Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery.
New assay designs are needed to improve the predictive value of the Trypanosoma cruzi in vitro tests used as part of the Chagas' disease drug development pipeline. Here, we employed a green fluorescent protein (eGFP)-expressing parasite line and live high-content imaging to monitor the growth o...
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ftdoajarticles:oai:doaj.org/article:7b65573ff19249218f435af9c19dad98 2023-05-15T15:13:31+02:00 Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. Anna F Fesser Olivier Braissant Francisco Olmo John M Kelly Pascal Mäser Marcel Kaiser 2020-07-01T00:00:00Z https://doi.org/10.1371/journal.pntd.0008487 https://doaj.org/article/7b65573ff19249218f435af9c19dad98 EN eng Public Library of Science (PLoS) https://doi.org/10.1371/journal.pntd.0008487 https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0008487 https://doaj.org/article/7b65573ff19249218f435af9c19dad98 PLoS Neglected Tropical Diseases, Vol 14, Iss 7, p e0008487 (2020) Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 article 2020 ftdoajarticles https://doi.org/10.1371/journal.pntd.0008487 2022-12-31T11:51:07Z New assay designs are needed to improve the predictive value of the Trypanosoma cruzi in vitro tests used as part of the Chagas' disease drug development pipeline. Here, we employed a green fluorescent protein (eGFP)-expressing parasite line and live high-content imaging to monitor the growth of T. cruzi amastigotes in mouse embryonic fibroblasts. A novel assay design allowed us to follow parasite numbers over 6 days, in four-hour intervals, while occupying the microscope for only 24 hours per biological replicate. Dose-response curves were calculated for each time point after addition of test compounds, revealing how EC50 values first decreased over the time of drug exposure, and then leveled off. However, we observed that parasite numbers could vary, even in the untreated controls, and at different sites in the same well, which caused variability in the EC50 values. To overcome this, we established that fold change in parasite number per hour is a more robust and informative measure of drug activity. This was calculated based on an exponential growth model for every biological sample. The net fold change per hour is the result of parasite replication, differentiation, and death. The calculation of this fold change enabled us to determine the tipping point of drug action, i.e. the time point when the death rate of the parasites exceeded the growth rate and the fold change dropped below 1, depending on the drug concentration and exposure time. This revealed specific pharmacodynamic profiles of the benchmark drugs benznidazole and posaconazole. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic PLOS Neglected Tropical Diseases 14 7 e0008487 |
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Open Polar |
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Directory of Open Access Journals: DOAJ Articles |
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ftdoajarticles |
language |
English |
topic |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
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Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 Anna F Fesser Olivier Braissant Francisco Olmo John M Kelly Pascal Mäser Marcel Kaiser Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
description |
New assay designs are needed to improve the predictive value of the Trypanosoma cruzi in vitro tests used as part of the Chagas' disease drug development pipeline. Here, we employed a green fluorescent protein (eGFP)-expressing parasite line and live high-content imaging to monitor the growth of T. cruzi amastigotes in mouse embryonic fibroblasts. A novel assay design allowed us to follow parasite numbers over 6 days, in four-hour intervals, while occupying the microscope for only 24 hours per biological replicate. Dose-response curves were calculated for each time point after addition of test compounds, revealing how EC50 values first decreased over the time of drug exposure, and then leveled off. However, we observed that parasite numbers could vary, even in the untreated controls, and at different sites in the same well, which caused variability in the EC50 values. To overcome this, we established that fold change in parasite number per hour is a more robust and informative measure of drug activity. This was calculated based on an exponential growth model for every biological sample. The net fold change per hour is the result of parasite replication, differentiation, and death. The calculation of this fold change enabled us to determine the tipping point of drug action, i.e. the time point when the death rate of the parasites exceeded the growth rate and the fold change dropped below 1, depending on the drug concentration and exposure time. This revealed specific pharmacodynamic profiles of the benchmark drugs benznidazole and posaconazole. |
format |
Article in Journal/Newspaper |
author |
Anna F Fesser Olivier Braissant Francisco Olmo John M Kelly Pascal Mäser Marcel Kaiser |
author_facet |
Anna F Fesser Olivier Braissant Francisco Olmo John M Kelly Pascal Mäser Marcel Kaiser |
author_sort |
Anna F Fesser |
title |
Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
title_short |
Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
title_full |
Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
title_fullStr |
Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
title_full_unstemmed |
Non-invasive monitoring of drug action: A new live in vitro assay design for Chagas' disease drug discovery. |
title_sort |
non-invasive monitoring of drug action: a new live in vitro assay design for chagas' disease drug discovery. |
publisher |
Public Library of Science (PLoS) |
publishDate |
2020 |
url |
https://doi.org/10.1371/journal.pntd.0008487 https://doaj.org/article/7b65573ff19249218f435af9c19dad98 |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
PLoS Neglected Tropical Diseases, Vol 14, Iss 7, p e0008487 (2020) |
op_relation |
https://doi.org/10.1371/journal.pntd.0008487 https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0008487 https://doaj.org/article/7b65573ff19249218f435af9c19dad98 |
op_doi |
https://doi.org/10.1371/journal.pntd.0008487 |
container_title |
PLOS Neglected Tropical Diseases |
container_volume |
14 |
container_issue |
7 |
container_start_page |
e0008487 |
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1766344063291752448 |