Enhanced ABL-inhibitor-induced MAPK-activation in T315I-BCR-ABL-expressing cells: a potential mechanism of altered leukemogenicity

Härtel, Nicolai; Klag, Thomas; Hanfstein, Benjamin; Mueller, Martin; Schenk, Thomas; Erben, Philipp; Hochhaus, Andreas; Rosée, Paul La

doi:10.1007/s00432-011-1086-x

Cited by 7 publications

(9 citation statements)

References 46 publications

(51 reference statements)

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“…In contrast to congruent signaling inhibition of the BCR‐ABL‐dependent CRKL‐protein in response to TKI, paradoxical activation of the BCR‐ABL‐dependent MAP‐kinases ERK1/2 has been reported in cell lines and primary CML samples exposed to TKIs in vitro (). This was confirmed by several studies investigating BCR‐ABL‐dependent signaling in response to all clinically available TKIs (). Recent experimental evidence suggests that Nilotinib inhibits RAF but triggers aberrant, RAS‐dependent RAF dimerization in TKI‐resistant cells, which itself triggers MAPK‐activation ().…”

supporting

confidence: 55%

“…Our previous results suggested that nilotinib induces off target activation of Erk1/2 in Ba/F3p210 T315I cells expressing the TKI‐resistant BCR‐ABL mutant T315I (). Our results were recently confirmed by Packer etal.…”

Section: Resultsmentioning

confidence: 99%

“…As a result of the screening efforts, we decided to use the mouse monocloncal antiphospho‐p44/42 MAPK (ERK1/2) (Thr202/Tyr204) (Cell Signaling #9106). This antibody has also been used in our previous studies on MAPK‐activation that were based on Western blot analysis (). For flow analysis the antibody was labeled with a fluorophore conjugated antibody.…”

Section: Resultsmentioning

confidence: 99%

“…). Mutant Ba/F3p210 T315I cells were used, as this TKI‐resistant subline showed maximum nilotinib induced paradoxical ERK1/2‐activation in previous experiments ().…”

Section: Resultsmentioning

confidence: 99%

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Paradoxical MAPK‐activation in response to treatment with tyrosine kinase inhibitors in CML: Flow cytometry loses track

Schnetzke

Fischer

Frietsch

et al. 2013

Cytometry Part B Clinical

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Background: Paradoxical activation of the MAP-kinases, ERK1, and ERK2 (ERK1/2) is observed in CML cell lines and primary CML patient cells treated with tyrosine kinase inhibitors (TKI) in vitro. The commonly accepted assumption is that activated ERK1/2 is key regulators of survival of leukemic cells treated with kinase inhibitors. Hence, paradoxical ERK1/2-activation may trigger resistance in vivo, which yet has to be shown. We therefore sought to establish a flow cytometric assay that enables us to measure paradoxical TKI-induced ERK1/2-activation on a single cell basis in primary CML cells.Methods: Side-by-side Western blot and intracellular flow cytometry (FCM) after in vitro exposure of cell lines and primary cells to nilotinib were performed. Detailed analysis of pre-analytical factors and the issue of compartmentalization of phosphorylated ERK1/2 by confocal laser scanning microscopy were performed.Result: Results were conflicting in that pERK-activation was robustly detected in Western blot assays, but not when cells were analyzed by FCM despite well functioning positive and negative controls. This is in contrast to experiments on other targets such as phospho-CrkL, where also in our hands TKI-dependent inhibition of phosphorylation is trackable by both Western blot and FCM assays.Conclusions: To our knowledge this is the first report of discordant results in phospho-protein analysis in TKI-treated cells analyzed by Western blot vs. FCM. We speculate that a substance specific interaction interferes with fluorescence dependent methods seeking to track phosphorylated ERK1/2 in TKItreated cells.

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supporting

confidence: 55%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

“…). Mutant Ba/F3p210 T315I cells were used, as this TKI‐resistant subline showed maximum nilotinib induced paradoxical ERK1/2‐activation in previous experiments ().…”

Section: Resultsmentioning

confidence: 99%

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Paradoxical MAPK‐activation in response to treatment with tyrosine kinase inhibitors in CML: Flow cytometry loses track

Schnetzke

Fischer

Frietsch

et al. 2013

Cytometry Part B Clinical

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“…9 We have shown that Ba/F3-BCR-ABL cell survival is associated with MAPK-activation in the presence of IL3. 10 Additionally, Ba/F3-BCR-ABL cells rapidly upregulate the common b-chain of the IL3-receptor (cCRbc) to compensate for loss of BCR-ABL activity during TKI-treatment. 11 Thus disruption of these signaling pathways could be an attractive target to combat resistance against TKI-treatment.…”

Section: Introductionmentioning

confidence: 99%

Omacetaxine mepesuccinate prevents cytokine-dependent resistance to nilotinib in vitro: potential role of the common β-subunit c of cytokine receptors

et al. 2012

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Varying effects of tyrosine kinase inhibitors on platelet function—A need for individualized CML treatment to minimize the risk for hemostatic and thrombotic complications?

et al. 2019

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Since their introduction, tyrosine kinase inhibitors (TKIs, eg, imatinib, nilotinib, dasatinib, bosutinib, ponatinib) have revolutionized the treatment of chronic myeloid leukemia (CML). However, long‐term treatment with TKIs is associated with serious adverse events including both bleeding and thromboembolism. Experimental studies have shown that TKIs can cause platelet dysfunction. Herein, we present the first side‐by‐side investigation comparing the effects of currently used TKIs on platelet function and thrombin generation when used in clinically relevant concentrations. A flow cytometry multiparameter protocol was used to study a range of significant platelet activation events (fibrinogen receptor activation, alpha granule, and lysosomal exocytosis, procoagulant membrane exposure, and mitochondrial permeability changes). In addition, thrombin generation was measured in the presence of TKIs to assess the effects on global hemostasis. Results show that dasatinib generally inhibited platelet function, while bosutinib, nilotinib, and ponatinib showed less consistent effects. In addition to these general trends for each TKI, we observed a large degree of interindividual variability in the effects of the different TKIs. Interindividual variation was also observed when blood from CML patients was studied ex vivo with whole blood platelet aggregometry, free oscillation rheometry (FOR), and flow cytometry. Based on the donor responses in the side‐by‐side TKI study, a TKI sensitivity map was developed. We propose that such a sensitivity map could potentially become a valuable tool to help in decision‐making regarding the choice of suitable TKIs for a CML patient with a history of bleeding or atherothrombotic disease.

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Enhanced ABL-inhibitor-induced MAPK-activation in T315I-BCR-ABL-expressing cells: a potential mechanism of altered leukemogenicity

Cited by 7 publications

References 46 publications

Paradoxical MAPK‐activation in response to treatment with tyrosine kinase inhibitors in CML: Flow cytometry loses track

Paradoxical MAPK‐activation in response to treatment with tyrosine kinase inhibitors in CML: Flow cytometry loses track

Omacetaxine mepesuccinate prevents cytokine-dependent resistance to nilotinib in vitro: potential role of the common β-subunit c of cytokine receptors

Varying effects of tyrosine kinase inhibitors on platelet function—A need for individualized CML treatment to minimize the risk for hemostatic and thrombotic complications?

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