A Flow Cytometric Clonogenic Assay Reveals the Single‐Cell Potency of Doxorubicin. Issue 12 (7th September 2015)

• Record Type: Journal Article
• Authors: Maass, Katie F.; Kulkarni, Chethana; Quadir, Mohiuddin A.; Hammond, Paula T.; Betts, Alison M.; Wittrup, Karl Dane
• Is Part Of: Journal of pharmaceutical sciences. Volume 104:Issue 12(2015:Dec.)
• Page Start: 4409
• Page End: 4416
• Publication Date: 2015-09-07
• Subjects: cancer chemotherapy -- pharmacodynamics -- drug effects -- efflux pumps -- cell lines; Pharmacy -- Periodicals; 615.1
• DOI: 10.1002/jps.24631
• Languages: English
• ISSNs: 0022-3549

Abstract

Abstract : 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 5 –10 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 IC50 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. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 104:4409–4416, 2015