Safety and immunogenicity of NVX-CoV2373 (TAK-019) vaccine in healthy Japanese adults: Interim report of a phase I/II randomized controlled trial

BackgroundWe evaluated the safety and immunogenicity of NVX-CoV2373, a recombinant SARS-CoV-2 nanoparticle vaccine, in healthy Japanese participants.MethodsThis phase 1/2, randomized, observer-blind, placebo-controlled trial conducted in Japan (two sites), enrolled healthy Japanese adults aged ≥ 20 years with no history/risk of SARS-CoV-2 infection and no prior exposure to other approved/investigational SARS-CoV-2 vaccines or treatments. Participants were stratified by age (< 65 or ≥ 65 years) and randomized to receive two doses of either NVX-CoV2373 (5 μg SARS-CoV-2 rS; 50 μg Matrix-M1) or placebo, 21 days apart. Primary outcomes were safety and immunogenicity assessed by serum IgG antibody levels against SARS-CoV-2 rS protein on day 36. Herein, we report the primary data analysis at 4 weeks after the second dose, ahead of 12-month follow-up completion (data cut-off: 8 May 2021).ResultsBetween 12 February 2021 and 17 March 2021, 326 subjects were screened, and 200 participants enrolled and randomized: NVX-CoV2373, n = 150; placebo, n = 50. Solicited adverse events (AEs) through 7 days after each injection occurred in 121/150 (80.7%) and 11/50 (22.0%) participants in the NVX-CoV2373 and placebo arms, respectively. In the NVX-CoV2373 arm, tenderness and injection site pain were the most frequently reported solicited AEs after each vaccination, irrespective of age. Robust immune responses occurred with NVX-CoV2373 (n = 150) by day 36: IgG geometric mean fold rise (95% confidence interval) 259 (219, 306); seroconversion rate 100% (97.6, 100). No such response occurred with placebo (n = 49).ConclusionTwo doses of NVX-CoV2373 given with a 21-day interval demonstrated acceptable safety and induced robust anti-SARS-CoV-2 immune responses in healthy Japanese adults. Funding: Takeda Pharmaceutical Company Limited and Japan Agency for Medical Research and Development (AMED). ClinicalTrials.gov identifier: NCT04712110.     

菊花脑芳樟醇合酶基因CnTPS1的克隆与原核表达分析＊

目的 克隆菊花脑芳樟醇合酶基因CnTPS1的全长编码序列，利用原核系统表达融合蛋白，为进一步研究该基因在菊花脑萜类合成中的功能提供理论依据。方法 以菊花脑基因组数据为基础，设计特异性引物，PCR扩增CnTPS1的全长编码序列，利用生物信息学分析软件分析序列特征。利用qRT-PCR技术分析CnTPS1基因在不同组织中的基因表达量。构建原核表达载体，体外诱导目的蛋白表达。结果 CnTPS1编码序列全长1749bp，编码582个氨基酸；基因表达模式分析表明该基因在茎和管状花中表达量较高；原核表达系统能诱导出67.58kDa大小的蛋白。结论 首次从菊花脑中克隆得到一个芳樟醇合酶基因CnTPS1，运用生物信息学方法对其编码蛋白的理化性质、结构特征等进行了分析预测，分析了该基因的组织表达模式，并在原核表达系统中成功诱导表达出目标蛋白，这些结果将为菊花脑萜类合酶基因的功能以及萜类物质生物合成途径的解析提供理论依据。     

一测多评法在中药质量控制中的应用及研究进展＊

一测多评法（quantitative analysis of multi-components by single marker，QAMS）已成熟应用于中药材、中药提取物及中成药领域等中药复杂体系的质量评价研究中，有效解决了因对照品缺乏而导致的多指标质控技术推广难问题。本文查阅近五年的国内外研究文献，重点总结了高效液相色谱、气相色谱和质谱等技术在QAMS法中药质量控制中的应用。高效液相色谱联用紫外检测器（UV）技术应用最为广泛，最适合于中药QAMS法定量，结果稳定、准确；而高效液相色谱联用蒸发光检测器在中药QAMS法应用中，其测定准确性和适用性仍然需进一步探索和验证。液质联用技术虽然具有灵敏度高、线性范围宽等优点，但易受仪器参数、基质效应等的影响。进一步发展HPLC-UV技术在中成药制剂质量评价中的应用，是QAMS法多成分定量值得关注的方向；探索LC-MS和GC技术在中药QAMS质量评价中的稳定性和普适性是未来将面临的挑战。     

Mechanism of action of Orthosiphon stamineus against non-alcoholic fatty liver disease: Insights from systems pharmacology and molecular docking approaches

Non-alcoholic fatty liver disease (NAFLD) is one of the most common complications of a metabolic syndrome caused by excessive accumulation of fat in the liver. Orthosiphon stamineus also known as Orthosiphon aristatus is a medicinal plant with possible potential beneficial effects on various metabolic disorders. This study aims to investigate the in vitro inhibitory effects of O. stamineus on hepatic fat accumulation and to further use the computational systems pharmacology approach to identify the pharmacokinetic properties of the bioactive compounds of O. stamineus and to predict their molecular mechanisms against NAFLD. Methods: The effects of an ethanolic extract of O. stamineus leaves on cytotoxicity, fat accumulation and antioxidant activity were assessed using HepG2 cells. The bioactive compounds of O. stamineus were identified using LC/MS and two bioinformatics databases, namely the Traditional Chinese Medicine Integrated Database (TCMID) and the Bioinformatics Analysis Tool for the Molecular Mechanism of Traditional Chinese Medicine (BATMAN-TCM). Pathway enrichment analysis was performed on the predicted targets of the bioactive compounds to provide a systematic overview of the molecular mechanism of action, while molecular docking was used to validate the predicted targets. Results: A total of 27 bioactive compounds corresponding to 50 potential NAFLD-related targets were identified. O. stamineus exerts its anti-NAFLD effects by modulating a variety of cellular processes, including oxidative stress, mitochondrial β-oxidation, inflammatory signalling pathways, insulin signalling, and fatty acid homeostasis pathways. O. stamineus is significantly targeting many oxidative stress regulators, including JNK, mammalian target of rapamycin (mTOR), NFKB1, PPAR, and AKT1. Molecular docking analysis confirmed the expected high affinity for the potential targets, while the in vitro assay indicates the ability of O. stamineus to inhibit hepatic fat accumulation. Conclusion: Using the computational systems pharmacology approach, the potentially beneficial effect of O. stamineus in NAFLD was indicated through the combination of multiple compounds, multiple targets, and multicellular components.     

In silico screening and analysis of single-nucleotide polymorphic variants of the ABCC2 gene affecting Dubin–Johnson syndrome

Background and Study AimsDubin–Johnson syndrome (DJS) is a benevolent genetic disorder of the liver with autosomal inheritance. It is a rare disorder characterized by an increase in conjugated bilirubin and anomaly in coproporphyrin clearance. DJS is caused by deleterious mutations in the ABCC2 gene. A polymorphism in the ABCC2 gene causes malfunctions in its ability to regulate the efflux of different organic anions, such as bilirubin, from hepatocytes to the canaliculi. Multidrug resistance protein 2 (MRP2) encoded by the ABCC2 gene is one of the main regulators of the export of bilirubin to respective sites. ABCC2 gene mutations have widely drawn attention in the pathology of DJS in various populations.Patients and MethodsThe ABCC2 gene was subjected to the National Center for Biotechnology Information (NCBI) database in 2020, and non-synonymous single-nucleotide polymorphisms (nsSNPs) and variants in untranslated regions were studied using different computational servers. SIFT, Protein variation effect analyzer, and PolyPhen-2 were used to retrieve the damaging Single-nucleotide polymorphisms (SNPs); PhD-SNP, SNPs&GO, and Protein Analysis Through Evolutionary Relationships were used to predict the association of nsSNPs with DJS; Mutation3D illustrated the location of variants in the protein; SNAP2, MutPred2, ELASPIC, and HOPE were used to predict the structural and functional effects of these mutations on MRP2; and I-mutant 3.0 and MuPro were used to determine the effects of polymorphism on the function of MRP2.ResultsIn this study, 18,947 SNPs were screened from the NCBI database, followed by a series of refinement of variants using online available servers. We concluded that 41 ABCC2 gene variants are vital etiological candidates for DJS in humans. These 41 variants had highly damaging effects on the MRP2 protein, which may lead to deficient transportation capacity, thereby affecting the efflux of bilirubin across the canalicular membrane.ConclusionIn silico tools are an alternative approach for predicting the target SNPs. Hence, previously unreported variants can be considered strong etiological candidates for diseases related to MRP2.     

Immunogenicity and protective efficacy of adenoviral and subunit RSV vaccines based on stabilized prefusion F protein in pre-clinical models

Respiratory Syncytial Virus (RSV) remains a leading cause of severe respiratory disease for which no licensed vaccine is available. We have previously described the derivation of an RSV Fusion protein (F) stabilized in its prefusion conformation (preF) as vaccine immunogen and demonstrated superior immunogenicity in naive mice of preF versus wild type RSV F protein, both as protein and when expressed from an Ad26 vaccine vector. Here we address the question if there are qualitative differences between the two vaccine platforms for induction of protective immunity. In naïve mice, both Ad26.RSV.preF and preF protein induced humoral responses, whereas cellular responses were only elicited by Ad26.RSV.preF. In RSV pre-exposed mice, a single dose of either vaccine induced cellular responses and strong humoral responses. Ad26-induced RSV-specific cellular immune responses were detected systemically and locally in the lungs. Both vaccines showed protective efficacy in the cotton rat model, but Ad26.RSV.preF conferred protection at lower virus neutralizing titers in comparison to RSV preF protein. Factors that may contribute to the protective capacity of Ad26.RSV.preF elicited immunity are the induced IgG2a antibodies that are able to engage Fcγ receptors mediating Antibody Dependent Cellular Cytotoxicity (ADCC), and the induction of systemic and lung resident RSV specific CD8 + T cells. These data demonstrate qualitative improvement of immune responses elicited by an adenoviral vector based vaccine encoding the RSV preF antigen compared to the subunit vaccine in small animal models which may inform RSV vaccine development.