzed by gene chip, exosome expression was verified by electron microscopy and Western blotting, and the expression of exosomal circRNA in pancreatic cancer cells, tissues, and plasma were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: Compared with healthy controls, hsa_circ_0006220 and hsa_circ_0001666 were highly expressed in exosomes in the plasma of pancreatic cancer patients. The AUC values were 0.7817 for hsa_circ_0006220, 0.8062 for hsa_circ_0001666, and 0.884 for the combined diagnosis. In addition, clinicopathological features revealed that the expression of hsa_circ_0006220 in plasma exosomes from pancreatic cancer patients was associated with CA19-9 levels (p = 0.0001) and lymph node metastasis (p = 0.0005). The expression of hsa_circ_0001666 was correlated with both tumor size (p = 0.0157) and CA19-9 level (p = 0.0001). CONCLUSIONS: The high expression of exosomal hsa_circ_0001666 and hsa_circ_0006220 suggests that these can be used as new biomarkers for the diagnosis and treatment of pancreatic cancer.

Synergy between Toxoplasma gondii type I ΔGRA17 immunotherapy and PD-L1 checkpoint inhibition triggers the regression of targeted and distal tumors.

BACKGROUND: In this study, we hypothesize that the ability of the protozoan Toxoplasma gondii to modulate immune response within the tumor might improve the therapeutic effect of immune checkpoint blockade. We examined the synergetic therapeutic activity of attenuated T. gondii RH ΔGRA17 strain and programmed death ligand-1 (PD-L1) treatment on both targeted and distal tumors in mice. METHODS: The effects of administration of T. gondii RH ΔGRA17 strain on the tumor volume and survival rate of mice bearing flank B16-F10, MC38, or LLC tumors were studied. We characterized the effects of ΔGRA17 on tumor biomarkers' expression, PD-L1 expression, immune cells infiltrating the tumors, and expression of immune-related genes by using immunohistochemistry, immunofluorescence, flow cytometry, NanoString platform, and real-time quantitative PCR, respectively. The role of immune cells in the efficacy of ΔGRA17 plus PD-L1 blockade therapy was determined via depletion of immune cell subtypes. RESULTS: Treatment with T. gondii ΔGRA17 tachyzoites and anti-PD-L1 therapy significantly extended the survival of mice and suppressed tumor growth in preclinical mouse models of melanoma, Lewis lung carcinoma, and colon adenocarcinoma. Attenuation of the tumor growth was detected in the injected and distant tumors, which was associated with upregulation of innate and adaptive immune pathways. Complete regression of tumors was underpinned by late interferon-gamma-producing CD8+ cytotoxic T cells. CONCLUSION: The results from these models indicate that intratumoral injection of ΔGRA17 induced a systemic effect, improved mouse immune response, and sensitized immunologically 'cold' tumors and rendered them sensitive to immune checkpoint blockade therapy.

Protective Immunity Induced by TgMIC5 and TgMIC16 DNA Vaccines Against Toxoplasmosis.

Toxoplasma gondii is an obligate intracellular parasite, which is responsible for a widely distributed zoonosis. Effective vaccines against toxoplasmosis are necessary to protect the public health. The aim of this study is to evaluate the immune efficacy of DNA vaccines encoding TgMIC5 and TgMIC16 genes against T. gondii infection. The recombinant plasmid pVAX-MIC5 and pVAX-MIC16 were constructed and injected intramuscularly in mice. The specific immune responses and protection against challenge with T. gondii RH tachyzoites were evaluated by measuring the cytokine levels, serum antibody concentrations, lymphocyte proliferation, lymphocyte populations, and the survival time. The protection against challenge with the T. gondii RH tchyzoites and PRU cysts was examined by evaluation of the reduction in the brain cyst burden. The results indicated that immunized mice showed significantly increased levels of IgG, IFN-γ, IL-2, IL-12p70, and IL-12p40 and percentages of CD4+ and CD8+ T cells. Additionally, vaccination prolonged the mouse survival time and reduced brain cysts compared with controls. Mouse groups immunized with a two-gene cocktail of pVAX-MIC5 + pVAX-MIC16 were more protected than mouse groups immunized with a single gene of pVAX-MIC5 or pVAX-MIC16. These results demonstrate that TgMIC5 and TgMIC16 induce effective immunity against toxoplasmosis and may serve as a good vaccine candidate against T. gondii infection.

Immunization With a DNA Vaccine Encoding the Toxoplasma gondii' s GRA39 Prolongs Survival and Reduce Brain Cyst Formation in a Murine Model.

Toxoplasma gondii, an obligate intracellular protozoan parasite, can cause infect almost all warm-blooded animals and humans. To evaluate the immunogenicity and protective efficacy of T. gondii GRA39 (TgGRA39) in mice by using DNA immunization, we constructed a recombinant eukaryotic plasmid pVAX-TgGRA39. The specific immune responses in immunized mice were analyzed by serum antibody and cytokine measurements, lymphocyte proliferation assays and flow cytometry of T lymphocyte subclasses. Also, protective efficacy against acute and chronic T. gondii infection was assessed by observing the survival time after challenge with the highly virulent T. gondii RH strain (Genotype I) and counting the number of cyst-forming in brain at 4 weeks post-infection with the cyst-forming PRU strain of T. gondii (Genotype II), respectively. Our results showed that DNA immunization with pVAX-GRA39 via intramuscular injection three times, at 2-week intervals could elicit humoral and cellular immune response, indicated by enhanced levels of IgG and IgG2a antibodies (a slightly elevated IgG2a to IgG1 ratio), and increased levels of cytokines IFN-γ, IL-2, IL-12, IL-17A, IL-17F, IL-22 and IL-23 and percentages of CD3+ CD4+ CD8- and CD3+ CD8+ CD4- T cells, in contrast to non-immunized mice. The significant increase in the expression levels of IL-6, TGF-ß1, IL-1ß, and the transcription factor factors RORγt, RORα, and STAT3 involved in the activation and pathway of Th17 and Tc17 cells, were also observed. However, no significant difference was detected in level of IL-4 and IL-10 (p > 0.05). These effective immune responses had mounted protective immunity against T. gondii infection, with a prolonged survival time (16.80 ± 3.50 days) and reduced cyst numbers (44.5%) in comparison to the control mice. Our data indicated that pVAX-TgGRA39 could induce effective humoral, and Th1-type, Th17, and Tc17 cellular immune responses, and may represent a promising vaccine candidate against both acute and chronic T. gondii infection.

DMSO-Enabled Selective Radical O-H Activation of 1,3(4)-Diols.

Control of selectivity is one of the central topics in organic chemistry. Although unprecedented alkoxyl-radical-induced transformations have drawn a lot of attention, compared to selective C-H activation, selective radical O-H activation remains less explored. Herein, we report a novel selective radical O-H activation strategy of diols by combining spatial effects with proton-coupled electron transfer (PCET). It was found that DMSO is an essential reagent that enables the regioselective transformation of diols. Mechanistic studies indicated the existence of the alkoxyl radical and the selective interaction between DMSO and hydroxyl groups. Moreover, the distal C-C cleavage was realized by this selective alkoxyl-radical-initiation protocol.

Adjuvantic cytokine IL-33 improves the protective immunity of cocktailed DNA vaccine of ROP5 and ROP18 against toxoplasma gondii infection in mice.

Toxoplasma gondii is a threat for immunocompromized individuals, and no treatment is available for enhancing immunity against infection. Molecular adjuvants may improve the efficacy of DNA vaccine-induced T cell immunity. Here, we report that cocktailed DNA immunization with ROP5 and ROP18 boosted immune responses induced by a single DNA immunization with ROP5 or ROP18, but also that co-administration of molecular adjuvant IL-33 enhanced immune efficacy induced by this cocktailed DNA vaccination. These improved immune responses were characterized by higher Toxoplasma-specific IgG2a titers, Th1 responses associated with the production of IFN-γ, IL-2, IL-12, as well as cell-mediated activity with higher frequencies of CD8+ and CD4+ T cells. More importantly, this enhanced immunity has the ability to confer remarkable protection against a high dose lethal challenge of the T. gondii RH strain and thus against chronic infection with the T. gondii PRU strain. These data show that IL-33 is a promising immunoadjuvant to facilitate humoral as well as cellular immunity in a vaccine setting against T. gondii, and suggest that it should be evaluated in strategies against other apicomplexan parasites.

TITLE: La cytokine IL-33 utilisée comme adjuvant améliore l'immunité protectrice du vaccin à cocktail d'ADN de ROP5 et ROP18 contre l'infection à Toxoplasma gondii chez la souris. ABSTRACT: Toxoplasma gondii est une menace pour les individus immunodéprimés et aucun traitement n'est disponible pour renforcer l'immunité contre l'infection. Les adjuvants moléculaires peuvent améliorer l'efficacité de l'immunité des cellules T induite par un vaccin à ADN. Ici, nous rapportons que l'immunisation par le cocktail d'ADN de ROP5 et ROP18 a stimulé les réponses immunitaires induites par une seule immunisation par l'ADN de ROP5 ou ROP18, mais aussi que la co-administration de l'adjuvant moléculaire IL-33 a amélioré l'efficacité immunitaire induite par cette vaccination par cocktail d'ADN. Ces réponses immunitaires améliorées ont été caractérisées par des titres d'IgG2a spécifiques à Toxoplasma plus élevés, des réponses Th1 associées à la production d'IFN-γ, IL-2, IL-12 ainsi qu'une activité à médiation cellulaire où les fréquences des cellules T CD8+ et CD4+ étaient plus élevées. Plus important encore, cette immunité renforcée a la capacité de conférer une protection remarquable contre une provocation létale par haute dose de la souche RH de T. gondii et donc contre une infection chronique par la souche PRU de T. gondii. Ces données montrent qu'IL-33 est un immunoadjuvant prometteur pour faciliter l'immunité humorale et cellulaire dans un contexte de vaccination contre T. gondii et suggèrent qu'il devrait être évalué dans des stratégies contre d'autres parasites apicomplexes.

[Spray drying process of extract of Wenjing Zhitong Prescription based on mixture design experiment].

To improve the spray drying effect of extract of Wenjing Zhitong Prescription, this study takes the yield, hygroscopic property and the fluidity of dry powder as indexes to screen out auxiliary materials, and the proportion of the auxiliary materials was optimized based on the mixing design experiment; based on that, HPLC method was established for the determination of glycyrrhizin and 6-gingerol in spray powder, the yield of spray powder and the retention rate of the two index components were taken as indexes to further optimize the spray drying parameters. The finally selected auxiliary materials were light magnesium oxide, maltodextrin and silica, and regression equations of dry powder yield, moisture absorption rate, angle of rest with proportion of auxiliary materials were established, and the optimized proportion of auxiliary materials was dry paste-light magnesium oxide-maltodextrin-silica=0.5∶0.305∶0.145∶0.05; according to the optimized drying process parameters of Wenjing Zhitong Prescription, initial temperature was 60 ℃, air inlet temperature was 130 ℃, air flow rate was 35 m~3·h~(-1), atomizing pressure was 40 mm, and liquid inlet speed was 4.5 mL·min~(-1). Under these conditions, the dry powder yield was 90.28%, the retention rate of glycyrrhizin was 74.51%, and the retention rate of 6-gingerol was 72.10%. In this study, optimized auxiliary materials can improve the yield of spray drying and the property of spray powder, and the optimized processing conditions were good for retaining the unstable gingerol components, which can lay a foundation for the further preparation research of meridian warming and pain relieving prescriptions, and provide reference for extract of other traditional Chinese medicine extracts that are difficult to spray drying.

Selective Aerobic Oxygenation of Tertiary Allylic Alcohols with Molecular Oxygen.

Aerobic epoxidation of tertiary allylic alcohols remains a significant challenge. Reported here is an efficient and highly chemoselective copper-catalyzed epoxidation and semipinacol rearrangement reaction of tertiary allylic alcohols with molecular oxygen. The solvent 1,4-dioxane activates dioxygen, thereby precluding the addition of a sacrificial reductant.

Acetonitrile Activation: An Effective Two-Carbon Unit for Cyclization.

A novel activation of acetonitrile for the construction of cyclobutenones by [2+2] cyclization was developed. Acetonitrile is utilized for the first time as two-carbon (C2) cyclization building block. The present protocol successfully inhibits the competitive cycloaddition with the C≡N bond of acetonitrile, but enables the in situ formation of an unsaturated carbon-carbon bond and the subsequent cycloaddition as a C2 unit. This chemistry features simple reaction conditions, high chemoselectivities, wide substrate scope, and offers a new and practical approach to cyclobutenones and cyclobuteneimines.

Author Correction: From alkylarenes to anilines via site-directed carbon-carbon amination.

The version of this Article originally published online did not include a caution statement relating to safety concerns over some of the reagents used. All versions of the Article now have the following text included at the start of the Methods section "Caution: Sodium azide (NaN3) is highly toxic and also a potential explosion hazard; it can also react with organohalides to form explosive organic azides. Under acidic conditions, sodium azide can form hydrazoic acid (HN3) which is highly toxic. Considering these hazards, appropriate safety precautions should be taken when undertaking the C-C amination reactions reported in this Article." Furthermore, in Fig. 2a, in the reaction conditions on the first equation it should have read DDQ not DDG; this has also been amended.

From alkylarenes to anilines via site-directed carbon-carbon amination.

Anilines are fundamental motifs in various chemical contexts, and are widely used in the industrial production of fine chemicals, polymers, agrochemicals and pharmaceuticals. A recent development for the synthesis of anilines uses the primary amination of C-H bonds in electron-rich arenes. However, there are limitations to this strategy: the amination of electron-deficient arenes remains a challenging task and the amination of electron-rich arenes has a limited control over regioselectivity-the formation of meta-aminated products is especially difficult. Here we report a site-directed C-C bond primary amination of simple and readily available alkylarenes or benzyl alcohols for the direct and efficient preparation of anilines. This chemistry involves a novel C-C bond transformation and offers a versatile protocol for the synthesis of substituted anilines. The use of O2 as an environmentally benign oxidant is demonstrated, and studies on model compounds suggest that this method may also be used for the depolymerization of lignin.

Silver-catalyzed remote Csp3-H functionalization of aliphatic alcohols.

Aliphatic alcohols are common and bulk chemicals in organic synthesis. The site-selective functionalization of non-activated aliphatic alcohols is attractive but challenging. Herein, we report a silver-catalyzed δ-selective Csp3-H bond functionalization of abundant and inexpensive aliphatic alcohols. Valuable oximonitrile substituted alcohols are easily obtained by using well-designed sulphonyl reagents under simple and mild conditions. This protocol realizes the challenging δ-selective C-C bond formation of simple alkanols.