BCR-ABL activity and its response to drugs can be determined in CD34+ CML stem cells by CrkL phosphorylation status using flow cytometry.

Hamilton, A, Elrick, L, Mysina, Svetlana, Copland, M, Jørgensen, H, Melo, J V and Holyoake, T (2006) BCR-ABL activity and its response to drugs can be determined in CD34+ CML stem cells by CrkL phosphorylation status using flow cytometry. Leukemia, 20 (6). pp. 1035-9. ISSN 0887-6924

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Abstract

In chronic myeloid leukaemia, CD34(+) stem/progenitor cells appear resistant to imatinib mesylate (IM) in vitro and in vivo. To investigate the underlying mechanism(s) of IM resistance, it is essential to quantify Bcr-Abl kinase status at the stem cell level. We developed a flow cytometry method to measure CrkL phosphorylation (P-CrkL) in samples with <10(4) cells. The method was first validated in wild-type (K562) and mutant (BAF3) BCR-ABL(+) as well as BCR-ABL(-) (HL60) cell lines. In response to increasing IM concentration, there was a linear reduction in P-CrkL, which was Bcr-Abl specific and correlated with known resistance. The results were comparable to those from Western blotting. The method also proved to be reproducible with small samples of normal and Ph(+) CD34(+) cells and was able to discriminate between Ph(-), sensitive and resistant Ph(+) cells. This assay should now enable investigators to unravel the mechanism(s) of IM resistance in stem cells.

Item Type: Article
Subjects: Sciences > Biomedical Sciences
Depositing User: Svetlana Mysina
Date Deposited: 08 Oct 2018 09:53
Last Modified: 09 Oct 2018 10:27
URI: http://sure.sunderland.ac.uk/id/eprint/10016

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