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Ponatinib efficiently kills imatinib-resistant chronic eosinophilic leukemia cells harboring gatekeeper mutant T674I FIP1L1-PDGFRα: roles of Mcl-1 and β-catenin.

Jin Y, Ding K, Li H, Xue M, Shi X, Wang C, Pan J - Mol. Cancer (2014)

Bottom Line: Therefore, novel TKIs effective against T674I FIP1L1-PDGFRα are needed.The purpose of this study was to examine the effect of ponatinib on T674I FIP1L1-PDGFRα.It induced apoptosis in CEL cells with caspase-3-dependent cleavage of Mcl-1, and inhibited tyrosine phosphorylation of β-catenin to decrease its stability and pro-survival functions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathophysiology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China. panjx2@mail.sysu.edu.cn.

ABSTRACT

Background: T674I FIP1L1-PDGFRα in a subset of chronic eosinophilic leukemia (CEL) is a gatekeeper mutation that is resistant to many tyrosine kinase inhibitors (TKIs) (e.g., imatinib, nilotinib and dasatinib), similar to T315I Bcr-Abl. Therefore, novel TKIs effective against T674I FIP1L1-PDGFRα are needed. Ponatinib (AP24534) is a novel orally bioavailable TKI against T315I Bcr-Abl, but it is not clear whether ponatinib is effective against T674I FIP1L1-PDGFRα. The purpose of this study was to examine the effect of ponatinib on T674I FIP1L1-PDGFRα.

Methods: Molecular docking analysis in silico was performed. The effects of ponatinib on PDGFRα signaling pathways, apoptosis and cell cycling were examined in EOL-1, BaF3 cells expressing either wild type (WT) or T674I FIP1L1-PDGFRα. The in vivo antitumor activity of ponatinib was evaluated with xenografted BaF3-T674I FIP1L1-PDGFRα cells in nude mice models.

Results: Molecular docking analysis revealed that ponatinib could bind to the DFG (Asp-Phe-Gly)-out state of T674I PDGFRα. Ponatinib potently inhibited the phosphorylation of WT and T674I FIP1L1-PDGFRα and their downstream signaling molecules (e.g., Stat3, Stat5). Ponatinib strikingly inhibited the growth of both WT and T674I FIP1L1-PDGFRα-carrying CEL cells (IC50: 0.004-2.5 nM). It induced apoptosis in CEL cells with caspase-3-dependent cleavage of Mcl-1, and inhibited tyrosine phosphorylation of β-catenin to decrease its stability and pro-survival functions. In vivo, ponatinib abrogated the growth of xenografted BaF3-T674I FIP1L1-PDGFRα cells in nude mice.

Conclusions: Ponatinib is a pan-FIP1L1-PDGFRα inhibitor, and clinical trials are warranted to investigate its efficacy in imatinib-resistant CEL.

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Ponatinib inhibits the growth of neoplastic cells expressing PDGFRα. (A) Ponatinib inhibited the cell viability of FIP1LI-PDGFRα-expressing cells. EOL-1 and BaF3-WT or -T674I FIP1L1-PDGFRα cells were exposed to increasing concentrations of ponatinib, sorafenib or imatinib for 72 h, and cell viability was evaluated by MTS assay. Graphs show data from 3 independent experiments; error bars represent 95% confidence intervals. (B) Clonogenicity of BaF3-WT or -T674I FIP1L1-PDGFRα cells was inhibited by ponatinib in a concentration-dependent manner. Error bars represent 95% confidence intervals. (C) Effect of ponatinib on cell cycle distribution in CEL cells. CEL cells were exposed to ponatinib for 24 h. Cells were fixed and analyzed by FACScalibur after staining with propidium iodide. Histograms are from representative experiments.
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Figure 2: Ponatinib inhibits the growth of neoplastic cells expressing PDGFRα. (A) Ponatinib inhibited the cell viability of FIP1LI-PDGFRα-expressing cells. EOL-1 and BaF3-WT or -T674I FIP1L1-PDGFRα cells were exposed to increasing concentrations of ponatinib, sorafenib or imatinib for 72 h, and cell viability was evaluated by MTS assay. Graphs show data from 3 independent experiments; error bars represent 95% confidence intervals. (B) Clonogenicity of BaF3-WT or -T674I FIP1L1-PDGFRα cells was inhibited by ponatinib in a concentration-dependent manner. Error bars represent 95% confidence intervals. (C) Effect of ponatinib on cell cycle distribution in CEL cells. CEL cells were exposed to ponatinib for 24 h. Cells were fixed and analyzed by FACScalibur after staining with propidium iodide. Histograms are from representative experiments.

Mentions: We examined the effect of TKIs on cell viability (MTS assay). The three lines of FIP1L1-PDGFRα-expressing cells were incubated with or without increasing concentrations of ponatinib, sorafenib, or imatinib for 72 h; log concentration-response curves are shown in Figure 2A. EOL-1 and BaF3-WT FIP1L1-PDGFRα cells were sensitive to imatinib, with IC50 values of 0.3 and 2.8 nM, respectively. BaF3-T674I FIP1L1-PDGFRα cells were resistant to imatinib, similar to previous reports [1,2]. Ponatinib inhibited the growth of all three FIP1L1-PDGFRα-expressing cell lines, with IC50 values of 0.004-2.5 nM. Notably, BaF3-T674I FIP1L1-PDGFRα cells were about 100-fold more sensitive to ponatinib than to sorafenib (IC50 = 2.5 versus 250 nM, respectively).


Ponatinib efficiently kills imatinib-resistant chronic eosinophilic leukemia cells harboring gatekeeper mutant T674I FIP1L1-PDGFRα: roles of Mcl-1 and β-catenin.

Jin Y, Ding K, Li H, Xue M, Shi X, Wang C, Pan J - Mol. Cancer (2014)

Ponatinib inhibits the growth of neoplastic cells expressing PDGFRα. (A) Ponatinib inhibited the cell viability of FIP1LI-PDGFRα-expressing cells. EOL-1 and BaF3-WT or -T674I FIP1L1-PDGFRα cells were exposed to increasing concentrations of ponatinib, sorafenib or imatinib for 72 h, and cell viability was evaluated by MTS assay. Graphs show data from 3 independent experiments; error bars represent 95% confidence intervals. (B) Clonogenicity of BaF3-WT or -T674I FIP1L1-PDGFRα cells was inhibited by ponatinib in a concentration-dependent manner. Error bars represent 95% confidence intervals. (C) Effect of ponatinib on cell cycle distribution in CEL cells. CEL cells were exposed to ponatinib for 24 h. Cells were fixed and analyzed by FACScalibur after staining with propidium iodide. Histograms are from representative experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3928078&req=5

Figure 2: Ponatinib inhibits the growth of neoplastic cells expressing PDGFRα. (A) Ponatinib inhibited the cell viability of FIP1LI-PDGFRα-expressing cells. EOL-1 and BaF3-WT or -T674I FIP1L1-PDGFRα cells were exposed to increasing concentrations of ponatinib, sorafenib or imatinib for 72 h, and cell viability was evaluated by MTS assay. Graphs show data from 3 independent experiments; error bars represent 95% confidence intervals. (B) Clonogenicity of BaF3-WT or -T674I FIP1L1-PDGFRα cells was inhibited by ponatinib in a concentration-dependent manner. Error bars represent 95% confidence intervals. (C) Effect of ponatinib on cell cycle distribution in CEL cells. CEL cells were exposed to ponatinib for 24 h. Cells were fixed and analyzed by FACScalibur after staining with propidium iodide. Histograms are from representative experiments.
Mentions: We examined the effect of TKIs on cell viability (MTS assay). The three lines of FIP1L1-PDGFRα-expressing cells were incubated with or without increasing concentrations of ponatinib, sorafenib, or imatinib for 72 h; log concentration-response curves are shown in Figure 2A. EOL-1 and BaF3-WT FIP1L1-PDGFRα cells were sensitive to imatinib, with IC50 values of 0.3 and 2.8 nM, respectively. BaF3-T674I FIP1L1-PDGFRα cells were resistant to imatinib, similar to previous reports [1,2]. Ponatinib inhibited the growth of all three FIP1L1-PDGFRα-expressing cell lines, with IC50 values of 0.004-2.5 nM. Notably, BaF3-T674I FIP1L1-PDGFRα cells were about 100-fold more sensitive to ponatinib than to sorafenib (IC50 = 2.5 versus 250 nM, respectively).

Bottom Line: Therefore, novel TKIs effective against T674I FIP1L1-PDGFRα are needed.The purpose of this study was to examine the effect of ponatinib on T674I FIP1L1-PDGFRα.It induced apoptosis in CEL cells with caspase-3-dependent cleavage of Mcl-1, and inhibited tyrosine phosphorylation of β-catenin to decrease its stability and pro-survival functions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathophysiology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China. panjx2@mail.sysu.edu.cn.

ABSTRACT

Background: T674I FIP1L1-PDGFRα in a subset of chronic eosinophilic leukemia (CEL) is a gatekeeper mutation that is resistant to many tyrosine kinase inhibitors (TKIs) (e.g., imatinib, nilotinib and dasatinib), similar to T315I Bcr-Abl. Therefore, novel TKIs effective against T674I FIP1L1-PDGFRα are needed. Ponatinib (AP24534) is a novel orally bioavailable TKI against T315I Bcr-Abl, but it is not clear whether ponatinib is effective against T674I FIP1L1-PDGFRα. The purpose of this study was to examine the effect of ponatinib on T674I FIP1L1-PDGFRα.

Methods: Molecular docking analysis in silico was performed. The effects of ponatinib on PDGFRα signaling pathways, apoptosis and cell cycling were examined in EOL-1, BaF3 cells expressing either wild type (WT) or T674I FIP1L1-PDGFRα. The in vivo antitumor activity of ponatinib was evaluated with xenografted BaF3-T674I FIP1L1-PDGFRα cells in nude mice models.

Results: Molecular docking analysis revealed that ponatinib could bind to the DFG (Asp-Phe-Gly)-out state of T674I PDGFRα. Ponatinib potently inhibited the phosphorylation of WT and T674I FIP1L1-PDGFRα and their downstream signaling molecules (e.g., Stat3, Stat5). Ponatinib strikingly inhibited the growth of both WT and T674I FIP1L1-PDGFRα-carrying CEL cells (IC50: 0.004-2.5 nM). It induced apoptosis in CEL cells with caspase-3-dependent cleavage of Mcl-1, and inhibited tyrosine phosphorylation of β-catenin to decrease its stability and pro-survival functions. In vivo, ponatinib abrogated the growth of xenografted BaF3-T674I FIP1L1-PDGFRα cells in nude mice.

Conclusions: Ponatinib is a pan-FIP1L1-PDGFRα inhibitor, and clinical trials are warranted to investigate its efficacy in imatinib-resistant CEL.

Show MeSH
Related in: MedlinePlus