Resistance to mitogen-activated protein kinase kinase (MEK) inhibitors correlates with up-regulation of the MEK/extracellular signal-regulated kinase pathway in hepatocellular carcinoma cells

Michele Yip-Schneider, Patrick J. Klein, Sabrina C. Wentz, Amer Zeni, Alex Menze, C. Schmidt

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Abstract

The extracellular signal-regulated (ERK), mitogen-activated protein kinase (p42/p44 MAPK) pathway is up-regulated in hepatocellular carcinoma (HCC). Molecular targeting of this critical mitogenic pathway may have therapeutic potential for the treatment of HCC; however, chemoresistance to long-term therapy may develop. In the present study, we employed small-molecule MAPK kinase (MEK) inhibitors, including U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2- aminophynyltio)butadiene] and PD184161 (Neoplasia 8:1-8, 2006), in HepG2 and Hep3B human HCC cell lines to identify potential mechanism(s) of resistance. U0126 dose-dependently suppressed ERK phosphorylation at both 1- and 24-h time points in HepG2 cells, previously shown to be sensitive to growth inhibition by U0126. In contrast, ERK phosphorylation was only decreased at the 1-h time point but not at 24 h in the more resistant Hep3B cells. It is interesting that the lack of prolonged phospho-ERK suppression was associated with MEK hyperphosphorylation in Hep3B cells. Several MEK/ERK pathway intermediates were up-regulated in Hep3B cells; furthermore, transfection of Raf-1 small interfering RNA to suppress MEK/ERK pathway activation sensitized Hep3B cells to U0126. MEK inhibitor resistance was independent of p53 or hepatitis Bx protein status. Finally, we showed that combining two chemically distinct MEK inhibitors enhanced growth inhibition and apoptosis compared with the single agents. Taken together, these results suggest that upregulated expression or activity of the MEK/ERK pathway contributes to MEK inhibitor resistance in HCC cells. Our findings also provide preclinical evidence suggesting that the status of the MEK/ERK pathway in patients may predict response to MEK/ERK-targeted therapeutics.

Original languageEnglish
Pages (from-to)1063-1070
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume329
Issue number3
DOIs
StatePublished - Jun 2009

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Mitogen-Activated Protein Kinase Kinases
Extracellular Signal-Regulated MAP Kinases
Hepatocellular Carcinoma
Up-Regulation
MAP Kinase Signaling System
Phosphorylation
Growth Inhibitors
Critical Pathways
Mitogen-Activated Protein Kinase 3
Mitogen-Activated Protein Kinase 1
Hep G2 Cells
Therapeutics
Mitogen-Activated Protein Kinases
Small Interfering RNA
Hepatitis
Transfection
Apoptosis
Cell Line
U 0126

ASJC Scopus subject areas

  • Pharmacology
  • Molecular Medicine

Cite this

@article{41733242621248fd8c406b800cbd6b71,
title = "Resistance to mitogen-activated protein kinase kinase (MEK) inhibitors correlates with up-regulation of the MEK/extracellular signal-regulated kinase pathway in hepatocellular carcinoma cells",
abstract = "The extracellular signal-regulated (ERK), mitogen-activated protein kinase (p42/p44 MAPK) pathway is up-regulated in hepatocellular carcinoma (HCC). Molecular targeting of this critical mitogenic pathway may have therapeutic potential for the treatment of HCC; however, chemoresistance to long-term therapy may develop. In the present study, we employed small-molecule MAPK kinase (MEK) inhibitors, including U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2- aminophynyltio)butadiene] and PD184161 (Neoplasia 8:1-8, 2006), in HepG2 and Hep3B human HCC cell lines to identify potential mechanism(s) of resistance. U0126 dose-dependently suppressed ERK phosphorylation at both 1- and 24-h time points in HepG2 cells, previously shown to be sensitive to growth inhibition by U0126. In contrast, ERK phosphorylation was only decreased at the 1-h time point but not at 24 h in the more resistant Hep3B cells. It is interesting that the lack of prolonged phospho-ERK suppression was associated with MEK hyperphosphorylation in Hep3B cells. Several MEK/ERK pathway intermediates were up-regulated in Hep3B cells; furthermore, transfection of Raf-1 small interfering RNA to suppress MEK/ERK pathway activation sensitized Hep3B cells to U0126. MEK inhibitor resistance was independent of p53 or hepatitis Bx protein status. Finally, we showed that combining two chemically distinct MEK inhibitors enhanced growth inhibition and apoptosis compared with the single agents. Taken together, these results suggest that upregulated expression or activity of the MEK/ERK pathway contributes to MEK inhibitor resistance in HCC cells. Our findings also provide preclinical evidence suggesting that the status of the MEK/ERK pathway in patients may predict response to MEK/ERK-targeted therapeutics.",
author = "Michele Yip-Schneider and Klein, {Patrick J.} and Wentz, {Sabrina C.} and Amer Zeni and Alex Menze and C. Schmidt",
year = "2009",
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T1 - Resistance to mitogen-activated protein kinase kinase (MEK) inhibitors correlates with up-regulation of the MEK/extracellular signal-regulated kinase pathway in hepatocellular carcinoma cells

AU - Yip-Schneider, Michele

AU - Klein, Patrick J.

AU - Wentz, Sabrina C.

AU - Zeni, Amer

AU - Menze, Alex

AU - Schmidt, C.

PY - 2009/6

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N2 - The extracellular signal-regulated (ERK), mitogen-activated protein kinase (p42/p44 MAPK) pathway is up-regulated in hepatocellular carcinoma (HCC). Molecular targeting of this critical mitogenic pathway may have therapeutic potential for the treatment of HCC; however, chemoresistance to long-term therapy may develop. In the present study, we employed small-molecule MAPK kinase (MEK) inhibitors, including U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2- aminophynyltio)butadiene] and PD184161 (Neoplasia 8:1-8, 2006), in HepG2 and Hep3B human HCC cell lines to identify potential mechanism(s) of resistance. U0126 dose-dependently suppressed ERK phosphorylation at both 1- and 24-h time points in HepG2 cells, previously shown to be sensitive to growth inhibition by U0126. In contrast, ERK phosphorylation was only decreased at the 1-h time point but not at 24 h in the more resistant Hep3B cells. It is interesting that the lack of prolonged phospho-ERK suppression was associated with MEK hyperphosphorylation in Hep3B cells. Several MEK/ERK pathway intermediates were up-regulated in Hep3B cells; furthermore, transfection of Raf-1 small interfering RNA to suppress MEK/ERK pathway activation sensitized Hep3B cells to U0126. MEK inhibitor resistance was independent of p53 or hepatitis Bx protein status. Finally, we showed that combining two chemically distinct MEK inhibitors enhanced growth inhibition and apoptosis compared with the single agents. Taken together, these results suggest that upregulated expression or activity of the MEK/ERK pathway contributes to MEK inhibitor resistance in HCC cells. Our findings also provide preclinical evidence suggesting that the status of the MEK/ERK pathway in patients may predict response to MEK/ERK-targeted therapeutics.

AB - The extracellular signal-regulated (ERK), mitogen-activated protein kinase (p42/p44 MAPK) pathway is up-regulated in hepatocellular carcinoma (HCC). Molecular targeting of this critical mitogenic pathway may have therapeutic potential for the treatment of HCC; however, chemoresistance to long-term therapy may develop. In the present study, we employed small-molecule MAPK kinase (MEK) inhibitors, including U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2- aminophynyltio)butadiene] and PD184161 (Neoplasia 8:1-8, 2006), in HepG2 and Hep3B human HCC cell lines to identify potential mechanism(s) of resistance. U0126 dose-dependently suppressed ERK phosphorylation at both 1- and 24-h time points in HepG2 cells, previously shown to be sensitive to growth inhibition by U0126. In contrast, ERK phosphorylation was only decreased at the 1-h time point but not at 24 h in the more resistant Hep3B cells. It is interesting that the lack of prolonged phospho-ERK suppression was associated with MEK hyperphosphorylation in Hep3B cells. Several MEK/ERK pathway intermediates were up-regulated in Hep3B cells; furthermore, transfection of Raf-1 small interfering RNA to suppress MEK/ERK pathway activation sensitized Hep3B cells to U0126. MEK inhibitor resistance was independent of p53 or hepatitis Bx protein status. Finally, we showed that combining two chemically distinct MEK inhibitors enhanced growth inhibition and apoptosis compared with the single agents. Taken together, these results suggest that upregulated expression or activity of the MEK/ERK pathway contributes to MEK inhibitor resistance in HCC cells. Our findings also provide preclinical evidence suggesting that the status of the MEK/ERK pathway in patients may predict response to MEK/ERK-targeted therapeutics.

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