Development of an in-vitro High-Throughput Screening Assay for the Identification of Modulators of the Blood-Brain Barrier Endothelium Integrity

In Alzheimer’s disease (AD), several studies have reported blood-brain barrier (BBB) breakdown with compromised function. P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) are transport proteins localized at the BBB luminal membrane and play an important role in the clearance of amyloid-β (Aβ). The purpose of this study was to investigate the effect of pharmacological inhibition of Aβ efflux transporters on BBB function and Aβ accumulation and related pathology. Recently, we have developed an in vitro high-throughput screening assay to screen for compounds that modulate the integrity of a cell-based BBB model, which identified elacridar as a disruptor of the monolayer integrity. Elacridar, an investigational compound known for its P-gp and BCRP inhibitory effect and widely used in cancer research. Therefore, it was used as a model compound for further evaluation in a mouse model of AD, namely TgSwDI. TgSwDI mouse is also used as a model for cerebral amyloid angiopathy (CAA). Results showed that P-gp and BCRP inhibition by elacridar disrupted the BBB integrity as measured by increased IgG extravasation and reduced expression of tight junction proteins, increased amyloid deposition due to P-gp, and BCRP downregulation and receptor for advanced glycation end products (RAGE) upregulation, increased CAA and astrogliosis. Further studies revealed the effect was mediated by activation of NF-κB pathway. In conclusion, results suggest that BBB disruption by inhibiting P-gp and BCRP exacerbates AD pathology in a mouse model of AD, and indicate that therapeutic drugs that inhibit P-gp and BCRP could increase the risk for AD.

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DDIS-12. A FACILE, ROBUST AND PREDICTIVE IN VITRO BLOOD-BRAIN-BARRIER MODEL FOR HIGH-THROUGHPUT SCREENING AND DISCOVERY OF BRAIN-PENETRATING AGENTS

Neuro-Oncology ◽

10.1093/neuonc/now212.203 ◽

2016 ◽

Vol 18 (suppl_6) ◽

pp. vi49-vi50

Author(s):

Choi-Fong Cho ◽

Justin Wolfe ◽

Colin Fazden ◽

Kalvis Hornburg ◽

E. Antonio Chiocca ◽

...

Keyword(s):

Blood Brain Barrier ◽

High Throughput ◽

High Throughput Screening ◽

Brain Barrier ◽

Blood Brain ◽

Blood Brain Barrier Model ◽

Barrier Model

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Closing the empty anti-Babesia gibsoni drug pipeline in vitro using fluorescence-based high throughput screening assay

Parasitology International ◽

10.1016/j.parint.2020.102054 ◽

2020 ◽

Vol 75 ◽

pp. 102054 ◽

Cited By ~ 3

Author(s):

Mohamed Abdo Rizk ◽

Shengwei Ji ◽

Mingming Liu ◽

Shimaa Abd El-Salam El-Sayed ◽

Yongchang Li ◽

...

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Babesia Gibsoni ◽

Screening Assay ◽

High Throughput Screening Assay ◽

Drug Pipeline

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An In Vitro Förster Resonance Energy Transfer-Based High-Throughput Screening Assay for Inhibitors of Protein–Protein Interactions in SUMOylation Pathway

Assay and Drug Development Technologies ◽

10.1089/adt.2011.0394 ◽

2012 ◽

Vol 10 (4) ◽

pp. 336-343 ◽

Cited By ~ 11

Author(s):

Yang Song ◽

Jiayu Liao

Keyword(s):

Energy Transfer ◽

High Throughput ◽

Protein Interactions ◽

High Throughput Screening ◽

Resonance Energy Transfer ◽

Resonance Energy ◽

Protein Protein Interactions ◽

Screening Assay ◽

High Throughput Screening Assay

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Establishment of an In Vitro High-Throughput Screening Assay for Detecting Phospholipidosis-Inducing Potential

Toxicological Sciences ◽

10.1093/toxsci/kfj067 ◽

2005 ◽

Vol 90 (1) ◽

pp. 133-141 ◽

Cited By ~ 59

Author(s):

Toshihiko Kasahara ◽

Kazuo Tomita ◽

Hiroyuki Murano ◽

Tsuyoshi Harada ◽

Keisuke Tsubakimoto ◽

...

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Screening Assay ◽

High Throughput Screening Assay

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Lab-on-a-Chip Platforms as Tools for Drug Screening in Neuropathologies Associated with Blood–Brain Barrier Alterations

Biomolecules ◽

10.3390/biom11060916 ◽

2021 ◽

Vol 11 (6) ◽

pp. 916

Author(s):

Cristina Elena Staicu ◽

Florin Jipa ◽

Emanuel Axente ◽

Mihai Radu ◽

Beatrice Mihaela Radu ◽

...

Keyword(s):

Blood Brain Barrier ◽

High Throughput ◽

High Throughput Screening ◽

Biomedical Applications ◽

Neurovascular Unit ◽

Lab On A Chip ◽

Brain Barrier ◽

Blood Brain ◽

Pharmacological Screening ◽

The Brain

Lab-on-a-chip (LOC) and organ-on-a-chip (OOC) devices are highly versatile platforms that enable miniaturization and advanced controlled laboratory functions (i.e., microfluidics, advanced optical or electrical recordings, high-throughput screening). The manufacturing advancements of LOCs/OOCs for biomedical applications and their current limitations are briefly discussed. Multiple studies have exploited the advantages of mimicking organs or tissues on a chip. Among these, we focused our attention on the brain-on-a-chip, blood–brain barrier (BBB)-on-a-chip, and neurovascular unit (NVU)-on-a-chip applications. Mainly, we review the latest developments of brain-on-a-chip, BBB-on-a-chip, and NVU-on-a-chip devices and their use as testing platforms for high-throughput pharmacological screening. In particular, we analyze the most important contributions of these studies in the field of neurodegenerative diseases and their relevance in translational personalized medicine.

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Practical High-Throughput Method to Screen Compounds for Anthelmintic Activity against Caenorhabditis elegans

Molecules ◽

10.3390/molecules26144156 ◽

2021 ◽

Vol 26 (14) ◽

pp. 4156

Author(s):

Aya C. Taki ◽

Joseph J. Byrne ◽

Peter R. Boag ◽

Abdul Jabbar ◽

Robin B. Gasser

Keyword(s):

Caenorhabditis Elegans ◽

Small Molecules ◽

High Throughput ◽

High Throughput Screening ◽

Anthelmintic Activity ◽

Infrared Light ◽

Parasitic Nematodes ◽

Screening Assay ◽

High Throughput Screening Assay

In the present study, we established a practical and cost-effective high throughput screening assay, which relies on the measurement of the motility of Caenorhabditis elegans by infrared light-interference. Using this assay, we screened 14,400 small molecules from the “HitFinder” library (Maybridge), achieving a hit rate of 0.3%. We identified small molecules that reproducibly inhibited the motility of C. elegans (young adults) and assessed dose relationships for a subset of compounds. Future work will critically evaluate the potential of some of these hits as candidates for subsequent optimisation or repurposing as nematocides or nematostats. This high throughput screening assay has the advantage over many previous assays in that it is cost- and time-effective to carry out and achieves a markedly higher throughput (~10,000 compounds per week); therefore, it is suited to the screening of libraries of tens to hundreds of thousands of compounds for subsequent evaluation and development. The present phenotypic whole-worm assay should be readily adaptable to a range of socioeconomically important parasitic nematodes of humans and animals, depending on their dimensions and motility characteristics in vitro, for the discovery of new anthelmintic candidates. This focus is particularly important, given the widespread problems associated with drug resistance in many parasitic worms of livestock animals globally.

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A Repurposing Approach Identifies Off-Patent Drugs with Fungicidal Cryptococcal Activity, a Common Structural Chemotype, and Pharmacological Properties Relevant to the Treatment of Cryptococcosis

Eukaryotic Cell ◽

10.1128/ec.00314-12 ◽

2012 ◽

Vol 12 (2) ◽

pp. 278-287 ◽

Cited By ~ 44

Author(s):

Arielle Butts ◽

Louis DiDone ◽

Kristy Koselny ◽

Bonnie K. Baxter ◽

Yeissa Chabrier-Rosello ◽

...

Keyword(s):

Antifungal Activity ◽

Blood Brain Barrier ◽

High Throughput Screening ◽

Fungicidal Activity ◽

Bioactive Molecules ◽

Brain Barrier ◽

Content Type ◽

Blood Brain ◽

Hydrophobic Moiety ◽

High Throughput Screening Assay

ABSTRACT New, more accessible therapies for cryptococcosis represent an unmet clinical need of global importance. We took a repurposing approach to identify previously developed drugs with fungicidal activity toward Cryptococcus neoformans , using a high-throughput screening assay designed to detect drugs that directly kill fungi. From a set of 1,120 off-patent medications and bioactive molecules, we identified 31 drugs/molecules with fungicidal activity, including 15 drugs for which direct antifungal activity had not previously been reported. A significant portion of the drugs are orally bioavailable and cross the blood-brain barrier, features key to the development of a widely applicable anticryptococcal agent. Structural analysis of this set revealed a common chemotype consisting of a hydrophobic moiety linked to a basic amine, features that are common to drugs that cross the blood-brain barrier and access the phagolysosome, two important niches of C. neoformans . Consistent with their fungicidal activity, the set contains eight drugs that are either additive or synergistic in combination with fluconazole. Importantly, we identified two drugs, amiodarone and thioridazine, with activity against intraphagocytic C. neoformans . Finally, the set of drugs is also enriched for molecules that inhibit calmodulin, and we have confirmed that seven drugs directly bind C. neoformans calmodulin, providing a molecular target that may contribute to the mechanism of antifungal activity. Taken together, these studies provide a foundation for the optimization of the antifungal properties of a set of pharmacologically attractive scaffolds for the development of novel anticryptococcal therapies.

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