Abstract
As the signaling pathways involved in leukemogenesis are being elucidated, several proteins have emerged as potential targets for therapy. Downstream from those targets are numerous intracellular factors that are constantly modulated. Monitoring those factors could provide insight into the potential efficacy of therapies by predicting which patients will respond to them and by determining the optimal dosage that will inhibit the target protein. We describe a flow cytometry method for quantitation of total and phosphorylated intracellular proteins. Compared with Western blot analysis, this technique dramatically decreases time and labor while providing multiparameter information on specific cell populations. As an example, total and phosphorylated CRKL is quantitated. The methodology has the potential for widespread application in the monitoring of targeted therapy.
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References
Chopra, R., Pu, Q. Q., and Elefanty, A. G. (1999) Biology of BCR-ABL. Blood Rev. 13, 211–229.
Kantarjian, H. M., Talpaz, M., Cortes J., et al. (2003) Quantitative polymerase chain reaction monitoring of BCR-ABL during therapy with imatinib mesylate (STI571; Gleevec) in chronic-phase chronic myelogenous leukemia. Clin. Cancer Res. 9, 160–166.
O’Brien, S. G., Guilhot, E., Larson, R. A., et al. (2003) Imatinib compared with interferon and low-dose cytarabine for newly diagnosed chronic-phase chronic myeloid leukemia. N. Engl. J. Med. 348, 994–1004.
Deininger, M., Buchdunger, E., and Druker, B. J. (2003) The development of imatinib as a therapeutic agent for chronic myeloid leukemia. Blood 105, 1321–1331.
Gorre, M. E., Mohammed, M., Ellwood, K., et al. (2001) Clinical resistance to STI-571 cancer therapy caused by BCR-ABL gene mutation or amplification. Science 293, 876–880.
ten Hoeve, J., Arlinghaus, R. B., Guo, J. Q., Heisterkamp, N., and Groffen, J. (1994) Tyrosine phosphorylation of CRKL in Philadelphia-!-leukemia. Blood 84, 1731–1736.
Oda, T., Heaney, C., Hagopian, J. R., Okuda, K., Griffin, J. D., and Druker, B. J. (1994) Crkl is the major tyrosine-phophorylated protein in neutrophils from patients with chronic myelogenous leukemia. J. Biol. Chem. 269, 22,925–22,928.
Nichols, G. L., Raines, M. A., Vera, J. C., Lacomis, L., Tempst, P., anf Golde, D. W. (1994) Identification of CRKL as the constitutively phosphorylated 39-kD tyrosine phosphoprotein in chronic myelogenous leukemia cells. Blood 84, 2912–2918.
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© 2007 Humana Press Inc., Totowa, NJ
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Chan, H.E.H., Jilani, I., Chang, R., Albitar, M. (2007). Monitoring Cell Signaling Pathways by Quantitative Flow Cytometry. In: Albitar, M. (eds) Monoclonal Antibodies. Methods in Molecular Biology, vol 378. Humana Press. https://doi.org/10.1007/978-1-59745-323-3_7
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DOI: https://doi.org/10.1007/978-1-59745-323-3_7
Publisher Name: Humana Press
Print ISBN: 978-1-58829-567-5
Online ISBN: 978-1-59745-323-3
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