Abstract
Laser flow cytometry has been used for monitoring cellular retention of fluorescent drugs such as fluorescent anticancer antibiotics (e.g., doxorubicin) and fluorochromes used for the detection of cellular drug efflux and resistance (e.g., rhodamine 123, Hoechst 33342). Multi parametric flow cytometry can be used for identification of tumor cell subpopulations based on their drug retention profiles with or without the presence of an efflux blocker. This rapid proce dure can be used for identification of tumor cells with the drug-resistance phenotype based on drug efflux as well as for efflux blockers that may block efflux of a chemotherapeutic agent and thus increase cellular retention and sensitivity. It has been reported recently that some of the bone marrow stem cells (SP cells) efflux the Hoechst 33342 fluorochrome and thus can be rapidly identified by comparing red vs blue fluorescence in the presence or absence of an efflux blocker such as verapamil. The present chapter discusses some of the flow cytometric methods used for the study of cellular drug retention and the artifacts that may arise in such analysis.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Goodell, M. A., Brose, K., Paradis, G., Conner, A. S., and Mulligan, R. C. (1996) Isolation and functional properties of murine hematopoietic stem cells that are replicating in vivo. J. Exp. Med. 183, 1797ā1806.
Storms, R. W., Goodell, M. A., Fisher, A., Mulligan, R. C., and Smith, C. (2000) Hoechst dye efflux reveals a novel CD7(+)CD34(-) lymphoid progenitor in human umbilical cord blood. Blood 96, 2125ā2133.
Ling, V. (1992) P-glycoprotein and resistance to anticancer drugs. Cancer 69, 2603ā2609.
Biedler, J. L. (1994) Drug resistance: genotype versus phenotype. Cancer Res. 54, 666ā678.
Dano, K. (1973) Active outward transport of daunomycin in resistant Ehrlich ascites tumor cells. Biochim Biophys. Acta 323, 466ā483.
Kessel, D., Botterill, V., and Wodinsky, I. (1968) Uptake and retention of daunomycin by mouse leukemic cells as factors in drug response. Cancer Res. 28, 938ā941.
Gottesman, M. M. (1993) How cancer cells evade chemotherapy: Sixteenth Richard and Hinda Rosenthal Foundation Award Lecture. Cancer Res. 53, 747ā754.
Gottesman, M. M. and Pastan, I. (1989) Clinical trials of agents that reverse multidrug-resistance. J. Clin. Oncol. 7, 409ā411.
Cole, S. P., Bhardwaj, G., Gerlach, J. H., et al. (1992) Overexpression of a trans porter gene in a multidrug-resistant human lung cancer cell line. Science 258, 1650ā1654.
Izquierdo, M. A., Scheffer, G. L., Flens, M. J., et al. (1996) Broad distribution of the multidrug resistance-related vault lung resistance protein in normal human tissues and tumors. Am. J. Pathol. 148, 877ā887.
Krishan, A. and Ganapathi, R. (1980) Laser flow cytometric studies on the intracellular fluorescence of anthracyclines. Cancer Res. 40, 3895ā3900.
Durand, R. E. and Olive, P. L. (1981) Flow cytometry studies of intracellular adriamycin in single cells in vitro. Cancer Res. 41, 3489ā3494.
Krishan, A. and Bourguignon, L.Y. (1984) Cell cycle related phenothiazine effects on adriamycin transport. Cell. Biol. Int. Rep. 8, 449ā456.
Krishan, A., Sauerteig, A., and Wellham, L. L. (1985) Flow cytometric studies on modulation of cellular adriamycin retention by phenothiazines. Cancer Res. 45, 1046ā1051.
Krishan, A., Sridhar, K. S., Davila, E., Vogel, C., and Sternheim, W. (1987) Pat terns of anthracycline retention modulation in human tumor cells. Cytometry 8, 306ā314.
Nair, S., Singh, S. V., Samy, T. S., and Krishan, A. (1990) Anthracycline resis tance in murine leukemic P388 cells. Role of drug efflux and glutathione related enzymes. Biochem. Pharmacol. 39, 723ā728.
Inaba, M., Kobayashi, H., Sakurai, Y., and Johnson, R. K. (1979) Active efflux of daunorubicin and adriamycin in sensitive and resistant sublines of P388 leukemia. Cancer Res. 39, 2200ā2203.
Beck, W. T., Mueller, T. J., and Tanzer, L. R. (1979) Altered surface membrane glycoproteins in Vinca alkaloid-resistant human leukemic lymphoblasts. Cancer Res. 39, 2070ā2076.
Leibovitz, A., Stinson, J. C., McCombs, W. B. III, McCoy, C. E., Mazur, K. C., and Mabry, N. D. (1976) Classification of human colorectal adenocarcinoma cell lines. Cancer Res. 36, 4562ā4569.
Krishan, A., Sauerteig, A., Andritsch, I., and Wellham, L. (1997) Flow cytometric analysis of the multiple drug resistance phenotype. Leukemia 11, 1138ā1146.
Lampidis, T. J., Munck, J. N., Krishan, A., and Tapiero, H. (1985) Reversal of resistance to rhodamine 123 in adriamycin-resistant Friend leukemia cells. Cancer Res. 45, 2626ā2631.
Tsuruo, T., Iida, H., Tsukagoshi, S., and Sakurai, Y. (1981) Overcoming of vincristine resistance in P388 leukemia in vivo and in vitro through enhanced cyto toxicity of vincristine and vinblastine by verapamil. Cancer Res. 41, 1967ā1972.
Ganapathi, R., Grabowski, D., Rouse, W., and Riegler, F. (1984) Differential effect of the calmodulin inhibitor trifluoperazine on cellular accumulation, retention, and cytotoxicity of anthracyclines in doxorubicin (adriamycin)-resistant P388 mouse leukemia cells. Cancer Res. 44, 5056ā5061.
Slater, L. M., Sweet, P., Stupecky, M., Wetzel, M. W., and Gupta, S. (1986) Cyclosporin A corrects daunorubicin resistance in Ehrlich ascites carcinoma. Br. J. Cancer 54, 235ā238.
Stein, W. D. (1997) Kinetics of the multidrug transporter (P-glycoprotein) and its reversal. Physiol. Rev. 77, 545ā590.
Krishan, A., Sridhar, K. S., Mou, C., et al. (2000) Synergistic effect of prochlorperazine and dipyridamole on the cellular retention and cytotoxicity of doxoru bicin. Clin. Cancer Res. 6, 1508ā1517.
Kim, M., Turnquist, H., Jackson, J., et al. (2002) The multidrug resistance transporter ABCG2 (breast cancer resistance protein 1) effluxes Hoechst 33342 and is overexpressed in hematopoietic stem cells. Clin. Cancer Res. 8, 22ā28.
Darzynkiewicz, Z., Traganos, F., Staiano-Coico, L., Kapuscinski, J., and Melamed, M. R. (1982) Interaction of rhodamine 123 with living cells studied by flow cytom etry. Cancer Res. 42, 799ā806.
Alabaster, O., Woods, T., Ortiz-Sanchez, V., and Jahangeer, S. (1989) Influence of microenvironmental pH on adriamycin resistance. Cancer Res. 49, 5638ā5643.
Krishan, A. (2001) Monitoring of cellular resistance to cancer chemotherapy: drug retention and efflux, in Methods in Cell Biology, 64 (Darzynkiewicz, Z., Criss man, H. A., and Robinson, J. P., eds.), Academic, San Diego, CA, pp. 193ā209.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
Ā© 2005 Humana Press Inc.
About this protocol
Cite this protocol
Krishan, A., Hamelik, R.M. (2005). Flow Cytometric Monitoring of Fluorescent Drug Retention and Efflux. In: Blumenthal, R.D. (eds) Chemosensitivity: Volume II. Methods in Molecular Medicineā¢, vol 111. Humana Press, Totowa, NJ. https://doi.org/10.1385/1-59259-889-7:149
Download citation
DOI: https://doi.org/10.1385/1-59259-889-7:149
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-58829-586-6
Online ISBN: 978-1-59259-889-2
eBook Packages: Springer Protocols