Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors
Promiscuous inhibition due to aggregate formation has been recognized as a major concern in drug discovery campaigns. Here, we report some aggregators identified in a virtual screening (VS) protocol to search for inhibitors of human ecto-5′-nucleotidase (ecto-5′-NT/CD73), a promi...
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MDPI AG
2018-07-01
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| Series: | Molecules |
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| Online Access: | http://www.mdpi.com/1420-3049/23/8/1876 |
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doaj-2be8da1c474d48bca2d8e63868a1ffdf2020-11-24T23:11:34ZengMDPI AGMolecules1420-30492018-07-01238187610.3390/molecules23081876molecules23081876Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase InhibitorsLucas G. Viviani0Erika Piccirillo1Arquimedes Cheffer2Leandro de Rezende3Henning Ulrich4Ana Maria Carmona-Ribeiro5Antonia T.-do Amaral6Departamento de Química Fundamental, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Química Fundamental, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Química Fundamental, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilDepartamento de Química Fundamental, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo 05508-000, BrazilPromiscuous inhibition due to aggregate formation has been recognized as a major concern in drug discovery campaigns. Here, we report some aggregators identified in a virtual screening (VS) protocol to search for inhibitors of human ecto-5′-nucleotidase (ecto-5′-NT/CD73), a promising target for several diseases and pathophysiological events, including cancer, inflammation and autoimmune diseases. Four compounds (A, B, C and D), selected from the ZINC-11 database, showed IC50 values in the micromolar range, being at the same time computationally predicted as potential aggregators. To confirm if they inhibit human ecto-5′-NT via promiscuous mechanism, forming aggregates, enzymatic assays were done in the presence of 0.01% (v/v) Triton X-100 and an increase in the enzyme concentration by 10-fold. Under both experimental conditions, these four compounds showed a significant decrease in their inhibitory activities. To corroborate these findings, turbidimetric assays were performed, confirming that they form aggregate species. Additionally, aggregation kinetic studies were done by dynamic light scattering (DLS) for compound C. None of the identified aggregators has been previously reported in the literature. For the first time, aggregation and promiscuous inhibition issues were systematically studied and evaluated for compounds selected by VS as potential inhibitors for human ecto-5′-NT. Together, our results reinforce the importance of accounting for potential false-positive hits acting by aggregation in drug discovery campaigns to avoid misleading assay results.http://www.mdpi.com/1420-3049/23/8/1876aggregationpromiscuous mechanismhuman ecto-5′-nucleotidasevirtual screeningenzymatic assaysturbidimetrydynamic light scattering |
| collection |
DOAJ |
| language |
English |
| format |
Article |
| sources |
DOAJ |
| author |
Lucas G. Viviani Erika Piccirillo Arquimedes Cheffer Leandro de Rezende Henning Ulrich Ana Maria Carmona-Ribeiro Antonia T.-do Amaral |
| spellingShingle |
Lucas G. Viviani Erika Piccirillo Arquimedes Cheffer Leandro de Rezende Henning Ulrich Ana Maria Carmona-Ribeiro Antonia T.-do Amaral Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors Molecules aggregation promiscuous mechanism human ecto-5′-nucleotidase virtual screening enzymatic assays turbidimetry dynamic light scattering |
| author_facet |
Lucas G. Viviani Erika Piccirillo Arquimedes Cheffer Leandro de Rezende Henning Ulrich Ana Maria Carmona-Ribeiro Antonia T.-do Amaral |
| author_sort |
Lucas G. Viviani |
| title |
Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors |
| title_short |
Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors |
| title_full |
Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors |
| title_fullStr |
Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors |
| title_full_unstemmed |
Be Aware of Aggregators in the Search for Potential Human ecto-5′-Nucleotidase Inhibitors |
| title_sort |
be aware of aggregators in the search for potential human ecto-5′-nucleotidase inhibitors |
| publisher |
MDPI AG |
| series |
Molecules |
| issn |
1420-3049 |
| publishDate |
2018-07-01 |
| description |
Promiscuous inhibition due to aggregate formation has been recognized as a major concern in drug discovery campaigns. Here, we report some aggregators identified in a virtual screening (VS) protocol to search for inhibitors of human ecto-5′-nucleotidase (ecto-5′-NT/CD73), a promising target for several diseases and pathophysiological events, including cancer, inflammation and autoimmune diseases. Four compounds (A, B, C and D), selected from the ZINC-11 database, showed IC50 values in the micromolar range, being at the same time computationally predicted as potential aggregators. To confirm if they inhibit human ecto-5′-NT via promiscuous mechanism, forming aggregates, enzymatic assays were done in the presence of 0.01% (v/v) Triton X-100 and an increase in the enzyme concentration by 10-fold. Under both experimental conditions, these four compounds showed a significant decrease in their inhibitory activities. To corroborate these findings, turbidimetric assays were performed, confirming that they form aggregate species. Additionally, aggregation kinetic studies were done by dynamic light scattering (DLS) for compound C. None of the identified aggregators has been previously reported in the literature. For the first time, aggregation and promiscuous inhibition issues were systematically studied and evaluated for compounds selected by VS as potential inhibitors for human ecto-5′-NT. Together, our results reinforce the importance of accounting for potential false-positive hits acting by aggregation in drug discovery campaigns to avoid misleading assay results. |
| topic |
aggregation promiscuous mechanism human ecto-5′-nucleotidase virtual screening enzymatic assays turbidimetry dynamic light scattering |
| url |
http://www.mdpi.com/1420-3049/23/8/1876 |
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