The Use of Genetically Engineered Cell-Based Assays in in-vitro Drug Discovery

2007 ◽  
pp. 247-262
Keyword(s):  
Drug Discovery ◽  
Genetically Engineered

scholarly journals Identification of a proteasome-targeting arylsulfonamide with potential for the treatment of Chagas’ disease

2021 ◽  
Author(s):  
Marta Lima ◽  
Lindsay B Tulloch ◽  
Victoriano Corpas-Lopez ◽  
Sandra Carvalho ◽  
Richard J. Wall ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Causative Agent ◽  
Mode Of Action ◽  
Malic Enzyme ◽  
Proteasome Inhibitors ◽  
Genetically Engineered ◽  
Phenotypic Screening

Phenotypic screening identified an arylsulfonamide compound with activity against Trypanosoma cruzi , the causative agent of Chagas’ disease. Comprehensive mode of action studies revealed that this compound primarily targets the T. cruzi proteasome, binding at the interface between β4 and β5 subunits that catalyse chymotrypsin-like activity. A mutation in the β5 subunit of the proteasome was associated with resistance to compound 1 , while overexpression of this mutated subunit also reduced susceptibility to compound 1 . Further genetically engineered and in vitro selected clones resistant to proteasome inhibitors known to bind at the β4/β5 interface were cross-resistant to compound 1 . Ubiquitinylated proteins were additionally found to accumulate in compound 1 -treated epimastigotes. Finally, thermal proteome profiling identified malic enzyme as a secondary target of compound 1 , although malic enzyme inhibition was not found to drive potency. These studies identify a novel pharmacophore capable of inhibiting the T. cruzi proteasome that may be exploitable for anti-chagasic drug discovery.

scholarly journals Development of a miniaturized 3D organoid culture platform for ultra-high-throughput screening

2020 ◽  
Vol 12 (8) ◽  
pp. 630-643 ◽  
Author(s):  
Yuhong Du ◽  
Xingnan Li ◽  
Qiankun Niu ◽  
Xiulei Mo ◽  
Min Qui ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Large Scale ◽  
Genetically Engineered ◽  
High Density ◽  
Organoid Culture ◽  
3D Architecture

Abstract The recent advent of robust methods to grow human tissues as 3D organoids allows us to recapitulate the 3D architecture of tumors in an in vitro setting and offers a new orthogonal approach for drug discovery. However, organoid culturing with extracellular matrix to support 3D architecture has been challenging for high-throughput screening (HTS)-based drug discovery due to technical difficulties. Using genetically engineered human colon organoids as a model system, here we report our effort to miniaturize such 3D organoid culture with extracellular matrix support in high-density plates to enable HTS. We first established organoid culturing in a 384-well plate format and validated its application in a cell viability HTS assay by screening a 2036-compound library. We further miniaturized the 3D organoid culturing in a 1536-well ultra-HTS format and demonstrated its robust performance for large-scale primary compound screening. Our miniaturized organoid culturing method may be adapted to other types of organoids. By leveraging the power of 3D organoid culture in a high-density plate format, we provide a physiologically relevant screening platform to model tumors to accelerate organoid-based research and drug discovery.

Effect of C6-Volatiles on Bioluminescent Plant Pathogens

HortScience ◽  
1998 ◽  
Vol 33 (3) ◽  
pp. 557d-557
Author(s):  
Jennifer Warr ◽  
Fenny Dane ◽  
Bob Ebel
Keyword(s):  
Biological Activity ◽  
Bacterial Growth ◽  
Xanthomonas Campestris ◽  
Plant Pathogens ◽  
Genetically Engineered ◽  
The Other ◽  
Computer Assisted ◽  
Signal Molecules ◽  
Low Concentrations

C6 volatile compounds are known to be produced by the plant upon pathogen attack or other stress-related events. The biological activity of many of these substances is poorly understood, but some might produce signal molecules important in host–pathogen interactions. In this research we explored the possibility that lipid-derived C6 volatiles have a direct effect on bacterial plant pathogens. To this purpose we used a unique tool, a bacterium genetically engineered to bioluminesce. Light-producing genes from a fish-associated bacterium were introduced into Xanthomonas campestris pv. campestris, enabling nondestructive detection of bacteria in vitro and in the plant with special computer-assisted camera equipment. The effects of different C6 volatiles (trans-2 hexanal, trans-2 hexen-1-ol and cis-3 hexenol) on growth of bioluminescent Xanthomonas campestris were investigated. Different volatile concentrations were used. Treatment with trans-2 hexanal appeared bactericidal at low concentrations (1% and 10%), while treatments with the other volatiles were not inhibitive to bacterial growth. The implications of these results with respect to practical use of trans-2 hexanal in pathogen susceptible and resistant plants will be discussed.

Current Approaches to Drug Discovery for Chagas Disease: Methodological Advances

2019 ◽  
Vol 22 (8) ◽  
pp. 509-520
Author(s):  
Cauê B. Scarim ◽  
Chung M. Chin
Keyword(s):  
Drug Discovery ◽  
Chagas Disease ◽  
Drug Candidates ◽  
Lead Compounds ◽  
New Drug ◽  
Compound Libraries ◽  
Screening Assays ◽  
Cruzi Infection

Background: In recent years, there has been an improvement in the in vitro and in vivo methodology for the screening of anti-chagasic compounds. Millions of compounds can now have their activity evaluated (in large compound libraries) by means of high throughput in vitro screening assays. Objective: Current approaches to drug discovery for Chagas disease. Method: This review article examines the contribution of these methodological advances in medicinal chemistry in the last four years, focusing on Trypanosoma cruzi infection, obtained from the PubMed, Web of Science, and Scopus databases. Results: Here, we have shown that the promise is increasing each year for more lead compounds for the development of a new drug against Chagas disease. Conclusion: There is increased optimism among those working with the objective to find new drug candidates for optimal treatments against Chagas disease.

scholarly journals Ultra-strong bio-glue from genetically engineered polypeptides

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Chao Ma ◽  
Jing Sun ◽  
Bo Li ◽  
Yang Feng ◽  
Yao Sun ◽  
...  
Keyword(s):  
Soft Tissues ◽  
Covalent Bond ◽  
Genetically Engineered ◽  
Supramolecular Interactions ◽  
Molar Ratio ◽  
High Adhesion ◽  
Strong Adhesion ◽  
Order Of Magnitude

AbstractThe development of biomedical glues is an important, yet challenging task as seemingly mutually exclusive properties need to be combined in one material, i.e. strong adhesion and adaption to remodeling processes in healing tissue. Here, we report a biocompatible and biodegradable protein-based adhesive with high adhesion strengths. The maximum strength reaches 16.5 ± 2.2 MPa on hard substrates, which is comparable to that of commercial cyanoacrylate superglue and higher than other protein-based adhesives by at least one order of magnitude. Moreover, the strong adhesion on soft tissues qualifies the adhesive as biomedical glue outperforming some commercial products. Robust mechanical properties are realized without covalent bond formation during the adhesion process. A complex consisting of cationic supercharged polypeptides and anionic aromatic surfactants with lysine to surfactant molar ratio of 1:0.9 is driven by multiple supramolecular interactions enabling such strong adhesion. We demonstrate the glue’s robust performance in vitro and in vivo for cosmetic and hemostasis applications and accelerated wound healing by comparison to surgical wound closures.

scholarly journals Rapid target validation in a Cas9-inducible hiPSC derived kidney model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yasaman Shamshirgaran ◽  
Anna Jonebring ◽  
Anna Svensson ◽  
Isabelle Leefa ◽  
Mohammad Bohlooly-Y ◽  
...  
Keyword(s):  
High Efficiency ◽  
Disease Modeling ◽  
Genetic Manipulation ◽  
Disease Model ◽  
Genetically Engineered ◽  
Model Systems ◽  
Target Validation ◽  
Direct Differentiation ◽  
Kidney Organoids

AbstractRecent advances in induced pluripotent stem cells (iPSCs), genome editing technologies and 3D organoid model systems highlight opportunities to develop new in vitro human disease models to serve drug discovery programs. An ideal disease model would accurately recapitulate the relevant disease phenotype and provide a scalable platform for drug and genetic screening studies. Kidney organoids offer a high cellular complexity that may provide greater insights than conventional single-cell type cell culture models. However, genetic manipulation of the kidney organoids requires prior generation of genetically modified clonal lines, which is a time and labor consuming procedure. Here, we present a methodology for direct differentiation of the CRISPR-targeted cell pools, using a doxycycline-inducible Cas9 expressing hiPSC line for high efficiency editing to eliminate the laborious clonal line generation steps. We demonstrate the versatile use of genetically engineered kidney organoids by targeting the autosomal dominant polycystic kidney disease (ADPKD) genes: PKD1 and PKD2. Direct differentiation of the respective knockout pool populations into kidney organoids resulted in the formation of cyst-like structures in the tubular compartment. Our findings demonstrated that we can achieve > 80% editing efficiency in the iPSC pool population which resulted in a reliable 3D organoid model of ADPKD. The described methodology may provide a platform for rapid target validation in the context of disease modeling.

scholarly journals Arrayed CRISPR reveals genetic regulators of tau aggregation, autophagy and mitochondria in Alzheimer’s disease model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lishu Duan ◽  
Mufeng Hu ◽  
Joseph A. Tamm ◽  
Yelena Y. Grinberg ◽  
Fang Shen ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Drug Discovery ◽  
Disease Model ◽  
Mitochondrial Morphology ◽  
Individual Gene ◽  
Human Genes ◽  
Future Drug ◽  
Tau Aggregation

AbstractAlzheimer’s disease (AD) is a common neurodegenerative disease with poor prognosis. New options for drug discovery targets are needed. We developed an imaging based arrayed CRISPR method to interrogate the human genome for modulation of in vitro correlates of AD features, and used this to assess 1525 human genes related to tau aggregation, autophagy and mitochondria. This work revealed (I) a network of tau aggregation modulators including the NF-κB pathway and inflammatory signaling, (II) a correlation between mitochondrial morphology, respiratory function and transcriptomics, (III) machine learning predicted novel roles of genes and pathways in autophagic processes and (IV) individual gene function inferences and interactions among biological processes via multi-feature clustering. These studies provide a platform to interrogate underexplored aspects of AD biology and offer several specific hypotheses for future drug discovery efforts.

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