A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin

作者：Maass Katie F; Kulkarni Chethana; Quadir Mohiuddin A; Hammond Paula T; Betts Alison M; Wittrup Karl Dane^*

来源：Journal of Pharmaceutical Sciences, 2015, 104(12): 4409-4416.

DOI：10.1002/jps.24631

• cancer chemotherapy
• pharmacodynamics
• drug effects
• efflux pumps
• cell lines

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.

• 出版日期2015-12
• 单位MIT