Ibrutinib selectively targets flt3-itd in mutant flt3-positive aml

Access through your institution Buy or subscribe Ibrutinib (PCI-32765) is an irreversible BTK (Bruton’s tyrosine kinase) kinase inhibitor that has been extensively used as a tool compound to

validate the role of BTK kinase in B cell related malignances.1, 2 Ibrutinib has been shown in preclinical studies to inhibit the proliferation of diffuse large B-cell lymphoma cells,

mantle cell lymphoma cells, chronic lymphocytic leukemia cells and multiple myeloma cells by blocking BTK kinase activity; ibrutinib was recently approved for the clinical application on

mantle cell lymphoma and chronic lymphocytic leukemia cells.3, 4, 5, 6, 7 Ibrutinib has also exhibited anti-inflammatory effects in preclinical models.8, 9 Recently, it has been reported

that ibrutinib is also effective against epidermal growth factor receptor mutant-positive non-small cell lung cancers through inhibition of epidermal growth factor receptor kinase

activities.10 In addition, there is evidence showing that BTK is also an important target for Acute Myeloid Leukemia (AML).11 As all of these cell lines express BTK kinase, we then studied

the effect of ibrutinib on BTK signaling. Upon anti-immunoglobulin M (IgM) stimulation of B-cell receptor to active the BTK kinase signaling pathway, we found that only in the FLT3 wild-type

(wt)-expressing cell lines (U937 and OCI-AML3), downstream mediators of BTK kinase were affected despite BTK phosphorylation having been inhibited in both FLT3 wt and FLT3-ITD cell lines.9

This suggests that FLT3-ITD mutant cell lines might not rely on BTK signaling for growth (Supplementary Figure 3A). Interestingly, except NOMO-1 and U937 cells, the phosphorylation of

BTKY223 of MOLM14, MOLM13, MV4-11, OCI-AML-3 and HL-60 cells did not respond to anti-IgM stimulation apparently, which might be due to the basal-level expression of the IgM in these AML

cells (Supplementary Figure 3B). AC220 did not affect BTK kinase activity and its direct downstream target PLCγ but instead inhibited phosphorylation of protein kinase B and

extracellular-signal-regulated kinase. The BTK inhibitors, CGI-1746 and AVL-292, were effective against BTK kinase and PLCγ kinase phosphorylation, but had no effect on the phosphorylation

of protein kinase B and extracellular-signal-regulated kinase. These results suggest that ibrutinib may mainly exert its inhibitory activity through FLT3 kinase but not BTK kinase in the

FLT3-ITD positive cells. To further confirm this, we knocked down BTK kinase in MOLM14 (FLT3-ITD), MOLM13 (FLT3-ITD), human erythroleukemia cell (HEL) (FLT3 wt) and NB4 (FLT3 wt) cells and

found that the growth of MOLM14 and MOLM13 was only minimally affected; however NB4 cells were significantly inhibited and HEL cells were moderately inhibited (Figures 2a and b). The fact

that BTK knockdown in HEL cells did not lead to suppressed growth may be due to the fact that mutant JAK2 is the oncogenic driver in this cell line.15 In addition, the finding that ibrutinib

still significantly blocked the growth of BTK-knockdown MOLM14 and MOLM13 cells further validated that the growth inhibitory effect of ibrutinib is likely dependent mainly on FLT3-ITD

inhibition. However, BTK kinase inhibition may also exert a combinatorial effect (Figure 2c). It has recently been reported that BTK kinase is activated by FLT3-ITD in FLT3-ITD positive cell

lines and dual inhibition of BTK and FLT3 kinase demonstrates combinatorial effects; these findings further confirm this observation.16 This is a preview of subscription content, access via

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88–94. Article  CAS  Google Scholar  Download references ACKNOWLEDGEMENTS JL, QL and WW are supported by the grant of ‘Cross-disciplinary Collaborative Teams Program for Science, Technology

and Innovation (2014-2016)’ from Chinese Academy of Sciences. ZZ is supported by Anhui Province Natural Science Foundation Annual Key Program (grant number: 1301023011). We thank China

‘Thousand Talents Program’ support for QL and ‘Hundred Talents Program’ of The Chinese Academy of Sciences support for JL and WW. AUTHOR INFORMATION Author notes * H Wu, C Hu, A Wang and E L

Weisberg: These authors contribute equally to this work. AUTHORS AND AFFILIATIONS * High Magnetic Field Laboratory, Chinese Academy of Sciences, Hefei, Anhui, China H Wu, C Hu, A Wang, W

Wang, C Chen, Z Zhao, K Yu, J Liu, J Wu, L Wang, B Wang, J Liu & Q Liu * University of Science and Technology of China, Hefei, Anhui, China H Wu, A Wang, J Wu & Q Liu * Department of

Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, USA E L Weisberg, A Nonami, R M Stone, S Liu & J D Griffin Authors * H Wu View author publications

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A. _et al._ Ibrutinib selectively targets FLT3-ITD in mutant FLT3-positive AML. _Leukemia_ 30, 754–757 (2016). https://doi.org/10.1038/leu.2015.175 Download citation * Published: 03 July

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