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Omeprazole inhibits proliferation and modulates autophagy in pancreatic cancer cells.

Udelnow A, Kreyes A, Ellinger S, Landfester K, Walther P, Klapperstueck T, Wohlrab J, Henne-Bruns D, Knippschild U, Würl P - PLoS ONE (2011)

Bottom Line: Gene expression of the autophagy-related LC3 gene as well as of Bad, Mdr-1, Atg12 and the vATPase was analysed after treatment of cells with 5-fluorouracil and omeprazole and confirmed the above mentioned results.We hypothesise that omeprazole interacts with the regulatory functions of the vATPase without inhibiting its pump function.Since omeprazole use has already been established in clinical practice these results could lead to new clinical applications.

View Article: PubMed Central - PubMed

Affiliation: Department of General, Visceral and Transplantation Surgery, University Hospital of Ulm, Ulm, Germany.

ABSTRACT

Background: Omeprazole has recently been described as a modulator of tumour chemoresistance, although its underlying molecular mechanisms remain controversial. Since pancreatic tumours are highly chemoresistant, a logical step would be to investigate the pharmacodynamic, morphological and biochemical effects of omeprazole on pancreatic cancer cell lines.

Methodology/principal findings: Dose-effect curves of omeprazole, pantoprazole, gemcitabine, 5-fluorouracil and the combinations of omeprazole and 5-fluorouracil or gemcitabine were generated for the pancreatic cancer cell lines MiaPaCa-2, ASPC-1, Colo357, PancTu-1, Panc1 and Panc89. They revealed that omeprazole inhibited proliferation at probably non-toxic concentrations and reversed the hormesis phenomena of 5-fluorouracil. Electron microscopy showed that omeprazole led to accumulation of phagophores and early autophagosomes in ASPC-1 and MiaPaCa-2 cells. Signal changes indicating inhibited proliferation and programmed cell death were found by proton NMR spectroscopy of both cell lines when treated with omeprazole which was identified intracellularly. Omeprazole modulates the lysosomal transport pathway as shown by Western blot analysis of the expression of LAMP-1, Cathepsin-D and β-COP in lysosome- and Golgi complex containing cell fractions. Acridine orange staining revealed that the pump function of the vATPase was not specifically inhibited by omeprazole. Gene expression of the autophagy-related LC3 gene as well as of Bad, Mdr-1, Atg12 and the vATPase was analysed after treatment of cells with 5-fluorouracil and omeprazole and confirmed the above mentioned results.

Conclusions: We hypothesise that omeprazole interacts with the regulatory functions of the vATPase without inhibiting its pump function. A modulation of the lysosomal transport pathway and autophagy is caused in pancreatic cancer cells leading to programmed cell death. This may circumvent common resistance mechanisms of pancreatic cancer. Since omeprazole use has already been established in clinical practice these results could lead to new clinical applications.

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Identification of substances from a proton NMR spectrum of viable untreated MiaPaCa-2 cells.The cells were harvested from monolayer culture, kept and measured at 20°C. Measurement were performed by a 600 MHz Bruker spectrometer. For better visibility the part of the spectrum showing the protons of aliphatic groups is splitted into 2 parts - A and B. (A). aliphatic part I. The methyl and β- and γ- methylene groups of various fatty acids and amino acids are visible. In addition, isopropanol and tetrachlorethan (the external concentration standard) occured as pollutions. (B) Aliphatic part II. Phospholipid metabolites and the α-methylene groups of amino acids and lactate are visible. (C) Formula of OMP with numbering of the respective protons. The methyl groups (1–3, 9) and the methyl group (4) are covered by other metabolites in the aliphatic parts of the spectrum. In contrast, the aromatic protons are visible (H5, H8, H10). (D) Overlay of the aromatic parts of different spectra for intracellular identification of OMP. The singulet of the H5 proton and the dublets of the H8 and H10 protons can be identified when the medium and the cell spectra are compared to those without OMP treatment. Abbreviations: His - histidine, Tyr - tyrosine, Phe – phenylalanine, Leu, Ile, Val - Leucine, Isoleucine, Valine. Ala - Alanine. Glu - Glutamate, Gln – Glutamine, PC - phosphatidylcholine, Cho - Choline, GPC – glycerophosphocholine, Tau – taurine, Scyllo - scylloinositole.
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pone-0020143-g005: Identification of substances from a proton NMR spectrum of viable untreated MiaPaCa-2 cells.The cells were harvested from monolayer culture, kept and measured at 20°C. Measurement were performed by a 600 MHz Bruker spectrometer. For better visibility the part of the spectrum showing the protons of aliphatic groups is splitted into 2 parts - A and B. (A). aliphatic part I. The methyl and β- and γ- methylene groups of various fatty acids and amino acids are visible. In addition, isopropanol and tetrachlorethan (the external concentration standard) occured as pollutions. (B) Aliphatic part II. Phospholipid metabolites and the α-methylene groups of amino acids and lactate are visible. (C) Formula of OMP with numbering of the respective protons. The methyl groups (1–3, 9) and the methyl group (4) are covered by other metabolites in the aliphatic parts of the spectrum. In contrast, the aromatic protons are visible (H5, H8, H10). (D) Overlay of the aromatic parts of different spectra for intracellular identification of OMP. The singulet of the H5 proton and the dublets of the H8 and H10 protons can be identified when the medium and the cell spectra are compared to those without OMP treatment. Abbreviations: His - histidine, Tyr - tyrosine, Phe – phenylalanine, Leu, Ile, Val - Leucine, Isoleucine, Valine. Ala - Alanine. Glu - Glutamate, Gln – Glutamine, PC - phosphatidylcholine, Cho - Choline, GPC – glycerophosphocholine, Tau – taurine, Scyllo - scylloinositole.

Mentions: Proton NMR spectroscopy of viable cells was performed in MiaPaCa-2 and ASPC-1 cell lines in order to analyse the biochemical processes associated with the morphological changes described above. Various characteristic low-molecular intracellular metabolites such as fatty acids, amino acids, membrane associated phospholipid mebolites and intermediate citrate cycle and glycolysis metabolites were identified (Figure 5).


Omeprazole inhibits proliferation and modulates autophagy in pancreatic cancer cells.

Udelnow A, Kreyes A, Ellinger S, Landfester K, Walther P, Klapperstueck T, Wohlrab J, Henne-Bruns D, Knippschild U, Würl P - PLoS ONE (2011)

Identification of substances from a proton NMR spectrum of viable untreated MiaPaCa-2 cells.The cells were harvested from monolayer culture, kept and measured at 20°C. Measurement were performed by a 600 MHz Bruker spectrometer. For better visibility the part of the spectrum showing the protons of aliphatic groups is splitted into 2 parts - A and B. (A). aliphatic part I. The methyl and β- and γ- methylene groups of various fatty acids and amino acids are visible. In addition, isopropanol and tetrachlorethan (the external concentration standard) occured as pollutions. (B) Aliphatic part II. Phospholipid metabolites and the α-methylene groups of amino acids and lactate are visible. (C) Formula of OMP with numbering of the respective protons. The methyl groups (1–3, 9) and the methyl group (4) are covered by other metabolites in the aliphatic parts of the spectrum. In contrast, the aromatic protons are visible (H5, H8, H10). (D) Overlay of the aromatic parts of different spectra for intracellular identification of OMP. The singulet of the H5 proton and the dublets of the H8 and H10 protons can be identified when the medium and the cell spectra are compared to those without OMP treatment. Abbreviations: His - histidine, Tyr - tyrosine, Phe – phenylalanine, Leu, Ile, Val - Leucine, Isoleucine, Valine. Ala - Alanine. Glu - Glutamate, Gln – Glutamine, PC - phosphatidylcholine, Cho - Choline, GPC – glycerophosphocholine, Tau – taurine, Scyllo - scylloinositole.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3101238&req=5

pone-0020143-g005: Identification of substances from a proton NMR spectrum of viable untreated MiaPaCa-2 cells.The cells were harvested from monolayer culture, kept and measured at 20°C. Measurement were performed by a 600 MHz Bruker spectrometer. For better visibility the part of the spectrum showing the protons of aliphatic groups is splitted into 2 parts - A and B. (A). aliphatic part I. The methyl and β- and γ- methylene groups of various fatty acids and amino acids are visible. In addition, isopropanol and tetrachlorethan (the external concentration standard) occured as pollutions. (B) Aliphatic part II. Phospholipid metabolites and the α-methylene groups of amino acids and lactate are visible. (C) Formula of OMP with numbering of the respective protons. The methyl groups (1–3, 9) and the methyl group (4) are covered by other metabolites in the aliphatic parts of the spectrum. In contrast, the aromatic protons are visible (H5, H8, H10). (D) Overlay of the aromatic parts of different spectra for intracellular identification of OMP. The singulet of the H5 proton and the dublets of the H8 and H10 protons can be identified when the medium and the cell spectra are compared to those without OMP treatment. Abbreviations: His - histidine, Tyr - tyrosine, Phe – phenylalanine, Leu, Ile, Val - Leucine, Isoleucine, Valine. Ala - Alanine. Glu - Glutamate, Gln – Glutamine, PC - phosphatidylcholine, Cho - Choline, GPC – glycerophosphocholine, Tau – taurine, Scyllo - scylloinositole.
Mentions: Proton NMR spectroscopy of viable cells was performed in MiaPaCa-2 and ASPC-1 cell lines in order to analyse the biochemical processes associated with the morphological changes described above. Various characteristic low-molecular intracellular metabolites such as fatty acids, amino acids, membrane associated phospholipid mebolites and intermediate citrate cycle and glycolysis metabolites were identified (Figure 5).

Bottom Line: Gene expression of the autophagy-related LC3 gene as well as of Bad, Mdr-1, Atg12 and the vATPase was analysed after treatment of cells with 5-fluorouracil and omeprazole and confirmed the above mentioned results.We hypothesise that omeprazole interacts with the regulatory functions of the vATPase without inhibiting its pump function.Since omeprazole use has already been established in clinical practice these results could lead to new clinical applications.

View Article: PubMed Central - PubMed

Affiliation: Department of General, Visceral and Transplantation Surgery, University Hospital of Ulm, Ulm, Germany.

ABSTRACT

Background: Omeprazole has recently been described as a modulator of tumour chemoresistance, although its underlying molecular mechanisms remain controversial. Since pancreatic tumours are highly chemoresistant, a logical step would be to investigate the pharmacodynamic, morphological and biochemical effects of omeprazole on pancreatic cancer cell lines.

Methodology/principal findings: Dose-effect curves of omeprazole, pantoprazole, gemcitabine, 5-fluorouracil and the combinations of omeprazole and 5-fluorouracil or gemcitabine were generated for the pancreatic cancer cell lines MiaPaCa-2, ASPC-1, Colo357, PancTu-1, Panc1 and Panc89. They revealed that omeprazole inhibited proliferation at probably non-toxic concentrations and reversed the hormesis phenomena of 5-fluorouracil. Electron microscopy showed that omeprazole led to accumulation of phagophores and early autophagosomes in ASPC-1 and MiaPaCa-2 cells. Signal changes indicating inhibited proliferation and programmed cell death were found by proton NMR spectroscopy of both cell lines when treated with omeprazole which was identified intracellularly. Omeprazole modulates the lysosomal transport pathway as shown by Western blot analysis of the expression of LAMP-1, Cathepsin-D and β-COP in lysosome- and Golgi complex containing cell fractions. Acridine orange staining revealed that the pump function of the vATPase was not specifically inhibited by omeprazole. Gene expression of the autophagy-related LC3 gene as well as of Bad, Mdr-1, Atg12 and the vATPase was analysed after treatment of cells with 5-fluorouracil and omeprazole and confirmed the above mentioned results.

Conclusions: We hypothesise that omeprazole interacts with the regulatory functions of the vATPase without inhibiting its pump function. A modulation of the lysosomal transport pathway and autophagy is caused in pancreatic cancer cells leading to programmed cell death. This may circumvent common resistance mechanisms of pancreatic cancer. Since omeprazole use has already been established in clinical practice these results could lead to new clinical applications.

Show MeSH
Related in: MedlinePlus