Abstract
Flow cytometry can sensitively detect and efficiently sort cells based on fluorescent signals integrated into cellular markers of proteins or DNA. It has been broadly applied to assess cell division, apoptosis and to isolate cells including stem cells. As the seeds for tumorigenesis and metastasis, cancer stem cells (CSCs) are often more resistant to cytotoxins and anticancer agents than other heterogeneous cells in tumors. Analyzing CSCs under treatments is an effective way to evaluate new therapeutic agents for cancers. We introduce a method using flow cytometry to assess breast CSCs (CD44+/CD24−/low) in human MCF-7/Dox breast cancer cells, after the treatment of mixed-backbone oligonucleotide against glucosylceramide synthase. Flow cytometry analysis of CSCs is a reliable, effective, and easy-handling approach to screen agents targeting CSCs.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Reya, T., Morrison, S. J., Clarke, M. F., and Weissman, I. L. (2001) Stem cells, cancer, and cancer stem cells. Nature. 414: 105–11.
Dalerba, P., Cho, R. W., and Clarke, M. F. (2007) Cancer stem cells: models and concepts. Annu Rev Med. 58: 267–84.
Visvader, J. E., and Lindeman, G. J. (2008) Cancer stem cells in solid tumours: accumulating evidence and unresolved questions. Nat Rev Cancer. 8: 755–68.
Al-Hajj, M., Wicha, M. S., Benito-Hernandez, A., Morrison, S. J., and Clarke, M. F. (2003) Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci U S A. 100: 3983–8.
Fillmore, C. M., and Kuperwasser, C. (2008) Human breast cancer cell lines contain stem-like cells that self-renew, give rise to phenotypically diverse progeny and survive chemotherapy. Breast Cancer Res. 10: R25.
Phillips, T. M., McBride, W. H., and Pajonk, F. (2006) The response of CD24(-/low)/CD44+ breast cancer-initiating cells to radiation. J Natl Cancer Inst. 98: 1777–85.
Tang, C., Ang, B. T., and Pervaiz, S. (2007) Cancer stem cell: target for anti-cancer therapy. FASEB J. 21: 3777–85.
Korkaya, H., and Wicha, M. S. (2009) HER-2, notch, and breast cancer stem cells: targeting an axis of evil. Clin Cancer Res. 15: 1845–7.
Patwardhan, G. A., Zhang, Q. J., Yin, D., Gupta, V., Bao, J., Senkal, C. E., Ogretmen, B., Cabot, M. C., Shah, G. V., Sylvester, P. W., Jazwinski, S. M., and Liu, Y. Y. (2009) A new mixed-backbone oligonucleotide against glucosylceramide synthase Sensitizes multidrug-resistant tumors to apoptosis. PLoS One. 4: e6938.
Petriz, J. (2007) Flow cytometry of the side population (SP). Curr Protoc Cytom. Chapter 9, Unit9 23.
Gupta, P. B., Onder, T. T., Jiang, G., Tao, K., Kuperwasser, C., Weinberg, R. A., and Lander, E. S. (2009) Identification of selective inhibitors of cancer stem cells by high-throughput screening. Cell. 138: 645–59.
Yamashita, T., Wada, R., Sasaki, T., Deng, C., Bierfreund, U., Sandhoff, K., and Proia, R. L. (1999) A vital role for glycosphingolipid synthesis during development and differentiation. Proc Natl Acad Sci U S A. 96: 9142–7.
Liour, S. S., and Yu, R. K. (2002) Differential effects of three inhibitors of glycosphingolipid biosynthesis on neuronal differentiation of embryonal carcinoma stem cells. Neurochem Res. 27: 1507–12.
Salli, U., Fox, T. E., Carkaci-Salli, N., Sharma, A., Robertson, G. P., Kester, M., and Vrana, K. (2009) Propagation of undifferentiated human embryonic stem cells with nano-liposomal ceramide. Stem Cells Dev. 18: 55–66.
Mehta, K. (1994) High levels of transglutaminase expression in doxorubicin-resistant human breast carcinoma cells. Int J Cancer. 58: 400–6.
Liu, Y. Y., Gupta, V., Patwardhan, G. A., Bhinge, K., Zhao, Y., Bao, J., Mehendale, H., Cabot, M. C., Li, Y. T., Jazwinski, S. M. (2010) Glucosylceramide synthase upregulates MDR1 expression in the regulation of cancer drug resistance through cSrc and beta-catenin signaling. Mol Cancer. 9:145.
Acknowledgments
This work was supported by United State Public Health Service/NIH grant P20 RR16456 (Y.Y.L) and grant 5G12RR026260-02 (Q.J.Z) from NCRR. We thank Dr. Benny Blaylock and Dr. Sharon Meyer (College of Pharmacy, University of Louisiana at Monroe) for their kind advices on flow cytometer.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2011 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Gupta, V., Zhang, QJ., Liu, YY. (2011). Evaluation of Anticancer Agents Using Flow Cytometry Analysis of Cancer Stem Cells. In: Satyanarayanajois, S. (eds) Drug Design and Discovery. Methods in Molecular Biology, vol 716. Humana Press. https://doi.org/10.1007/978-1-61779-012-6_11
Download citation
DOI: https://doi.org/10.1007/978-1-61779-012-6_11
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-61779-011-9
Online ISBN: 978-1-61779-012-6
eBook Packages: Springer Protocols