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PI3K/mTOR pathway inhibitors sensitize chronic myeloid leukemia stem cells to nilotinib and restore the response of progenitors to nilotinib in the presence of stem cell factor.

Airiau K, Mahon FX, Josselin M, Jeanneteau M, Belloc F - Cell Death Dis (2013)

Bottom Line: We previously demonstrated that nilotinib-induced apoptosis was reduced by stem cell factor (SCF) addition.However, this pathway seems not to be involved in the nilotinib-induced resistance of the CML stem cell population.Conversely, PI3K inhibition sensitized both CML progenitors and stem cells to nilotinib, suggesting that, downstream PI3K, two different kinase pathways are activated in CML progenitor and stem cell populations.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratoire d'Hématopoïèse Leucémique et Cibles Thérapeutiques, INSERM 1035, Université Bordeaux Segalen, 146 Rue Léo Saignat, Bordeaux Cedex 33076, France [2] CHU Bordeaux, Hôpital Haut-Lévêque, Laboratoire d'Hématologie, Avenue Magellan 33604 Pessac, France.

ABSTRACT
Nilotinib is a second-generation tyrosine kinase inhibitor, designed to specifically inhibit break-point cluster region (BCR)-Abelson (ABL) and developed to treat chronic myeloid leukemia (CML) in patients showing a resistance to imatinib. We previously demonstrated that nilotinib-induced apoptosis was reduced by stem cell factor (SCF) addition. Here, the SCF-activated survival pathway was investigated. BCR-ABL expression was accompanied by the activation of the SCF receptor: c-KIT. Nilotinib inhibited this activation that was restored by SCF binding. Parallel variations were observed for mammaliam target of rapamycin (mTOR) kinase and mTOR complex 1 substrate S6K. The inhibition of mTORC1 restored the response of BCR-ABL cell lines to nilotinib in the presence of SCF. PI3K inhibition restored nilotinib-induced apoptosis. On hematopoietic progenitors from CML patient's bone marrows, mTORC1 inhibition also restored nilotinib sensitivity in the presence of SCF, confirming its involvement in SCF-activated survival pathway. However, this pathway seems not to be involved in the nilotinib-induced resistance of the CML stem cell population. Conversely, PI3K inhibition sensitized both CML progenitors and stem cells to nilotinib, suggesting that, downstream PI3K, two different kinase pathways are activated in CML progenitor and stem cell populations.

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Related in: MedlinePlus

SCF maintains the activation of the mTOR pathway without restoring the global tyrosine phosphorylation state. (a) Global tyrosine phosphorylation state was measured by flow cytometry in K562 and LAMA-84 cells treated or not with 20 nM nilotinib and/or 100 ng/ml SCF for 24 h. Results are expressed in percentage of the control value. (b and c) K562 and LAMA-84 cell lysates were analyzed by western blot after treatment with nilotinib and/or SCF as described above. The blots were probed with anti-phosphotyrosine and anti-phospho-c-kit (b) or anti-phospho-c-kit, anti-phospho-mTOR and anti-phospho-S6K (c). Anti-tubulin antibody was used to verify the loading homogeneity
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fig2: SCF maintains the activation of the mTOR pathway without restoring the global tyrosine phosphorylation state. (a) Global tyrosine phosphorylation state was measured by flow cytometry in K562 and LAMA-84 cells treated or not with 20 nM nilotinib and/or 100 ng/ml SCF for 24 h. Results are expressed in percentage of the control value. (b and c) K562 and LAMA-84 cell lysates were analyzed by western blot after treatment with nilotinib and/or SCF as described above. The blots were probed with anti-phosphotyrosine and anti-phospho-c-kit (b) or anti-phospho-c-kit, anti-phospho-mTOR and anti-phospho-S6K (c). Anti-tubulin antibody was used to verify the loading homogeneity

Mentions: We first studied the effect of SCF addition on tyrosine phosphorylation. As shown in figures 2a and b, SCF could only partly restore the level of tyrosine phosphorylation after nilotinib treatment. In the same time, as expected, the SCF did not modifiy the BCR-ABL dephosphorylation induced by nilotinib. The SCF receptor c-KIT was constitutively activated in both BCR-ABL-expressing cell lines (Figure 2b). Nilotinib treatment, using low concentration (i.e., 20 nM) specific to BCR-ABL inhibition, was associated with a loss of c-KIT constitutive activation, restored in a large part after c-KIT binding by SCF (Figure 2b). Altogether, these results suggest that c-KIT constitutive phosphorylation resulted from BCR-ABL activity.


PI3K/mTOR pathway inhibitors sensitize chronic myeloid leukemia stem cells to nilotinib and restore the response of progenitors to nilotinib in the presence of stem cell factor.

Airiau K, Mahon FX, Josselin M, Jeanneteau M, Belloc F - Cell Death Dis (2013)

SCF maintains the activation of the mTOR pathway without restoring the global tyrosine phosphorylation state. (a) Global tyrosine phosphorylation state was measured by flow cytometry in K562 and LAMA-84 cells treated or not with 20 nM nilotinib and/or 100 ng/ml SCF for 24 h. Results are expressed in percentage of the control value. (b and c) K562 and LAMA-84 cell lysates were analyzed by western blot after treatment with nilotinib and/or SCF as described above. The blots were probed with anti-phosphotyrosine and anti-phospho-c-kit (b) or anti-phospho-c-kit, anti-phospho-mTOR and anti-phospho-S6K (c). Anti-tubulin antibody was used to verify the loading homogeneity
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824646&req=5

fig2: SCF maintains the activation of the mTOR pathway without restoring the global tyrosine phosphorylation state. (a) Global tyrosine phosphorylation state was measured by flow cytometry in K562 and LAMA-84 cells treated or not with 20 nM nilotinib and/or 100 ng/ml SCF for 24 h. Results are expressed in percentage of the control value. (b and c) K562 and LAMA-84 cell lysates were analyzed by western blot after treatment with nilotinib and/or SCF as described above. The blots were probed with anti-phosphotyrosine and anti-phospho-c-kit (b) or anti-phospho-c-kit, anti-phospho-mTOR and anti-phospho-S6K (c). Anti-tubulin antibody was used to verify the loading homogeneity
Mentions: We first studied the effect of SCF addition on tyrosine phosphorylation. As shown in figures 2a and b, SCF could only partly restore the level of tyrosine phosphorylation after nilotinib treatment. In the same time, as expected, the SCF did not modifiy the BCR-ABL dephosphorylation induced by nilotinib. The SCF receptor c-KIT was constitutively activated in both BCR-ABL-expressing cell lines (Figure 2b). Nilotinib treatment, using low concentration (i.e., 20 nM) specific to BCR-ABL inhibition, was associated with a loss of c-KIT constitutive activation, restored in a large part after c-KIT binding by SCF (Figure 2b). Altogether, these results suggest that c-KIT constitutive phosphorylation resulted from BCR-ABL activity.

Bottom Line: We previously demonstrated that nilotinib-induced apoptosis was reduced by stem cell factor (SCF) addition.However, this pathway seems not to be involved in the nilotinib-induced resistance of the CML stem cell population.Conversely, PI3K inhibition sensitized both CML progenitors and stem cells to nilotinib, suggesting that, downstream PI3K, two different kinase pathways are activated in CML progenitor and stem cell populations.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratoire d'Hématopoïèse Leucémique et Cibles Thérapeutiques, INSERM 1035, Université Bordeaux Segalen, 146 Rue Léo Saignat, Bordeaux Cedex 33076, France [2] CHU Bordeaux, Hôpital Haut-Lévêque, Laboratoire d'Hématologie, Avenue Magellan 33604 Pessac, France.

ABSTRACT
Nilotinib is a second-generation tyrosine kinase inhibitor, designed to specifically inhibit break-point cluster region (BCR)-Abelson (ABL) and developed to treat chronic myeloid leukemia (CML) in patients showing a resistance to imatinib. We previously demonstrated that nilotinib-induced apoptosis was reduced by stem cell factor (SCF) addition. Here, the SCF-activated survival pathway was investigated. BCR-ABL expression was accompanied by the activation of the SCF receptor: c-KIT. Nilotinib inhibited this activation that was restored by SCF binding. Parallel variations were observed for mammaliam target of rapamycin (mTOR) kinase and mTOR complex 1 substrate S6K. The inhibition of mTORC1 restored the response of BCR-ABL cell lines to nilotinib in the presence of SCF. PI3K inhibition restored nilotinib-induced apoptosis. On hematopoietic progenitors from CML patient's bone marrows, mTORC1 inhibition also restored nilotinib sensitivity in the presence of SCF, confirming its involvement in SCF-activated survival pathway. However, this pathway seems not to be involved in the nilotinib-induced resistance of the CML stem cell population. Conversely, PI3K inhibition sensitized both CML progenitors and stem cells to nilotinib, suggesting that, downstream PI3K, two different kinase pathways are activated in CML progenitor and stem cell populations.

Show MeSH
Related in: MedlinePlus