A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance.
Kinase enzymes are an important class of drug targets, particularly in cancer. Cell-based kinase assays are needed to understand how potential kinase inhibitors act on their targets in a physiologically relevant context. Current cell-based kinase assays rely on antibody-based detection of endogenous...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2016-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC5012566?pdf=render |
| _version_ | 1818683458564653056 |
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| author | Steven B Ouellette Brett M Noel Laurie L Parker |
| author_facet | Steven B Ouellette Brett M Noel Laurie L Parker |
| author_sort | Steven B Ouellette |
| collection | DOAJ |
| description | Kinase enzymes are an important class of drug targets, particularly in cancer. Cell-based kinase assays are needed to understand how potential kinase inhibitors act on their targets in a physiologically relevant context. Current cell-based kinase assays rely on antibody-based detection of endogenous substrates, inaccurate disease models, or indirect measurements of drug action. Here we expand on previous work from our lab to introduce a 96-well plate compatible approach for measuring cell-based kinase activity in disease-relevant human chronic myeloid leukemia cell lines using an exogenously added, multi-functional peptide substrate. Our cellular models natively express the BcrAbl oncogene and are either sensitive or have acquired resistance to well-characterized BcrAbl tyrosine kinase inhibitors. This approach measures IC50 values comparable to established methods of assessing drug potency, and its robustness indicates that it can be employed in drug discovery applications. This medium-throughput assay could bridge the gap between single target focused, high-throughput in vitro assays and lower-throughput cell-based follow-up experiments. |
| first_indexed | 2024-12-17T10:35:04Z |
| format | Article |
| id | doaj.art-d5c437747f7e475699631d40e533356a |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-17T10:35:04Z |
| publishDate | 2016-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-d5c437747f7e475699631d40e533356a2022-12-21T21:52:25ZengPublic Library of Science (PLoS)PLoS ONE1932-62032016-01-01119e016174810.1371/journal.pone.0161748A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance.Steven B OuelletteBrett M NoelLaurie L ParkerKinase enzymes are an important class of drug targets, particularly in cancer. Cell-based kinase assays are needed to understand how potential kinase inhibitors act on their targets in a physiologically relevant context. Current cell-based kinase assays rely on antibody-based detection of endogenous substrates, inaccurate disease models, or indirect measurements of drug action. Here we expand on previous work from our lab to introduce a 96-well plate compatible approach for measuring cell-based kinase activity in disease-relevant human chronic myeloid leukemia cell lines using an exogenously added, multi-functional peptide substrate. Our cellular models natively express the BcrAbl oncogene and are either sensitive or have acquired resistance to well-characterized BcrAbl tyrosine kinase inhibitors. This approach measures IC50 values comparable to established methods of assessing drug potency, and its robustness indicates that it can be employed in drug discovery applications. This medium-throughput assay could bridge the gap between single target focused, high-throughput in vitro assays and lower-throughput cell-based follow-up experiments.http://europepmc.org/articles/PMC5012566?pdf=render |
| spellingShingle | Steven B Ouellette Brett M Noel Laurie L Parker A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. PLoS ONE |
| title | A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. |
| title_full | A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. |
| title_fullStr | A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. |
| title_full_unstemmed | A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. |
| title_short | A Cell-Based Assay for Measuring Endogenous BcrAbl Kinase Activity and Inhibitor Resistance. |
| title_sort | cell based assay for measuring endogenous bcrabl kinase activity and inhibitor resistance |
| url | http://europepmc.org/articles/PMC5012566?pdf=render |
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