Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2014 Oct 30;5(20):9930-8.
doi: 10.18632/onco_target.2479.

Ibrutinib inhibits SDF1/CXCR4 mediated migration in AML

Lyubov Zaitseva  1 , Megan Y Murray  1 , Manar S Shafat  1 , Matthew J Lawes  2 , David J MacEwan  3 , Kristian M Bowles  4 , Stuart A Rushworth  1
Affiliations

Ibrutinib inhibits SDF1/CXCR4 mediated migration in AML

Lyubov Zaitseva et al. Onco_target. .

Abstract

Pharmacological _targeting of BTK using ibrutinib has recently shown encouraging clinical activity in a range of lymphoid malignancies. Recently we reported that ibrutinib inhibits human acute myeloid leukemia (AML) blast proliferation and leukemic cell adhesion to the surrounding bone marrow stroma cells. Here we report that in human AML ibrutinib, in addition, functions to inhibit SDF1/CXCR4-mediated AML migration at concentrations achievable in vivo. It has previously been shown that SDF1/CXCR4-induced migration is dependent on activation of downstream BTK in chronic lymphocytic leukaemia (CLL) and multiple myeloma. Here we show that SDF-1 induces BTK phosphorylation and downstream MAPK signalling in primary AML blast. Furthermore, we show that ibrutinib can inhibit SDF1-induced AKT and MAPK activation. These results reported here provide a molecular mechanistic rationale for clinically evaluating BTK inhibition in AML patients and suggests that in some AML patients the blasts count may initially rise in response to ibrutinib therapy, analgous to similar clinical observations in CLL.

PubMed Disclaimer

Conflict of interest statement

Disclosure of Conflicts of Interest

Figures

Figure 1
Figure 1. Ibrutinib inhibits AML cell migration in response to SDF-1
(A) AML cell lines were examined for CXCR4 expression (n=4) using flow cytometry (B) AML cell lines were pretreated with 500nM of ibrutinib for 1 h before wash off and then placed in the upper well of a 8.0μM transwell plate. The lower chamber contained 500ul of serum free media supplemented with SDF1 (100 ng/ml) for 3 hours and then assessed for cell number using a flow cytometer. Data were normalised to DMSO treated cells. *statistical significance. (C) Primary AML blasts (n=12) were pretreated with ibrutinib (500 nM) for 1 h before wash off and then placed in the upper well of a 8.0μM transwell plate. The lower chamber contained 500ul of serum free media supplemented with SDF1 (100 ng/ml) for 3 hours and then assessed for cell number using a flow cytometer. Data were normalised to DMSO treated cells.
Figure 2
Figure 2. BTK is activated in response to SDF-1 in human AML
(A) AML blasts were treated with SDF1 (100 ng/ml) for indicated times. Protein extracts were obtained and Western blot analysis was conducted for pBTK, BTK, pMAPK, MAPK and β-actin protein levels. (B) AML blasts were pretreated with increasing concentrations of ibrutinib for 1h and then treated with SDF1 (100 ng/ml) for 10 mins. Protein extracts were obtained and Western blot analysis was conducted for pATK, ATK, pMAPK, MAPK and β-actin protein levels. These results are representative of experiments which were repeated three times with the bar graphs showing mean expression (as measured by densitometry) compared to control from all 3 experiments.
Figure 3
Figure 3. Pharmacological inhibition of G-proteins, AKT and ERK block SDF1 induced migration in AML
(A) MV4-11 cells were pretreated with pertussis toxin (100ng/ml) for 30 mins and then stimulated with SDF1 for 10mins. Protein extracts were obtained and Western blot analysis was conducted for pBTK, BTK and β-actin protein levels. These results are representative of experiments which were repeated three times. (B) MV4-11 cells were pretreated with pertussis toxin (100ng/ml), PD98059 (20 μM) and AKT inhibitor VIII (2 μM) for 30 mins and then placed in the upper well of a 8.0μM transwell plate. The lower chamber contained 500ul of serum free media supplemented with SDF1 (100 ng/ml) for 3 hours and then assessed for cell number using a flow cytometer. Data were normalised to DMSO treated cells. *statistical significance compared to SDF1 MV4-11 cells.
Figure 4
Figure 4. Knockdown of BTK inhibits SDF1 induced migration in AML
(A) AML cell lines (HL60 and THP-1) were transduced with BTK-_targeted miRNA (BTK-KD) or a negative-_targeted miRNA (NEG-KD) GFP-tagged lentiviral constructs for 72 h. Protein extracts were obtained and Western blot analysis was conducted for BTK and β-actin protein levels. These results are representative of experiments which were repeated three times (B) HL60 and THP-1 were transduced with BTK-KD and NEG-KD lentivirus for 72 h and then placed in the upper well of a 8.0μM transwell plate. The lower chamber contained 500ul of serum free media supplemented with SDF1 (100 ng/ml) for 3 hours and then assessed for cell number using a flow cytometer. Data were normalised to NEG-KD cells. *statistical significance. (C) Schematic to show the role of BTK inhibition by ibrutinib in preventing AML migration.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Juliusson G, Antunovic P, Derolf A, Lehmann S, Mollgard L, Stockelberg D, Tidefelt U, Wahlin A, Hoglund M. Age and acute myeloid leukemia: real world data on decision to treat and outcomes from the Swedish Acute Leukemia Registry. Blood. 2009;113:4179–4187. - PubMed
    1. Burnett A. Ham-Wasserman Lecture. 2012 https://ash.confex.com/ash/2012/webprogram/Session3976.html
    1. Ezell SA, Mayo M, Bihani T, Tepsuporn S, Wang S, Passino M, Grosskurth SE, Collins M, Parmentier J, Reimer C, Byth KF. Synergistic induction of apoptosis by combination of BTK and dual mTORC1/2 inhibitors in diffuse large B cell lymphoma. Onco_target. 2014;5:4990–5001. - PMC - PubMed
    1. Tai YT, Anderson KC. Bruton’s tyrosine kinase: onco_target in myeloma. Onco_target. 2012;3:913–914. - PMC - PubMed
    1. Rushworth SA, Macewan DJ, Bowles KM. Ibrutinib in Relapsed Chronic Lymphocytic Leukemia. New England Journal of Medicine. 2013;369:1278–1279. - PubMed

Publication types

MeSH terms

  NODES
twitter 2