A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin
Standard cell proliferation assays use bulk media drug concentration to ascertain the potency of chemotherapeutic drugs; however, the relevant quantity is clearly the amount of drug actually taken up by the cell. To address this discrepancy, we have developed a flow cytometric clonogenic assay to co...
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| Format: | Article |
| Language: | en_US |
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Wiley Blackwell
2017
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| Online Access: | http://hdl.handle.net/1721.1/107005 https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0002-5568-6455 https://orcid.org/0000-0003-2398-5896 |
| _version_ | 1826207045290819584 |
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| author | Kulkarni, Chethana Betts, Alison M. Maass, Katie F. Quadir, Mohiuddin Abdul Hammond, Paula T. Wittrup, Karl Dane |
| author2 | Massachusetts Institute of Technology. Department of Biological Engineering |
| author_facet | Massachusetts Institute of Technology. Department of Biological Engineering Kulkarni, Chethana Betts, Alison M. Maass, Katie F. Quadir, Mohiuddin Abdul Hammond, Paula T. Wittrup, Karl Dane |
| author_sort | Kulkarni, Chethana |
| collection | MIT |
| description | Standard cell proliferation assays use bulk media drug concentration to ascertain the potency of chemotherapeutic drugs; however, the relevant quantity is clearly the amount of drug actually taken up by the cell. To address this discrepancy, we have developed a flow cytometric clonogenic assay to correlate the amount of drug in a single cell with the cell’s ability to proliferate using a cell tracing dye and doxorubicin, a naturally fluorescent chemotherapeutic drug. By varying doxorubicin concentration in the media, length of treatment time, and treatment with verapamil, an efflux pump inhibitor, we introduced 10[superscript 5]–10[superscript 10] doxorubicin molecules per cell; then used a dye-dilution assay to simultaneously assess the number of cell divisions. We find that a cell’s ability to proliferate is a surprisingly conserved function of the number of intracellular doxorubicin molecules, resulting in single-cell IC[subscript 50] values of 4–12 million intracellular doxorubicin molecules. The developed assay is a straightforward method for understanding a drug’s single-cell potency and can be used for any fluorescent or fluorescently labeled drug, including nanoparticles or antibody–drug conjugates. |
| first_indexed | 2024-09-23T13:42:37Z |
| format | Article |
| id | mit-1721.1/107005 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T13:42:37Z |
| publishDate | 2017 |
| publisher | Wiley Blackwell |
| record_format | dspace |
| spelling | mit-1721.1/1070052022-10-01T16:42:00Z A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin Kulkarni, Chethana Betts, Alison M. Maass, Katie F. Quadir, Mohiuddin Abdul Hammond, Paula T. Wittrup, Karl Dane Massachusetts Institute of Technology. Department of Biological Engineering Massachusetts Institute of Technology. Department of Chemical Engineering Koch Institute for Integrative Cancer Research at MIT Maass, Katie F. Quadir, Mohiuddin Abdul Hammond, Paula T. Wittrup, Karl Dane Standard cell proliferation assays use bulk media drug concentration to ascertain the potency of chemotherapeutic drugs; however, the relevant quantity is clearly the amount of drug actually taken up by the cell. To address this discrepancy, we have developed a flow cytometric clonogenic assay to correlate the amount of drug in a single cell with the cell’s ability to proliferate using a cell tracing dye and doxorubicin, a naturally fluorescent chemotherapeutic drug. By varying doxorubicin concentration in the media, length of treatment time, and treatment with verapamil, an efflux pump inhibitor, we introduced 10[superscript 5]–10[superscript 10] doxorubicin molecules per cell; then used a dye-dilution assay to simultaneously assess the number of cell divisions. We find that a cell’s ability to proliferate is a surprisingly conserved function of the number of intracellular doxorubicin molecules, resulting in single-cell IC[subscript 50] values of 4–12 million intracellular doxorubicin molecules. The developed assay is a straightforward method for understanding a drug’s single-cell potency and can be used for any fluorescent or fluorescently labeled drug, including nanoparticles or antibody–drug conjugates. Hertz Foundation (Fellowship) National Science Foundation (U.S.). Graduate Research Fellowship Program Pfizer Inc. National Cancer Institute (U.S.) (David H. Koch Institute for Integrative Cancer Research at MIT. Support (Core) Grant P30-CA14051) 2017-02-21T17:05:29Z 2017-02-21T17:05:29Z 2016-01 2015-07 Article http://purl.org/eprint/type/JournalArticle 0022-3549 1520-6017 http://hdl.handle.net/1721.1/107005 Maass, Katie F. et al. “A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin.” Journal of Pharmaceutical Sciences 104.12 (2015): 4409–4416. https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0002-5568-6455 https://orcid.org/0000-0003-2398-5896 en_US http://dx.doi.org/10.1002/jps.24631 Journal of Pharmaceutical Sciences Creative Commons Attribution-Noncommercial-Share Alike http://creativecommons.org/licenses/by-nc-sa/4.0/ application/pdf Wiley Blackwell PMC |
| spellingShingle | Kulkarni, Chethana Betts, Alison M. Maass, Katie F. Quadir, Mohiuddin Abdul Hammond, Paula T. Wittrup, Karl Dane A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title | A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title_full | A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title_fullStr | A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title_full_unstemmed | A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title_short | A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin |
| title_sort | flow cytometric clonogenic assay reveals the single cell potency of doxorubicin |
| url | http://hdl.handle.net/1721.1/107005 https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0002-5568-6455 https://orcid.org/0000-0003-2398-5896 |
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