Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response
Common alkylating antitumor drugs, such as temozolomide, trigger their cytotoxicity by methylating the O6-position of guanosine in DNA. However, the therapeutic effect of these drugs is dampened by elevated levels of the DNA repair enzyme, O6-methylguanine DNA methyltransferase (MGMT), which directl...
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| Format: | Article |
| Language: | en_US |
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Public Library of Science
2017
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| Online Access: | http://hdl.handle.net/1721.1/108301 https://orcid.org/0000-0002-7112-1454 |
| _version_ | 1826209123280093184 |
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| author | Beharry, Andrew A. Kool, Eric T. Samson, Leona D Nagel, Zachary D. |
| author2 | Massachusetts Institute of Technology. Department of Biological Engineering |
| author_facet | Massachusetts Institute of Technology. Department of Biological Engineering Beharry, Andrew A. Kool, Eric T. Samson, Leona D Nagel, Zachary D. |
| author_sort | Beharry, Andrew A. |
| collection | MIT |
| description | Common alkylating antitumor drugs, such as temozolomide, trigger their cytotoxicity by methylating the O6-position of guanosine in DNA. However, the therapeutic effect of these drugs is dampened by elevated levels of the DNA repair enzyme, O6-methylguanine DNA methyltransferase (MGMT), which directly reverses this alkylation. As a result, assessing MGMT levels in patient samples provides an important predictor of therapeutic response; however, current methods available to measure this protein are indirect, complex and slow. Here we describe the design and synthesis of fluorescent chemosensors that report directly on MGMT activity in a single step within minutes. The chemosensors incorporate a fluorophore and quencher pair, which become separated by the MGMT dealkylation reaction, yielding light-up responses of up to 55-fold, directly reflecting repair activity. Experiments show that the best-performing probe retains near-native activity at mid-nanomolar concentrations. A nuclease-protected probe, NR-1, was prepared and tested in tumor cell lysates, demonstrating an ability to evaluate relative levels of MGMT repair activity in twenty minutes. In addition, a probe was employed to evaluate inhibitors of MGMT, suggesting utility for discovering new inhibitors in a high-throughput manner. Probe designs such as that of NR-1 may prove valuable to clinicians in selection of patients for alkylating drug therapies and in assessing resistance that arises during treatment. |
| first_indexed | 2024-09-23T14:17:55Z |
| format | Article |
| id | mit-1721.1/108301 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T14:17:55Z |
| publishDate | 2017 |
| publisher | Public Library of Science |
| record_format | dspace |
| spelling | mit-1721.1/1083012022-09-28T19:52:25Z Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response Beharry, Andrew A. Kool, Eric T. Samson, Leona D Nagel, Zachary D. Massachusetts Institute of Technology. Department of Biological Engineering Massachusetts Institute of Technology. Department of Biology Samson, Leona D Nagel, Zachary D. Common alkylating antitumor drugs, such as temozolomide, trigger their cytotoxicity by methylating the O6-position of guanosine in DNA. However, the therapeutic effect of these drugs is dampened by elevated levels of the DNA repair enzyme, O6-methylguanine DNA methyltransferase (MGMT), which directly reverses this alkylation. As a result, assessing MGMT levels in patient samples provides an important predictor of therapeutic response; however, current methods available to measure this protein are indirect, complex and slow. Here we describe the design and synthesis of fluorescent chemosensors that report directly on MGMT activity in a single step within minutes. The chemosensors incorporate a fluorophore and quencher pair, which become separated by the MGMT dealkylation reaction, yielding light-up responses of up to 55-fold, directly reflecting repair activity. Experiments show that the best-performing probe retains near-native activity at mid-nanomolar concentrations. A nuclease-protected probe, NR-1, was prepared and tested in tumor cell lysates, demonstrating an ability to evaluate relative levels of MGMT repair activity in twenty minutes. In addition, a probe was employed to evaluate inhibitors of MGMT, suggesting utility for discovering new inhibitors in a high-throughput manner. Probe designs such as that of NR-1 may prove valuable to clinicians in selection of patients for alkylating drug therapies and in assessing resistance that arises during treatment. National Institutes of Health (U.S.) (DP1-ES 022576) 2017-04-20T17:51:44Z 2017-04-20T17:51:44Z 2016-04 2015-12 Article http://purl.org/eprint/type/JournalArticle 1932-6203 http://hdl.handle.net/1721.1/108301 Beharry, Andrew A., Zachary D. Nagel, Leona D. Samson, and Eric T. Kool. “Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response.” Edited by Robert W Sobol. PLoS ONE 11, no. 4 (April 1, 2016): e0152684. https://orcid.org/0000-0002-7112-1454 en_US http://dx.doi.org/10.1371/journal.pone.0152684 PLOS ONE Creative Commons Attribution http://creativecommons.org/licenses/by/4.0/ application/pdf Public Library of Science Public Library of Science |
| spellingShingle | Beharry, Andrew A. Kool, Eric T. Samson, Leona D Nagel, Zachary D. Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title | Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title_full | Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title_fullStr | Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title_full_unstemmed | Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title_short | Fluorogenic Real-Time Reporters of DNA Repair by MGMT, a Clinical Predictor of Antitumor Drug Response |
| title_sort | fluorogenic real time reporters of dna repair by mgmt a clinical predictor of antitumor drug response |
| url | http://hdl.handle.net/1721.1/108301 https://orcid.org/0000-0002-7112-1454 |
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