Abstract：Objective To prepare human monoclonal antibody against spike protein（S protein）of severe acute respiratory syndrome coronavirus 2（SARS⁃CoV⁃2）by using single B cell，and determine its neutralizing activity. Methods Venous blood with high antibody level was collected from people immunized with inactivated SARS⁃CoV⁃2 vaccine（Vero cells） twice，of which peripheral blood mononuclear cells（PBMCs）were isolated by lymphocyte stratified fluid and used to isolate single B cell expressing S protein antibody by magnetic beads coupled with S1 protein. Variable region genes of IgG heavy chain and light chain were amplified by nested PCR after reverse transcription of single B cell，which were connected with CMV promoter，IgG leader sequence，IgG constant region and polyA sequence by overlapping PCR to construct antibody linear expression cassette. Linear expression cassette of the heavy chain and light chain from the same B cell was transfected to HEK293T cells to express human monoclonal antibody of SARS⁃CoV⁃2 S protein. Immunoreactivity was detected by immuno⁃ fluorescence while neutralizing activity by pseudovirus neutralization test. Results A total of 26 monoclonal antibodies against SARS⁃CoV⁃2 S protein were expressed，which showed heavy chain and light chain protein bands of IgG antibody at

This study aimed to investigate the effect and molecular mechanism of sinomenine on proliferation, apoptosis, metastasis, and combination with inhibitors in human hepatocellular carcinoma HepG2 cells and SK-HEP-1 cells. The effect of sinomenine on the growth ability of HepG2 and SK-HEP-1 cells were investigated by CCK-8 assay, colony formation assay, and BeyoClick~(TM) EdU-488 staining. The effect of sinomenine on DNA damage was detected by immunofluorescence assay, and the effect of sinomenine on apoptosis of human hepatocellular carcinoma cells was clarified by Hoechst 33258 staining and CellEvent~(TM) Cystein-3/7Green ReadyProbes~(TM) reagent assay. Cell invasion assay and 3D tumor cell spheroid invasion assay were performed to investigate the effect of sinomenine on the invasion ability of human hepatocellular carcinoma cells in vitro. The effect of sinomenine on the regulation of protein expression related to the protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/signal transducer and activator of transcription 3(STAT3) signaling pathway in HepG2 and SK-HEP-1 cells was examined by Western blot. Molecular docking was used to evaluate the strength of affinity of sinomenine to the target cysteinyl aspartate specific proteinase-3(caspase-3) and STAT3, and combined with CCK-8 assay to detect the changes in cell viability after combination with STAT3 inhibitor JSI-124 in combination with CCK-8 assay. The results showed that sinomenine could significantly reduce the cell viability of human hepatocellular carcinoma cells in a concentration-and time-dependent manner, significantly inhibit the clonogenic ability of human hepatocellular carcinoma cells, and weaken the invasive ability of human hepatocellular carcinoma cells in vitro. In addition, sinomenine could up-regulate the cleaved level of poly ADP-ribose polymerase(PARP), a marker of apoptosis, and down-regulate the protein levels of p-Akt, p-mTOR, and p-STAT3 in human hepatocellular carcinoma cells. Molecular docking results showed that sinomenine had good affinity with the targets caspase-3 and STAT3, and the sensitivity of sinomenine to hepatocellular carcinoma cells was diminished after STAT3 was inhibited. Therefore, sinomenine can inhibit the proliferation and invasion of human hepatocellular carcinoma cells and induce apoptosis, and the mechanism may be attributed to the activation of caspase-3 signaling and inhibition of the Akt/mTOR/STAT3 pathway. This study can provide a new reference for the in-depth research and clinical application of sinomenine and is of great significance to further promote the scientific development and utilization of sinomenine.
Humans ; Carcinoma, Hepatocellular/genetics* ; Proto-Oncogene Proteins c-akt/metabolism* ; Caspase 3/metabolism* ; Liver Neoplasms/genetics* ; Molecular Docking Simulation ; Sincalide/pharmacology* ; Cell Line, Tumor ; Cell Proliferation ; Hep G2 Cells ; TOR Serine-Threonine Kinases/metabolism* ; Apoptosis

Objective: To evaluate the long-term outcomes, failure patterns and prognostic factors of definitive radiotherapy in patients with cervical esophageal carcinoma (CEC). Methods: We retrospectively reviewed the clinical data of 148 CEC patients who treated with definitive radiotherapy in Cancer Hospital of Chinese Academy of Medical Sciences from January 2001 to December 2017. The median radiation dose was 66 Gy (59.4-70 Gy) and 33.1% of patients received concurrent chemotherapy. The Kaplan-Meier method was used to calculate survival rates. The log rank test was used for survival comparison and univariate prognostic analysis. The Cox model was used for multivariate prognostic analysis. Results: The median follow-up time was 102.6 months. The median survival time, 2- and 5-year overall survival (OS) were 22.7 months, 49.9% and 28.3%. The median, 2- and 5-year progression-free survival were 12.6 months, 35.8% and 25.8%. The 2- and 5-year locoregional recurrence-free survival were 59.1% and 50.8%. The 2- and 5-year distant metastases-free survival were 74.6% and 65.9%. Multivariate analysis showed that EQD(2)>66 Gy was the only independent prognostic indicator for OS (P=0.040). The median survival time and 5-year OS rate significantly improved in patients who received EQD(2)>66 Gy than those who received≤66 Gy (31.2 months vs. 19.2 months, 40.1% vs. 19.1%, P=0.027). A total of 87 patients (58.8%) developed tumor progression. There were 50 (33.8%), 23 (15.5%) and 39 (26.4%) patients developed local, regional recurrence and distant metastases, respectively. Eleven patients (7.4%) underwent salvage surgery, and the laryngeal preservation rate for entire group was 93.9%. Conclusions: Definitive radiotherapy is an effective treatment for cervical esophageal carcinoma with the advantage of larynx preservation. Local recurrence is the major failure pattern. EQD(2)>66 Gy is associated with the improved overall survival.
Humans ; Retrospective Studies ; Esophageal Neoplasms/pathology* ; Carcinoma/drug therapy* ; Prognosis ; Treatment Outcome ; Chemoradiotherapy/methods* ; Radiotherapy Dosage

Purpose: We sought to investigate the effectiveness and safety of dabrafenib in children with BRAFV600E-mutated Langerhans cell histiocytosis (LCH). Materials and Methods: A retrospective analysis was performed on 20 children with BRAFV600E-mutated LCH who were treated with dabrafenib. Results: The median age at which the patients started taking dabrafenib was 2.3 years old (range, 0.6 to 6.5 years). The ratio of boys to girls was 2.3:1. The median follow-up time was 30.8 months (range, 18.9 to 43.6 months). There were 14 patients (70%) in the risk organ (RO)+ group and six patients (30%) in the RO– group. All patients were initially treated with traditional chemotherapy and then shifted to targeted therapy due to poor control of LCH or intolerance to chemotherapy. The overall objective response rate and the overall disease control rate were 65% and 75%, respectively. During treatment, circulating levels of cell-free BRAFV600E (cfBRAFV600E) became negative in 60% of the patients within a median period of 3.0 months (range, 1.0 to 9.0 months). Grade 2 or 3 adverse effects occurred in five patients. Conclusion Some children with BRAFV600E-mutated LCH may benefit from monotherapy with dabrafenib, especially high-risk patients with concomitant hemophagocytic lymphohistiocytosis and intolerance to chemotherapy. The safety of dabrafenib is notable. A prospective study with a larger sample size is required to determine the optimal dosage and treatment duration.

A bioassay method for inhibiting platelet aggregation in vitro was established to quantify the pharmacological effects of Compound Danshen Tablets and support its quality control. The inhibition of platelet aggregation in rabbit plasma in vitro by Compound Danshen Tablets was used as the experimental system. The titer was calculated by using the method of dose-response parallel lines. As a result, a bioassay for the inhibition of platelet aggregation in vitro by Compound Danshen Tablets was established. Linearity was good in the concentration range of 0.128 g·mL^-1 to 0.205 g·mL^-1, and the titer of standard Compound Danshen tablets was 7 659 U·g^-1 according to the titer definition. This in vitro assay was simple, reliable, reproducible and convenient. The activity of Compound Danshen Tablets in inhibiting platelet aggregation was quantified by potency assay and the quality of different batches was evaluated. The method can be applied for the quality control of Compound Danshen Tablets.

In 2013,China pharmaceutical regulatory department issued guiduance on postmarketing drug safety monitoring for industry. It aimed to encourage industry to carry out postmarketing drug safety monitoring including hospital-based intensive monitoring of postmarketing Chinese patent medicine. Subsequently,more and more such kind of studies have been performed all over China. However,in view of the current situation in this field,the development of hospital-based intensive monitoring of postmarketing Chinese patent medicine lacks standardization,such as unreasonable design,omission of reports about adverse drug reactions,inadequate process of quality control,non-standardized interpretation of adverse reactions,etc. Therefore,it is necessary to formulate relevant technical specifications to guide this area. The developing of this technical specification refered to the international post-marketing safety monitoring model and advanced design concepts and methods. We developed it under the guidance of relevant laws,regulations and technical documents in China. Meanwhile the characteristics of Chinese patent medicines and the real situation in this area were considered. The aim of this technical specification is to obtain the incidence,type,degree and clinical manifestation of adverse drug reactions of Chinese patent medicines,to find new risk signals of adverse reactions,to identify risk factors,and to provide a basis for the formulation of risk management and control plans. This specification has been approved by China association of Chinese medicine which is numbered T/CACM011-2016.
Adverse Drug Reaction Reporting Systems ; China ; Hospitals ; Medicine, Chinese Traditional ; Nonprescription Drugs ; Product Surveillance, Postmarketing

OBJECTIVE: To investigate the characteristic changes of the plasma cytokine profile in Chinese patients with idiopathic multicentric Castleman diseases (iMCD). METHODS: The plasma samples from 22 patients with confirmed diagnosis of iMCD were collected before treatments; Specimens from 17 patients with newly diagnosed multiple myeloma, 10 non Hodgkin's lymphoma, and 15 healthy donors were used as control. Seventeen kinds of cytokines were measured by cytokine beads array (CBA) and ELISA respectively. RESULTS: Six cytokines were measured by ELISA. The concentrations of IL-2, IL-6, IL-21 and VEGF were significantly higher in the plasma of iMCD patients than those of the healthy donors (P＜0.01) and the level of IL-21 was highest in the iMCD group. There was no significant difference in the levels of IL-1β and IL-4 between the iMCD and healthy donor groups. Thirteen cytokines were measured by CBA assay, besides IL-6 level was confirmed to be higher in iMCD group than that in healthy controls (P＜0.01）, IL-12-p70 and IL-33 levels were also higher in iMCD group than those in control group (P＜0.05）, no significant difference of the rest cytokines was found between iMCD and the control group. CONCLUSION IL-6 and VEGF has shown to involved in the pathogenesis of iMCD, the results of preliminary study imply the role of IL-2 、IL-21、IL-12-p70 and IL-33 in this rare lymphoproliferative disease. Further studies are needed to elucidate the mechanism of these cytokines, which may shed some light on the identification of novel therapeutic targets against iMCD.
Castleman Disease ; Cytokines ; Humans ; Interleukin-12 ; Interleukin-1beta ; Plasma

OBJECTIVE: To investigate the use of antibiotics in children with community-acquired pneumonia (CAP) in multiple regions of China, and to provide a reference for CAP standard treatment and rational antibiotic use in children. METHODS: The medical data of 1 383 children with CAP who were hospitalized in the department of pediatrics in 10 grade A tertiary hospitals from 9 cities between April 14, 2014 and January 1, 2016 were reviewed, to analyze the status of antibiotic use in hospitalized children in North China, Northeast China, East China, and South China. RESULTS: The overall rate of antibiotic use in children with CAP was 89.08%, with 88.7% in North China, 95.5% in Northeast China, 83.3% in East China, and 86.6% in South China. The main types of antibiotics used were cephalosporins, macrolides, compound preparations of β-lactam antibiotics, polyphosphoric broad-spectrum antibiotics and other β-lactam antibiotics. The selection of antibiotics was generally rational, but antibiotics were still used in some patients with viral infection alone or a combined use of ≥2 kinds of antibiotics were noted in some patients with infection caused by one kind of pathogen. Irrational antibiotic use was observed in 131 children (10.63%). CONCLUSIONS There are high rates of antibiotic use and irrational use of antibiotics among children with CAP. Standard management of antibiotic use in children with CAP should be strengthened.
Anti-Bacterial Agents ; therapeutic use ; Child ; Child, Hospitalized ; China ; Community-Acquired Infections ; drug therapy ; Humans

Objective To evaluate endoscopic ultrasonography (EUS) in diagnosis of gastric submucosal tumors (SMTs), and analyze the characteristics of gastric SMTs under EUS. Methods Clinical data of 614 patients with gastric SMTs, who were evaluated by EUS and underwent endoscopic submucosal dissection (ESD) from September 2008 to December 2016, were retrospectively analyzed. The golden standard for lession origins was the intraoperative diagnosis of ESD, and that for pathological types was the combination of postoperative pathological and immunohistochemical findings. The consistency of diagnosis of EUS was evaluated, and the characteristics of lesions under EUS were analyzed. Results The total consistency in diagnosing lesion origins was 91.25％ between EUS and intraoperative results of ESD, and the consistency in diagnosing lesion originated from the muscularis mucosa, submucosa and muscularis propria was 66.67％, 80.85％ and 94.50％, respectively. The total consistency in pathological types was 65.99％ between EUS and postoperative pathological results, and the consistency of gastrointestinal stromal tumor (GIST), leiomyoma, ectopic pancreas and lipoma was 91.85％, 18.56％, 79.76％ and 90.70％, respectively. Conclusion EUS can initially determine the origins and types of gastric SMTs, providing a more accurate basis for endoscopic treatment, but there were some limitations on the diagnosis of leiomyoma and some rare lesions such as hamartoma, inflammatory fibrous polyps, carcinoid, fibroma, etc. Thus, if necessary, the lesion should be removed so as to make a definite diagnosis and prevent malignant change.

OBJECTIVETo investigate the relationship between inflammatory factors and two Chinese medicine (CM) syndrome types of qi stagnation and blood stasis (QSBS) and qi deficiency and blood stasis (QDBS) in patients with acute coronary syndrome (ACS).

METHODSSixty subjects with ACS, whose pathogenesis changes belongs to qi disturbance blood stasis syndrome, were divided into 2 groups: 30 in the QSBS group and 30 in the QDBS group. The comparative analysis on them was carried out through comparing general information, coronary angiography and inflammatory factors including intracellular adhesion molecule-1 (ICAM-1), chitinase-3-like protein 1 (YKL-40) and lipoprotein-associated phospholipase A2 (Lp-PLA2).

RESULTSCompared with the QSBS group, Lp-PLA2 and YKL-40 levels in the QDBS group showed no-significant difference (P>0.05); ICAM-1 was significantly higher in the QDBS group than in the QSBS group in the pathological processes of qi disturbance and blood stasis syndrome of ACS (P<0.05).

CONCLUSIONSInflammatory factor ICAM-1 may be an objective basis for syndrome typing of QSBS and QDBS, which provides a research direction for standardization research of CM syndrome types.