Solvent selection for insoluble ligands, a challenge for biological assay development: A TNF-α/SPD304 study

Christos P. Papaneophytou; Anthi K. Mettou; Vagelis Rinotas; Eleni Douni; George A. Kontopidis

doi:10.1021/ml300380h

Solvent selection for insoluble ligands, a challenge for biological assay development: A TNF-α/SPD304 study

Christos P. Papaneophytou, Anthi K. Mettou, Vagelis Rinotas, Eleni Douni, George A. Kontopidis

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Abstract

Many active compounds may be excluded from biological assays due to their low aqueous solubility. In this study, a simple method for the determination of the solubility of compounds containing aromatic rings is proposed. In addition to DMSO, five organic solvents for screening experiments of TNF-α inhibitors were explored. DMSO and PEG3350 were the most suitable for both protein stability and ligand-binding experiments. In addition, glycerol is a promising solvent for the screening of other compounds for which it might provide acceptable solubilization, due to its strong tendency to preserve the protein. Moreover, a fluorescence binding assay was developed using the TNF-α/SPD304 system, and a K_d of 5.36 ± 0.21 μM was determined. The results of this study could be used for the future screening of potential TNF-α inhibitors, while the protocols developed in this work could be applied to other proteins.

Original language	English
Pages (from-to)	137-141
Number of pages	5
Journal	ACS Medicinal Chemistry Letters
Volume	4
Issue number	1
DOIs	https://doi.org/10.1021/ml300380h
Publication status	Published - 10 Jan 2013

Keywords

aqueous solubility
fluorescence binding assay
insoluble ligands
protein stability
solvent selection

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10.1021/ml300380h

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title = "Solvent selection for insoluble ligands, a challenge for biological assay development: A TNF-α/SPD304 study",

abstract = "Many active compounds may be excluded from biological assays due to their low aqueous solubility. In this study, a simple method for the determination of the solubility of compounds containing aromatic rings is proposed. In addition to DMSO, five organic solvents for screening experiments of TNF-α inhibitors were explored. DMSO and PEG3350 were the most suitable for both protein stability and ligand-binding experiments. In addition, glycerol is a promising solvent for the screening of other compounds for which it might provide acceptable solubilization, due to its strong tendency to preserve the protein. Moreover, a fluorescence binding assay was developed using the TNF-α/SPD304 system, and a Kd of 5.36 ± 0.21 μM was determined. The results of this study could be used for the future screening of potential TNF-α inhibitors, while the protocols developed in this work could be applied to other proteins.",

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T1 - Solvent selection for insoluble ligands, a challenge for biological assay development

T2 - A TNF-α/SPD304 study

AU - Papaneophytou, Christos P.

AU - Mettou, Anthi K.

AU - Rinotas, Vagelis

AU - Douni, Eleni

AU - Kontopidis, George A.

PY - 2013/1/10

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N2 - Many active compounds may be excluded from biological assays due to their low aqueous solubility. In this study, a simple method for the determination of the solubility of compounds containing aromatic rings is proposed. In addition to DMSO, five organic solvents for screening experiments of TNF-α inhibitors were explored. DMSO and PEG3350 were the most suitable for both protein stability and ligand-binding experiments. In addition, glycerol is a promising solvent for the screening of other compounds for which it might provide acceptable solubilization, due to its strong tendency to preserve the protein. Moreover, a fluorescence binding assay was developed using the TNF-α/SPD304 system, and a Kd of 5.36 ± 0.21 μM was determined. The results of this study could be used for the future screening of potential TNF-α inhibitors, while the protocols developed in this work could be applied to other proteins.

AB - Many active compounds may be excluded from biological assays due to their low aqueous solubility. In this study, a simple method for the determination of the solubility of compounds containing aromatic rings is proposed. In addition to DMSO, five organic solvents for screening experiments of TNF-α inhibitors were explored. DMSO and PEG3350 were the most suitable for both protein stability and ligand-binding experiments. In addition, glycerol is a promising solvent for the screening of other compounds for which it might provide acceptable solubilization, due to its strong tendency to preserve the protein. Moreover, a fluorescence binding assay was developed using the TNF-α/SPD304 system, and a Kd of 5.36 ± 0.21 μM was determined. The results of this study could be used for the future screening of potential TNF-α inhibitors, while the protocols developed in this work could be applied to other proteins.

KW - aqueous solubility

KW - fluorescence binding assay

KW - insoluble ligands

KW - protein stability

KW - solvent selection

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Solvent selection for insoluble ligands, a challenge for biological assay development: A TNF-α/SPD304 study

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Keywords

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