Population-based carrier screening and prenatal diagnosis of fragile X syndrome in East Asian populations

Identification of carriers of fragile X syndrome (FXS) with the subsequent prenatal diagnosis and knowledge of FXS-associated genetic profiles are essential for intervention in specific populations. We report the results of carrier screening of 39,458 East Asian adult women and prenatal diagnosis from 87 FXS carriers. The prevalence of FXS carriers and full mutation fetuses was estimated to be 1/581 and 1/3124 in East Asian populations, respectively. We confirmed the validity of the current threshold of CGG trinucleotide repeats for FMR1 categorization; the integral risks of full mutation expansion were approximately 6.0%, 43.8%, and 100% for premutation alleles with 55–74, 75–89, and ≥ 90 CGG repeats, respectively. The protective effect of AGG (adenine-guanine-guanine nucleotides) interruption in East Asian populations was validated, which is important in protecting premutation alleles with 75–89 CGG repeats from full mutation expansion. Finally, family history was shown not an effective indicator for FXS carrier screening in East Asian populations, and population-based screening was more cost-effective. This study provides an insight into the largest carrier screening and prenatal diagnosis for FXS in East Asian populations to date. The FXS-associated genetic profiles of East Asian populations are delineated, and population-based carrier screening is shown to be promising for FXS intervention.     

A Novel Wick-Like Paper-Based Microfluidic Device for 3D Cell Culture and Anti-Cancer Drugs Screening

Paper is increasingly recognized as a portable substrate for cell culture, due to its low-cost, flexible, and special porous property, which provides a native cellular 3D microenvironment. Therefore, paper-based microfluidics has been developed for cell culture and biomedical analysis. However, the inability of continuous medium supply limits the wide application of paper devices for cell culture. Herein, a paper-based microfluidic device is developed with novel folded paper strips as wick-like structure, which is used for medium self-driven perfusion. The paper with patterns of hydrophilic channel, culture areas, and hydrophobic barrier could be easily fabricated through wax-printing. After printing, the hydrophilic paper strip at the periphery of the lower layer is then folded at 90° and extended into the medium container for continuous automatic supply of medium to the cell culture area. Tumor cells cultured in the paper device are tested for anti-cancer drug screening. Visualized cell viability and chemical sensitivity testing can be achieved by colorimetry combined with simple smartphone imaging, effectively reducing precision instrument dependence. The wick paper-based microfluidic device for cell culture endows the method the advantages of lower cost, ease-of-operation, miniaturization, and shows a great potential for large-scale cell culture, antibody drug production, and efficient screening.     

Microengineered systems with iPSC-derived cardiac and hepatic cells to evaluate drug adverse effects

Hepatic and cardiac drug adverse effects are among the leading causes of attrition in drug development programs, in part due to predictive failures of current animal or in vitro models. Hepatocytes and cardiomyocytes differentiated from human induced pluripotent stem cells (iPSCs) hold promise for predicting clinical drug effects, given their human-specific properties and their ability to harbor genetically determined characteristics that underlie inter-individual variations in drug response. Currently, the fetal-like properties and heterogeneity of hepatocytes and cardiomyocytes differentiated from iPSCs make them physiologically different from their counterparts isolated from primary tissues and limit their use for predicting clinical drug effects. To address this hurdle, there have been ongoing advances in differentiation and maturation protocols to improve the quality and use of iPSC-differentiated lineages. Among these are in vitro hepatic and cardiac cellular microsystems that can further enhance the physiology of cultured cells, can be used to better predict drug adverse effects, and investigate drug metabolism, pharmacokinetics, and pharmacodynamics to facilitate successful drug development. In this article, we discuss how cellular microsystems can establish microenvironments for these applications and propose how they could be used for potentially controlling the differentiation of hepatocytes or cardiomyocytes. The physiological relevance of cells is enhanced in cellular microsystems by simulating properties of tissue microenvironments, such as structural dimensionality, media flow, microfluidic control of media composition, and co-cultures with interacting cell types. Recent studies demonstrated that these properties also affect iPSC differentiations and we further elaborate on how they could control differentiation efficiency in microengineered devices. In summary, we describe recent advances in the field of cellular microsystems that can control the differentiation and maturation of hepatocytes and cardiomyocytes for drug evaluation. We also propose how future research with iPSCs within engineered microenvironments could enable their differentiation for scalable evaluations of drug effects.     

新型冠状病毒相关TMPRSS2蛋白结构特征和抗原表位分析

采用生物信息学方法分析预测新型冠状病毒(Severe acute respiratory syndrome coronavirus 2, SARS-CoV-2)跨膜蛋白酶丝氨酸2(transmembrane protease serine 2, TMPRSS2)的理化特性、结构特征和抗原表位,为抗SARS-CoV-2药物研发提供参考。利用ProtParam、ProtScale分析预测TMPRSS2蛋白酶的理化特性;利用COILS Server、SignalP、TMPred、TargetP Server、NetPhos Server、NetNGlyc Server服务器对TMPRSS2蛋白酶结构进行功能结构的分析预测;利用SOPMA、Pfam、SWISS-MODEL分析预测TMPRSS2蛋白酶高级结构;利用IEBD分析预测TMPRSS2蛋白酶B细胞、T细胞表位。TMPRSS2蛋白酶氨基酸组成数为492个,其中丝氨酸占比最高;亲水性较高,含10个跨膜螺旋区;具有4个磷酸化位点,3个糖基化修饰点;二级结构中无规则卷曲占据主导地位,三级结构能与已知的5ce.1.1.A(SMTL ID)模型同源建模;存在13个潜在的B细胞表位,12个得分较高的T细胞表位。     

A novel peptide specifically targeting ovarian cancer identified by in vivo phage display

Discovery of peptide ligands that can target human ovarian cancer and deliver chemotherapeutics offers new opportunity for cancer therapy. The advent of phage‐displayed peptide library facilitated the screening of such peptides. In vivo screening that set in a microanatomic and functional context was applied in our study, and a novel peptide WSGPGVWGASVK targeting ovarian cancer was isolated. The phage clone PC3‐1 displaying peptide WSGPGVWGASVK can gain effective access to accumulate in the tumor sites after intravenous injection while reducing its accumulation in normal organs. Positive immunostaining of PC3‐1 was located in both sites of tumor cells and tumor blood vessels, which resulted in a diffuse binding pattern through the tumor. In vitro study results confirmed the capability of peptide WSGPGVWGASVK binding to and being internalized by both tumor cells and angiogenic endothelial cells. Flow cytometry analysis revealed that the peptide bound to SKOV3 cells with Kd value of 5.43 ± 0.4 μM. Taken together, it suggested that peptide WSGPGVWGASVK is a lead candidate for delivering therapeutics to penetrate into tumors. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Pharmacophore mapping and in silico screening to identify new potent leads for A2A adenosine receptor as antagonists

A_2A adenosine receptor (AR) antagonists play an important role in neurodegenerative diseases like Parkinson’s disease. A 3D-QSAR study of A_2A AR antagonists, was taken up to design best pharmacophore model. The pharmacophoric features (ADHRR) containing a hydrogen bond acceptor (A), a hydrogen bond donor (D), a hydrophobic group (H) and two aromatic rings (R), is projected as the best predictive pharmacophore model. The QSAR model was further treated as a template for in silico search of databases to identify new scaffolds. The binding patterns of the leads with A_2A AR are analysed using docking studies and novel potent ligands of A_2A AR are projected.     

In silico insights into the identification of potential novel angiogenic inhibitors against human VEGFR-2: a new SAR-based hierarchical clustering approach

Abstract

In this study, binding efficiency of new pyrrolopyrimidine structural analogs against human vascular endothelial growth factor receptor-2 (VEGFR-2) were elucidated using integrated in silico methods. Optimized high-resolution model of VEGFR-2 was generated and adopted for structure-based virtual screening approaches. Pyrrolopyrimidine inhibitor (CP15) associated compounds were screened from PubChem database and subjected to virtual screening and comparative docking methods against the receptor ligand-binding domain. Accordingly, high efficient compounds were clustered with similarity indices through PubChem structure cluster module using single-linkage algorithm. Moreover, pharmacokinetics including drug metabolism activities of high-binding leads under investigation was portrayed using ADMET and similarity ensemble analysis. Optimal energy orientations of the selected protein model have been shown to be reliable, and highly recommended for screening and docking studies. Docking and clustering strategies were shown that nineteen candidates as most effective binders for VEGFR-2 than CP15, and are grouped as three classes. Lys⁸⁶⁸, Glu⁸⁸⁵, Cys⁹¹⁹, His¹⁰²⁶, Arg¹⁰²⁷, Asp¹⁰⁴⁶, and Gly¹⁰⁴⁸ residues were predicted as novel hotspot residues, and participate in H-bonds, π-cation, π-stacking, halogen bonds, and salt-bridges formation with ligands. These additional bonds are contributing extent stability that holds the receptor structure at flexible state, this make difficult to any further conformational changes for evoking angiogenic signals. The ADMET and similarity ensemble analysis results were strongly indicated that thirteen candidates as best ligands for angiogenesis targets. Altogether, these findings indicate potential angiogenic templates and their binding levels with VEGFR-2; sorted viewpoints could be useful as a promising way to describe potential angiogenesis inhibitors with related molecular targets.     

Virtual screening of LPXTG competitive SrtA inhibitors targeting signal transduction mechanism in Bacillus anthracis: a combined experimental and theoretical study

Abstract

Members of the sortase enzyme super family decorate the surfaces of Bacillus anthracis cell wall with proteins that play key roles in microbial pathogenesis and its biofilm formation. Bacillus anthracis Sortase-A (Ba-SrtA) is a potential target for new therapeutics as it is required for B. anthracis survival and replication within macrophages. An understanding of the binding site pocket and substrate recognition mechanism by SrtA enzymes may serve to be beneficial in the rational development of sortase inhibitors. Here, the LPXTG signal peptide-based competitive inhibitors are screened against the Ba-SrtA and compounds with reasonable inhibition, specificity, and mechanisms of inactivation of SrtA have been covered. The screened compounds are experimentally validated against the phylogenetically similar Gram-positive pathogen B. cereus. In situ microscopic visualizations suggest that these screened compounds showed the microbial and biofilm inhibitory activity against B. cereus. It facilitates the further development of these molecules into useful anti-infective agents to treat infections caused by B. anthracis and other Gram-positive pathogens. These results provide insight into basic design principles for generating new clinically relevant lead molecules. It also provides an alternative strategy where a screened ligand molecule can be used in combination to battle increasingly against the Gram-positive pathogens.     

Design,synthesis and biological evaluation of novel 1H-1,2,4-triazole,benzothiazole and indazole-based derivatives as potent FGFR1 inhibitors viafragment-based virtual screening

Abstract

Fibroblast growth-factor receptor (FGFR) is a potential target for cancer therapy. We designed three novel series of FGFR1 inhibitors bearing indazole, benzothiazole, and 1H-1,2,4-triazole scaffold via fragment-based virtual screening. All the newly synthesised compounds were evaluated in vitro for their inhibitory activities against FGFR1. Compound 9d bearing an indazole scaffold was first identified as a hit compound, with excellent kinase inhibitory activity (IC₅₀ = 15.0?nM) and modest anti-proliferative activity (IC₅₀ = 785.8?nM). Through two rounds of optimisation, the indazole derivative 9?u stood out as the most potent FGFR1 inhibitors with the best enzyme inhibitory activity (IC₅₀ = 3.3?nM) and cellular activity (IC₅₀ = 468.2?nM). Moreover, 9?u also exhibited good kinase selectivity. In addition, molecular docking study was performed to investigate the binding mode between target compounds and FGFR1.     

Identification of novel CDK2 inhibitors by a multistage virtual screening method based on SVM,pharmacophore and docking model

Abstract

Cyclin-dependent kinase 2 (CDK2) is the family of Ser/Thr protein kinases that has emerged as a highly selective with low toxic cancer therapy target. A multistage virtual screening method combined by SVM, protein-ligand interaction fingerprints (PLIF) pharmacophore and docking was utilised for screening the CDK2 inhibitors. The evaluation of the validation set indicated that this method can be used to screen large chemical databases because it has a high hit-rate and enrichment factor (80.1% and 332.83 respectively). Six compounds were screened out from NCI, Enamine and Pubchem database. After molecular dynamics and binding free energy calculation, two compounds had great potential as novel CDK2 inhibitors and they also showed selective inhibition against CDK2 in the kinase activity assay.