Hyperfractionation compared with standard fractionation in intensity-modulated radiotherapy for patients with locally advanced recurrent nasopharyngeal carcinoma: a multicentre, randomised, open-label, phase 3 trial.

BACKGROUND: Reirradiation in standard fractionation for locally advanced recurrent nasopharyngeal carcinoma after a previous course of high-dose radiotherapy is often associated with substantial late toxicity, negating its overall benefit. We therefore aimed to investigate the efficacy and safety of hyperfractionation compared with standard fractionation in intensity-modulated radiotherapy. METHODS: This multicentre, randomised, open-label, phase 3 trial was done in three centres in Guangzhou, China. Eligible patients were aged 18-65 years with histopathologically confirmed undifferentiated or differentiated, non-keratinising, advanced locally recurrent nasopharyngeal carcinoma. Participants were randomly assigned (1:1) to either receive hyperfractionation (65 Gy in 54 fractions, given twice daily with an interfractional time interval of at least 6 h) or standard fractionation (60 Gy in 27 fractions, given once a day). Intensity-modulated radiotherapy was used in both groups. A computer program generated the assignment sequence and randomisation was stratified by treatment centre, recurrent tumour stage (T2-T3 vs T4), and recurrent nodal stage (N0 vs N1-N2), determined at the time of randomisation. The two primary endpoints were the incidence of severe late complications defined as the incidence of grade 3 or worse late radiation-induced complications occurring 3 months after the completion of radiotherapy until the latest follow-up in the safety population, and overall survival defined as the time interval from randomisation to death due to any cause in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, NCT02456506. FINDINGS: Between July 10, 2015, and Dec 23, 2019, 178 patients were screened for eligibility, 144 of whom were enrolled and randomly assigned to hyperfractionation or standard fractionation (n=72 in each group). 35 (24%) participants were women and 109 (76%) were men. After a median follow-up of 45·0 months (IQR 37·3-53·3), there was a significantly lower incidence of grade 3 or worse late radiation-induced toxicity in the hyperfractionation group (23 [34%] of 68 patients) versus the standard fractionation group (39 [57%] of 68 patients; between-group difference -23% [95% CI -39 to -7]; p=0·023). Patients in the hyperfractionation group had better 3-year overall survival than those in the standard fractionation group (74·6% [95% CI 64·4 to 84·8] vs 55·0% [43·4 to 66·6]; hazard ratio for death 0·54 [95% CI 0·33 to 0·88]; p=0·014). There were fewer grade 5 late complications in the hyperfractionation group (five [7%] nasal haemorrhage) than in the standard fractionation group (16 [24%], including two [3%] nasopharyngeal necrosis, 11 [16%] nasal haemorrhage, and three [4%] temporal lobe necrosis). INTERPRETATION: Hyperfractionated intensity-modulated radiotherapy could significantly decrease the rate of severe late complications and improve overall survival among patients with locally advanced recurrent nasopharyngeal carcinoma. Our findings suggest that hyperfractionated intensity-modulated radiotherapy could be used as the standard of care for these patients. FUNDING: Key-Area Research and Development of Guangdong Province, the National Natural Science Foundation of China, the Special Support Program for High-level Talents in Sun Yat-sen University Cancer Center, the Guangzhou Science and Technology Plan Project, and the National Ten Thousand Talents Program Science and Technology Innovation Leading Talents, Sun Yat-Sen University Clinical Research 5010 Program.

Outcomes of patients in nasopharyngeal adenoid cystic carcinoma in the IMRT era: a single-center experience.

OBJECTIVE: Nasopharyngeal adenoid cystic carcinoma (NACC) is a rare malignancy with special biological features. Controversies exist regarding the treatment approach and prognostic factors in the IMRT era. This study aimed to evaluate the long-term outcomes and management approaches in NACC. METHODS: Fifty patients with NACC at our institution between 2010 and 2020 were reviewed. Sixteen patients received primary radiotherapy (RT), and 34 patients underwent primary surgery. RESULTS: Between January 2010 and October 2020, a total of 50 patients with pathologically proven NACC were included in our analysis. The median follow-up time was 58.5 months (range: 6.0-151.0 months). The 5-year overall survival rate (OS) and progression-free survival rate (PFS) were 83.9% and 67.5%, respectively. The 5-year OS rates of patients whose primary treatment was surgery and RT were 90.0% and 67.3%, respectively (log-rank P = 0.028). The 5-year PFS rates of patients whose primary treatment was surgery or RT were 80.8% and 40.7%, respectively (log-rank P = 0.024). Multivariate analyses showed that nerve invasion and the pattern of primary treatment were independent factors associated with PFS. CONCLUSIONS: Due to the relative insensitivity to radiation, primary surgery seemed to provide a better chance of disease control and improved survival in NACC. Meanwhile, postoperative radiotherapy should be performed for advanced stage or residual tumours. Cranial nerve invasion and treatment pattern might be important factors affecting the prognosis of patients with NACC.

Molecular PET/CT mapping of rhACE2 distribution and quantification in organs to aid in SARS-CoV-2 targeted therapy.

Coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, poses a significant threat to public health. Angiotensin-converting enzyme 2 (ACE2) is a key receptor for SARS-CoV-2 infection. Recombinant human ACE2 (RhACE2), as a soluble supplement for human ACE2, can competitively block SARS-CoV-2 infection. In this study, a mouse organ in situ rhACE2 high aggregation model was constructed for the first time, and in vivo real-time positron emission tomography (PET) imaging of rhACE2 in the mouse model was performed using an ACE2-specific agent 68 Ga-HZ20. This radiotracer exhibits reliable radiochemical properties in vitro and maintains a high affinity for rhACE2 in vivo. In terms of probe uptake, 68 Ga-HZ20 showed a good target-to-nontarget ratio and was rapidly cleared from the circulatory system and excreted by the kidneys and urinary system. PET imaging with this radiotracer can noninvasively and accurately monitor the content and distribution of rhACE2 in the body, which clarifies that rhACE2 can aggregate in multiple organs, suggesting the preventive and therapeutic potential of rhACE2 for SARS-CoV-2 and COVID-19.

Erianin Exerts Antineoplastic Effects on Esophageal Squamous Cell Carcinoma Cells by Activating the cGMP-PKG Signaling Pathway.

Erianin, a natural compound extracted from Dendrobium chrysotoxum Lindl, has potential therapeutic benefits against various tumors. However, its role in esophageal squamous cell carcinoma (ESCC) remains unclear. Cell proliferation was analyzed by CCK8, colony-formation, and EdU proliferation assays, while cell migration was evaluated through wound healing assays as well as determination of the protein expression levels of epithelial-to-mesenchymal transition (EMT) markers and ß-catenin. Apoptosis was measured by flow cytometry. RNA sequencing (RNA-seq) and bioinformatic analyses were performed to elucidate the underlying mechanisms of erianin in ESCC. Enzyme-linked immunosorbent assay (ELISA) was used to determine intracellular cGMP, cleaved-PARP, and caspase-3/7 activity, while mRNA and protein levels were determined by qRT-PCR and western blotting, respectively. Our results indicate that erianin significantly inhibited ESCC cell proliferation and migration while also promoting apoptosis. Mechanistically, RNA sequencing coupled with KEGG enrichment analysis and functional assays revealed that activation of the cGMP-PKG pathway contributed to the antitumor effects of erianin, whereas the c-GMP-dependent protein kinase inhibitor KT5823 significantly attenuated these effects. In conclusion, our results demonstrate that erianin suppresses the proliferation of ESCC cells by activating the cGMP-PKG pathway, suggesting that erianin could be a promising candidate for the treatment of ESCC.

Electronic Nose and Head Space GC-IMS Provide Insights into the Dynamic Changes and Regularity of Volatile Compounds in Zangju (Citrus reticulata cv. Manau Gan) Peel at Different Maturation Stages.

Zangju (Citrus reticulata cv. Manau Gan) is the main citrus cultivar in Derong County, China, with unique aroma and flavour characteristics, but the use of Zangju peel (CRZP) is limited due to a lack of research on its peel. In this study, electronic nose, headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS), and partial least squares-discriminant analysis (PLS-DA) methods were used to rapidly and comprehensively evaluate the volatile compounds of dried CRZP and to analyse the role of dynamic changes at different maturation stages. The results showed that seventy-eight volatile compounds, mainly aldehydes (25.27%) and monoterpenes (55.88%), were found in the samples at four maturity stages. The contents of alcohols and aldehydes that produce unripe fruit aromas are relatively high in the immature stage (October to November), while the contents of monoterpenoids, ketones and esters in ripe fruit aromas are relatively high in the full ripening stage (January to February). The PLS-DA model results showed that the samples collected at different maturity stages could be effectively discriminated. The VIP method identified 12 key volatile compounds that could be used as flavour markers for CRZP samples collected at different maturity stages. Specifically, the relative volatile organic compounds (VOCs) content of CRZP harvested in October is the highest. This study provides a basis for a comprehensive understanding of the flavour characteristics of CRZP in the ripening process, the application of CRZP as a byproduct in industrial production (food, cosmetics, flavour and fragrance), and a reference for similar research on other C. reticulata varieties.

[Assay of amino acids in leaves of Eucommia ulmoides under arbor forest mode and leaf-oriented cultivation mode by pre-column derivatization HPLC].

This study aims to develop the pre-column derivatization high performance liquid chromatography(HPLC) method for the determination of 16 kinds of amino acids in Eucommia ulmoides leaves, and compare the content of amino acids in the leaves harvested at different time and under leaf-oriented cultivation mode(LCM) and arbor forest mode(AFM). The HPLC conditions are as below: phenyl isothiocyanate(PITC) as pre-column derivatization agent, Agilent ZORBAX C_(18 )column(4.6 mm×250 mm, 5 µm), mobile phase A of acetonitrile-water(80∶20), mobile phase B of 0.1 mol·L~(-1) sodium acetate solution-acetonitrile(94∶6), gradient elution, flow rate of 1.0 mL·min~(-1), injection volume of 5 µL, column temperature of 40 ℃, and detection wavelength of 254 nm. The HPLC profile indicated well separation of 16 kinds of amino acids and the amino acid content in E. ulmoides leaves was up to 16.26%. In addition, the amino acid content in leaves of E. ulmoides under LCM was higher than under AFM. The amino acid content varied with the harvesting time. Through orthogonal partial least squares discriminant analysis, the amino acids of E. ulmoides under LCM and AFM were compared, which can distinguish the leaves under LCM from those under AFM. Principal component analysis was applied to comprehensively score the amino acids of E. ulmoides leaves. The results showed that the score of leaves under LCM was higher than that under AFM. Nutritional evaluation results indicated that the proteins in E. ulmoides leaves belonged to high-quality vegetable proteins. The established method for the determination of amino acid content is reliable. With the amino acid content as index, the leaf quality of E. ulmoides under LCM is better than that under AFM. This study lays a theoretical basis for the promotion of LCM for E. ulmoides and the development of medicinal and edible products from E. ulmoides leaves.

[Effects of different drying methods on quality of male flowers of Eucommia ulmoides based on color and chemical composition].

To study the effects of different drying methods on the quality of male flowers of Eucommia ulmoides(MFOEU), we treated fresh MFOEU samples with drying in the shade(DS), vacuum freeze drying(VFD), high-or low-temperature hot air drying(HTHAD, LTHAD), microwave drying(MD), and vacuum drying(VD), respectively. The color, total flavonoid content, total polysaccharide content, and main active components such as geniposide, geniposidic acid, rutin, chlorogenic acid, galuteolin, pinoresinol diglucoside, and aucubin in MFOEU were taken as the evaluation indicators. The quality of MFOEU was comprehensively evaluated by entropy weight method combined with color index method, partial least squares discriminant analysis and content clustering heat map. The experimental results showed that VFD and DS basically kept the original color of MFOEU. The MFOEU treated with MD had higher content of total polysaccharides, phenylpropanoids, lignans, and iridoids. The MFOEU treated with LTHAD had higher content of total flavonoids and that treated with VD had lower content of active components. According to the results of comprehensive evaluation, the quality of MFOEU dried with different methods followed the order of MD>HTHAD>VFD>LTHAD>DS>VD. Considering the color of MFOEU, the suitable drying methods were DS and VFD. Considering the color, active components, and economic benefits of MFOEU, MD was the suitable drying method. The results of this study are of a reference value for the determination of suitable methods for MFOEU processing in the producing areas.

[Spleen deficiency and phlegm dampness syndrome model in rats treated by Citri Reticulatae Pericarpium based on metabolomics].

The present study analyzed the effect of Citri Reticulatae Pericarpium on endogenous metabolites in spleen deficiency and phlegm dampness syndrome by metabolomics, and explored the underlying mechanism of Citri Reticulatae Pericarpium in the treatment of spleen deficiency and phlegm dampness syndrome.The model of spleen deficiency and phlegm dampness syndrome was induced in rats by the multi-factor modeling method.The intervention effects of Citri Reticulatae Pericarpium on rats with spleen deficiency and phlegm dampness syndrome were preliminarily evaluated by observing the pathological changes of rat liver tissues and measuring the plasma content of pathological and biochemical indexes such as triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), and high-density lipoprotein cholesterol(HDL-C).Immunohistochemistry was used to detect the expression of AQP2 in the kidney, AQP3 in the colon, and AQP5 in the submandibular gland, and the effect of Citri Reticulatae Pericarpium on aquaporin expression in rats with spleen deficiency and phlegm dampness syndrome was evaluated.Furthermore, UHPLC-ESI-MS/MS was used to analyze the metabolic profiles of rat plasma samples.Multiple methods, such as principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) were used for pattern recognition.Differential metabolites were screened out by t-test and variable importance in projection(VIP), followed by pathway analysis based on MetaboAnalyst 5.0.As revealed by experimental results, Citri Reticulatae Pericarpium could improve the pathological changes of liver tissues, increase the levels of HDL-C in the plasma, reduce the levels of TC, TG, and LDL-C, and enhance the expression of AQP2 in the kidney, AQP3 in the colon, and AQP5 in the submandibular gland of rats with spleen deficiency and phlegm dampness syndrome.In addition, 87 differential metabolites of spleen deficiency and phlegm dampness syndrome were screened out by UHPLC-ESI-MS/MS(the levels of 39 metabolites increased significantly and the levels of 48 metabolites decreased significantly), with the representatives of glycine, L-isoleucine, N-acetyl-L-tyrosine, xanthine, hypoxanthine, and trigonelline.The differential metabolites were mainly enriched in the pathways of steroid hormone biosynthesis, linoleic acid metabolism, and purine metabolism.This study distinguished and revealed the characteristic metabolic pattern of spleen deficiency and phlegm dampness syndrome by metabolomics.The preliminary construction of the OPLS-DA model provides an objective basis for the differentiation of spleen deficiency and phlegm dampness syndrome in traditional Chinese medi-cine(TCM), as well as ideas and methods for exploring the biological basis of TCM syndrome from the molecular level and the overall level.

[Toxicity mechanism of Rhododendri Mollis Flos: based on serum metabolomics and network toxicology].

This study aims to explore the toxicity mechanism of Rhododendri Mollis Flos(RMF) based on serum metabolomics and network toxicology. The toxic effect of RMF on normal rats was evaluated according to the symptoms, serum biochemical indexes, and histopathology. Serum metabolomics was combined with multivariate statistical analysis to search endogenous differential metabolites and related metabolic pathways. The toxic components, targets, and signaling pathways of RMF were screened by network toxicology technique, and the component-target-metabolite-metabolic pathway network was established with the help of serum metabolomics. The result suggested the neurotoxicity, hepatotoxicity, and cardiotoxicity of RMF. A total of 31 differential metabolites and 10 main metabolic pathways were identified by serum metabolomics, and 11 toxic components, 332 related target genes and 141 main signaling pathways were screened out by network toxicology. Further analysis yielded 7 key toxic components: grayanotoxin Ⅲ,grayanotoxinⅠ, rhodojaponin Ⅱ, rhodojaponin Ⅴ, rhodojaponin Ⅵ, rhodojaponin Ⅶ, and kalmanol, which acted on the following 12 key targets: androgen receptor(AR), albumin(ALB), estrogen receptor ß(ESR2), sex-hormone binding globulin(SHBG), type 11 hydroxysteroid(17-beta) dehydrogenase(HSD17 B11), estrogen receptor α(ESR1), retinoic X receptor-gamma(RXRG), lactate dehydrogenase type C(LDHC), Aldo-keto reductase(AKR) 1 C family member 3(AKR1 C3), ATP binding cassette subfamily B member 1(ABCB1), UDP-glucuronosyltransferase 2 B7(UGT2 B7), and glutamate-ammonia ligase(GLUL). These targets interfered with the metabolism of gamma-aminobutyric acid, estriol, testosterone, retinoic acid, 2-oxobutyric acid, and affected 4 key metabolic pathways of alanine, aspartate and glutamate metabolism, cysteine and methionine metabolism, steroid hormone biosynthesis, and retinol metabolism. RMF exerts toxic effect on multiple systems through multiple components, targets, and pathways. Through the analysis of key toxic components, target genes, metabolites, and metabolic pathways, this study unveiled the mechanism of potential neurotoxicity, cardiotoxicity, and hepatotoxicity of RMF, which is expected to provide a clue for the basic research on toxic Chinese medicinals.

Endoscopic surgery compared with intensity-modulated radiotherapy in resectable locally recurrent nasopharyngeal carcinoma: a multicentre, open-label, randomised, controlled, phase 3 trial.

BACKGROUND: The role of surgery compared with reirradiation in the primary treatment of patients with resectable, locally recurrent nasopharyngeal carcinoma (NPC) who have previously received radiotherapy is a matter of debate. In this trial, we compared the efficacy and safety outcomes of salvage endoscopic surgery versus intensity-modulated radiotherapy (IMRT) in patients with resectable locally recurrent NPC. METHODS: This multicentre, open-label, randomised, controlled, phase 3 trial was done in three hospitals in southern China. We included patients aged 18-70 years with a Karnofsky Performance Status score of at least 70 who were histopathologically diagnosed with undifferentiated or differentiated, non-keratinising, locally recurrent NPC with tumours confined to the nasopharyngeal cavity, the post-naris or nasal septum, the superficial parapharyngeal space, or the base wall of the sphenoid sinus. Eligible patients were randomly assigned (1:1) to receive either endoscopic nasopharyngectomy (ENPG group) or IMRT (IMRT group). Randomisation was done manually using a computer-generated random number code and patients were stratified by treatment centre. Treatment group assignment was not masked. The primary endpoint was overall survival, compared between the groups at 3 years. Efficacy analyses were done by intention to treat. Safety analysis was done in patients who received treatment according to the treatment they actually received. This trial was prospectively registered at the Chinese Clinical Trial Registry, ChiCTR-TRC-11001573, and is currently in follow-up. FINDINGS: Between Sept 30, 2011, and Jan 16, 2017, 200 eligible patients were randomly assigned to receive either ENPG (n=100) or IMRT (n=100). At a median follow-up of 56·0 months (IQR 42·0-69·0), 74 patients had died (29 [29%] of 100 patients in the ENPG group and 45 [45%] of 100 patients in the IMRT group). The 3-year overall survival was 85·8% (95% CI 78·9-92·7) in the ENPG group and 68·0% (58·6-77·4) in the IMRT group (hazard ratio 0·47, 95% CI 0·29-0·76; p=0·0015). The most common grade 3 or worse radiation-related late adverse event was pharyngeal mucositis (in five [5%] of 99 patients who underwent ENPG and 26 [26%] of 101 patients who underwent IMRT). Five [5%] of the 99 patients who underwent ENPG and 20 [20%] of the 101 patients who underwent IMRT died due to late toxic effects specific to radiotherapy; attribution to previous radiotherapy or trial radiotherapy is unclear due to the long-term nature of radiation-related toxicity. INTERPRETATION: Endoscopic surgery significantly improved overall survival compared with IMRT in patients with resectable locally recurrent NPC. These results suggest that ENPG could be considered as the standard treatment option for this patient population, although long-term follow-up is needed to further determine the efficacy and toxicity of this strategy. FUNDING: Sun Yat-sen University Clinical Research 5010 Program.

The role of the bacterial microbiome in the treatment of cancer.

The human microbiome is defined as the microorganisms that reside in or on the human body, such as bacteria, viruses, fungi, and protozoa, and their genomes. The human microbiome participates in the modulation of human metabolism by influencing several intricate pathways. The association between specific bacteria or viruses and the efficacy of cancer treatments and the occurrence of treatment-related toxicity in cancer patients has been reported. However, the understanding of the interaction between the host microbiome and the cancer treatment response is limited, and the microbiome potentially plays a greater role in the treatment of cancer than reported to date. Here, we provide a thorough review of the potential role of the gut and locally resident bacterial microbiota in modulating responses to different cancer therapeutics to demonstrate the association between the gut or locally resident bacterial microbiota and cancer therapy. Probable mechanisms, such as metabolism, the immune response and the translocation of microbiome constituents, are discussed to promote future research into the association between the microbiome and other types of cancer. We conclude that the interaction between the host immune system and the microbiome may be the basis of the role of the microbiome in cancer therapies. Future research on the association between host immunity and the microbiome may improve the efficacy of several cancer treatments and provide insights into the cause of treatment-related side effects.

Use of High-Throughput Sequencing to Identify Fungal Communities on the Surface of Citri Reticulatae Pericarpium During the 3-Year Aging Process.

Citri Reticulatae Pericarpium (CRP) is a natural product that is used widely in food and is an ingredient in traditional Chinese medicine. CRP improves gradually with aging; this process typically takes 3 years or more. During the aging process, CRP can be colonized with fungi and mildew. Molds and mildew may result in an increased flavonoid content; however, this has been observed only in response to fungi of the genera Penicillium and Aspergillus. As fungal colonization may alter the quality and properties of CRP, it is critical to have an understanding of the fungal communities detected on the surface of CRP during the aging process. We used a high-throughput sequencing (HiSeq) platform to sequence internal transcribed spacers (ITS) region to identify the contaminants associated with CRP during the 3-year aging process. We also evaluated the distribution of the dominant fungi of the genera Aspergillus and Penicillium over time. At the phylum level, we identified Ascomycota (36.26%) and Basidiomycota (18.98%), along with smaller populations of Mucoromycota, Glomeromycota, and Mortierellomycota. At the genus level, the fungi detected include Wallemia (12.40%), Cystofilobasidium (4.62%), Zasmidium (4.52%), Cladosporium (3.72%), Hanseniaspora (3.55%), Fusarium (3.49%), Kurtzmaniella (2.03%), Candida (1.74%), Passalora (1.47%), Ceramothyrium (1.33%), Mucor (1.07%), and Aspergillus (1.03%). Fungi of the genus Penicillium were detected primarily during the first year of storage. By contrast, fungi of the genus Aspergillus were not detected during the early stages (fresh peel-8 months), but appeared gradually at later stages of the aging process. Taken together, our results indicate that HiSeq is an effective method to study the changes in fungal communities that develop on the CRP surface over time. These findings provide a basis for further research into the correlation between dominant fungi and the mechanisms underlying the successful aging of CRP.

Fast Screening of Biomembrane-Permeable Compounds in Herbal Medicines Using Bubble-Generating Magnetic Liposomes Coupled with LC-MS.

A novel strategy based on the use of bionic membrane camouflaged magnetic particles and LC-MS was developed to quickly screen the biomembrane-permeable compounds in herbal medicines. The bionic membrane was constructed by bubble-generating magnetic liposomes loaded with NH4HCO3 (BMLs). The lipid bilayer structure of the liposomes enabled BMLs to capture biomembrane-permeable compounds from a herbal extract. The BMLs carrying the compounds were then separated from the extract by a magnetic field. Upon heat treatment, NH4HCO3 rapidly decomposed to form CO2 bubbles within the liposomal bilayer, and the captured compounds were released from BMLs and analyzed by LC-MS. Jinlingzi San (JLZS), which contains various natural ingredients, was chosen to assess the feasibility of the proposed method. As a result, nine potential permeable compounds captured by BMLs were identified for the first time. Moreover, an in vivo animal study found that most of the compounds screened out by the proposed method were absorbed into the blood. The study provides a powerful tool for rapid and simultaneous prediction of multiple biomembrane-permeable components.

Risk profiling using metabolomic characteristics for susceptible individuals of drug-induced liver injury caused by Polygonum multiflorum.

Idiosyncratic drug-induced liver injury (IDILI) is a rare but potentially severe adverse drug reaction. To date, identifying individuals at risk for IDILI remains challenging. This is a prospective study, where a nested case-control (1:5) design was adopted. For six patients who had abnormalities in liver function test after Polygonum multiflorum Thunb. (PM) ingestion (susceptible group), 30 patients with normal liver function were matched (tolerant group). Based on liquid chromatography-mass spectrometry, metabolomics analysis was done on serum samples prior to PM ingestion, to screen the differential metabolites and characterize metabolomic profiles of patient serum in the two groups. Multivariate analysis showed that there were remarkable separations between susceptible and tolerant groups. A total of 25 major differential metabolites were screened out, involving glycerophospholipid metabolism, sphingolipid metabolism, fatty acid metabolism, histidine metabolism and aromatic amino acid metabolism. Wherein, the area under the curve of the receiver operating characteristic curves of metabolites PE 22:6, crotonoyl-CoA, 2E-tetradecenoyl-CoA, phenyllactic acid, indole-5,6-quinone, phosphoribosyl-ATP were all greater than 0.9. The overall serum metabolic profile comprising of 25 metabolites could clearly distinguish susceptible and tolerant groups. This proof-of-concept study used metabolomics to characterize the metabolic profile of IDILI risk individuals before drug ingestion for the first time. The metabolome characteristics in patient serum before PM ingestion may predict the risk of liver injury after PM ingestion.

Components synergy between stilbenes and emodin derivatives contributes to hepatotoxicity induced by Polygonum multiflorum.

Polygonum multiflorum Thunb. (PM) is a famous traditional Chinese medicine with liver tonic effect, but arousing great concerns for hepatotoxicity issue. In this study, we elucidated the contribution of the two major compounds, emodin-8-O-ß-D-glucoside (EG) and 2,3,5,4´-tetrahydroxyl diphenylethylene-2-O-glucoside (TSG), in PM-induced liver injury.Based on LC-MS, the two concerned compounds were detected simultaneously in the sera of patients with PM-induced liver injury. In the lipopolysaccharide (LPS)-mediated inflammatory stress rat model, by the analysis of plasma biochemistry and liver histopathology, we observed that the solo treatment of EG, not TSG, could induce significant liver injury; and the combined administration of EG and TSG caused more severe liver injury than that of EG.Metabolomics analysis revealed that the EG-triggered liver injury was associated with significant disturbances of sphingolipids and primary bile acids metabolism pathways. In the combined administration group, much more disturbances in EG-triggered metabolic pathways, as well as alterations of several additional pathways such as retinol metabolism and vitamin B6 metabolism, were observed.Taken together, we considered EG was involved in the idiosyncratic liver injury of PM, and TSG played a synergetic role with EG, which contributed to the understanding of the hepatotoxic basis of PM.

Optimisation for simultaneous determination of iridoid glycosides and oligosaccharides in Radix Rehmannia by microwave assisted extraction and HILIC-UHPLC-TQ-MS/MS.

Hydrophilic iridoid glycosides and oligosaccharides are the major active ingredients of Radix Rehmanniae. Analysis of oligosaccharides is a challenging task because they are highly hydrophilic, with similar chemical structures and absence of chromophore and fluorophore groups. The difficulty for simultaneous analysis of iridoid glycosides and oligosaccharides in Radix Rehmanniae is increased due to the polarity difference between the two types of ingredients. OBJECTIVE: To develop a method for simultaneous determination of iridoid glycosides (ajugol, catalpol) and oligosaccharides (sucrose, melibiose, raffinose, mannotriose and stachyose) in Radix Rehmanniae. METHODOLOGY: Microwave-assisted extraction (MAE) was established to extract target analytes from Radix Rehmanniae samples using methanol-water (60:40, v/v) as the extraction solvent. Fast separation of seven analytes was achieved by hydrophilic interaction liquid chromatography (HILIC) using an Accucore-150-Amide-HILIC column. Sensitive and selective detection of the analytes was performed by triple quadrupole tandem mass spectrometry (TQ-MS/MS) using multiple reaction monitoring in positive electrospray ionisation mode. RESULTS: Good linearities were achieved for all the analytes with the correlation coefficients above 0.9991. The precisions resulted in deviations of less than 5.0% and the recoveries ranged from 93.8% to 105.5%. The established method was successfully applied to the analysis of iridoid glycosides and oligosaccharides in 12 samples of crude and processed Radix Rehmanniae. CONCLUSION: A simple, rapid and sensitive method based on MAE combined with HILIC-UHPLC-TQ-MS/MS was developed for simultaneous determination of iridoid glycosides and oligosaccharides in Radix Rehmanniae for the first time. The method exhibited excellent performance with simple sample preparation, short analysis time, high selectivity and sensitivity.

Relationship of circulating tumor cells and Epstein-Barr virus DNA to progression-free survival and overall survival in metastatic nasopharyngeal carcinoma patients.

We analyzed the number of circulating tumor cells (CTCs) and Epstein-Barr virus DNA (EBV DNA) for diagnosis, monitoring and prognosis of patients with metastatic nasopharyngeal carcinoma (mNPC). The levels of CTCs and EBV DNA were measured at baseline and after first-line chemotherapy in 148 mNPC patients prospectively enrolled between December 2014 and August 2016. We also collected 122 non-mNPC cases within the same time frame for examining CTCs and EBV DNA at baseline. In 270 NPC patients, we observed improved specificity (86.0% vs. 41.0%) and inferior sensitivity (42.3% vs. 81.3%) of CTCs as compared to EBV DNA for diagnosis of distant metastasis. mNPC patients were stratified into unfavorable and favorable prognostic groups, respectively, based on CTC of 12 at baseline and 1 after first-line chemotherapy and EBV DNA of 10,000 at baseline and 4,000 after first-line chemotherapy. Conversion of baseline unfavorable CTCs and EBV DNA to favorable after first-line chemotherapy was associated with significantly longer progression-free survival (PFS) and overall survival (OS) compared to patients with unfavorable CTCs and EBV DNA at both time points. Among patients with a complete/partial response as per imaging evaluation, favorable CTCs and EBV DNA levels after first-line chemotherapy were associated with significantly longer PFS and OS. In conclusion, our data demonstrated the number of CTCs and EBV DNA before, after and during first-line chemotherapy were strong predictive markers for mNPC patients. When utilized in conjunction with imaging studies, CTCs and EBV DNA could provide additional prognostic information.

Solid-phase extraction coupled with switchable hydrophilicity solvent-based homogeneous liquid-liquid microextraction for chloramphenicol enrichment in environmental water samples: a novel alternative to classical extraction techniques.

A simple and efficient method combining solid-phase extraction (SPE) with homogeneous liquid-liquid microextraction (HLLME) has been developed for fast pretreatment of chloramphenicol (CAP) from water samples prior to determination by high-performance liquid chromatography-ultraviolet detection. Oasis HLB sorbent was chosen for SPE. In HLLME, N,N-dimethylcyclohexylamine was used as a CO2-triggered switchable solvent that could switch reversibly between hydrophilic and hydrophobic forms. The parameters influencing both SPE and HLLME were investigated and optimized. Under the optimal conditions, the method exhibited low limit of detection (0.1 ng/mL), good linearity (0.5-50 ng/mL), acceptable precision (RSD <5.0%) and accuracy (RE <4.0%). An enrichment factor of 340 was obtained. The proposed method is simple, fast, cost-effective, and suitable for the determination of trace chloramphenicol in water matrices. Graphical abstract ᅟ.

Dissolvable layered double hydroxide as a sorbent in dispersive micro-solid phase extraction for the determination of acidic quinolones in honey by HPLC.

This study presents a simple and green dispersive micro-solid phase extraction method for preconcentration of acidic quinolones from honey prior to high performance liquid chromatography determination. A two-dimensional nanostructured zinc-aluminum layered double hydroxide was synthesized and used as the sorbent for dispersive micro-solid phase extraction. Its different characteristics from conventional sorbents is that it is dissolvable in acidic solution (pH < 4). After the extraction, the analyte elution step was omitted and thus the use of organic solvents was avoided. The key parameters influencing the extraction efficiency such as the amount of sorbent, pH of sample solution, vortex time, type and volume of acidic solution were investigated and optimized. The method exhibited low limits of detection (3.0-5.0 ng/g), good linearity (10-2000 ng/g) with coefficients of determinations higher than 0.9991, acceptable precision (RSD<9.1%) and accuracy (RE<5.8%). The proposed method is fast, efficient, eco-friendly, and suitable for the determination of acidic quinolones in honey samples.

Carbamazepine promotes specific stimuli-induced NLRP3 inflammasome activation and causes idiosyncratic liver injury in mice.

The occurrence of idiosyncratic drug-induced liver injury (IDILI) is a leading cause of post-marketing safety warnings and withdrawals of drugs. Carbamazepine (CBZ), widely used as an antiepileptic agent, could cause rare but severe idiosyncratic liver injury in humans. Although recent studies have shown that inflammasome is implicated in CBZ-induced hepatocellular injury in vitro, the precise pathogenesis of hepatotoxicity remains largely unexplored. Here we report that CBZ causes idiosyncratic liver injury through promoting specific stimuli-induced NLRP3 inflammasome activation. CBZ (40 µM) enhances NLRP3 inflammasome activation triggered by adenosine triphosphate (ATP) or nigericin, rather than SiO2, monosodium urate crystal or intracellular lipopolysaccharide (LPS). In addition, CBZ has no effect on NLRC4 or AIM2 inflammasome activation. Mechanistically, synergistic induction of mitochondrial reactive oxygen species (mtROS) is a crucial event in the enhancement effect of CBZ on ATP- or nigericin-induced NLRP3 inflammasome activation. Moreover, the "C=C" on the seven-membered ring and "C=O" on the nitrogen of CBZ may be contribute to NLRP3 inflammasome hyperactivation and hepatotoxicity. Notably, in vivo data indicate that CBZ (50 mg/kg) causes liver injury in an LPS (2 mg/kg)-mediated susceptibility mouse model of IDILI, accompanied by an increase in caspase-1 activity and IL-1ß production, whereas the combination of CBZ and LPS does not exhibit the effect in NLRP3-knockout mice. In conclusion, CBZ specifically promotes ATP- or nigericin-induced NLRP3 inflammasome activation and causes idiosyncratic liver injury. Our findings also suggest that CBZ may be avoided in patients with NLRP3 inflammasome activation-related diseases that are triggered by ATP or nigericin, which may be risk factors for IDILI.