Treatment of breast and lung cancer cells with a N-7 benzyl guanosine monophosphate tryptamine phosphoramidate pronucleotide (4Ei-1) results in chemosensitization to gemcitabine and induced eIF4E proteasomal degradation

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Abstract

The development of cancer and fibrotic diseases has been shown to be highly dependent on disregulation of cap-dependent translation. Binding protein eIF4E to N7-methylated guanosine capped mRNA has been found to be the rate-limiting step governing translation initiation, and therefore represents an attractive target for drug discovery. Our group has found that 7-benzyl guanosine monophosphate (7Bn-GMP) is a potent antagonist of eIF4E cap binding (Kd = 0.8 μM). Recent X-ray crystallographic studies have revealed that the cap-dependent pocket undergoes a unique structural change in order to accommodate the benzyl group. Unfortunately, 7Bn-GMP is not cell permeable. Recently, we have prepared a tryptamine phosphoramidate prodrug of 7Bn-GMP, 4Ei-1, and shown that it is a substrate for human histidine triad nucleotide binding protein (hHINT1) and inhibits eIF4E initiated epithelial-mesenchymal transition (EMT) by Zebra fish embryo cells. To assess the intracellular uptake of 4Ei-1 and conversion to 7Bn-GMP by cancer cells, we developed a sensitive assay using LC-ESI-MS/MS for the intracellular quantitation of 4Ei-1 and 7Bn-GMP. When incubated with the breast cancer cell line MDA-231 or lung cancer cell lines H460, H383 and H2009, 4Ei-1 was found to be rapidly internalized and converted to 7Bn-GMP. Since oncogenic mRNAs are predicted to have the highest eIF4E requirement for translation, we carried out chemosensitization studies with 4Ei-1. The prodrug was found to chemosensitize both breast and lung cancer cells to nontoxic levels of gemcitabine. Further mechanistic studies revealed that the expressed levels of eIF4E were substantially reduced in cells treated with 4Ei-1 in a dose-dependent manner. The levels of eI4E could be restored by treatment with the proteasome inhibitor MG-132. Taken together, our results demonstrate that 4Ei-1 is likely to inhibit translation initiation by eIF4E cap binding by both antagonizing eIF4E cap binding and initiating eIF4E proteasomal degradation.

Original languageEnglish (US)
Pages (from-to)523-531
Number of pages9
JournalMolecular Pharmaceutics
Volume10
Issue number2
DOIs
StatePublished - Feb 4 2013

Fingerprint

gemcitabine
Guanosine Monophosphate
Lung Neoplasms
Breast Neoplasms
Prodrugs
Carrier Proteins
Cell Line
Messenger RNA
Proteasome Inhibitors
Epithelial-Mesenchymal Transition
Guanosine
Zebrafish
Drug Discovery
Histidine
phosphoramidic acid
tryptamine
Neoplasms
Embryonic Structures
Nucleotides
X-Rays

Keywords

  • 4Ei-1
  • 7-benzyl guanosine monophosphate
  • cancer
  • eIF4E
  • fibrotic diseases

Cite this

@article{452f6e054a254c2cab55e23d36b85def,
title = "Treatment of breast and lung cancer cells with a N-7 benzyl guanosine monophosphate tryptamine phosphoramidate pronucleotide (4Ei-1) results in chemosensitization to gemcitabine and induced eIF4E proteasomal degradation",
abstract = "The development of cancer and fibrotic diseases has been shown to be highly dependent on disregulation of cap-dependent translation. Binding protein eIF4E to N7-methylated guanosine capped mRNA has been found to be the rate-limiting step governing translation initiation, and therefore represents an attractive target for drug discovery. Our group has found that 7-benzyl guanosine monophosphate (7Bn-GMP) is a potent antagonist of eIF4E cap binding (Kd = 0.8 μM). Recent X-ray crystallographic studies have revealed that the cap-dependent pocket undergoes a unique structural change in order to accommodate the benzyl group. Unfortunately, 7Bn-GMP is not cell permeable. Recently, we have prepared a tryptamine phosphoramidate prodrug of 7Bn-GMP, 4Ei-1, and shown that it is a substrate for human histidine triad nucleotide binding protein (hHINT1) and inhibits eIF4E initiated epithelial-mesenchymal transition (EMT) by Zebra fish embryo cells. To assess the intracellular uptake of 4Ei-1 and conversion to 7Bn-GMP by cancer cells, we developed a sensitive assay using LC-ESI-MS/MS for the intracellular quantitation of 4Ei-1 and 7Bn-GMP. When incubated with the breast cancer cell line MDA-231 or lung cancer cell lines H460, H383 and H2009, 4Ei-1 was found to be rapidly internalized and converted to 7Bn-GMP. Since oncogenic mRNAs are predicted to have the highest eIF4E requirement for translation, we carried out chemosensitization studies with 4Ei-1. The prodrug was found to chemosensitize both breast and lung cancer cells to nontoxic levels of gemcitabine. Further mechanistic studies revealed that the expressed levels of eIF4E were substantially reduced in cells treated with 4Ei-1 in a dose-dependent manner. The levels of eI4E could be restored by treatment with the proteasome inhibitor MG-132. Taken together, our results demonstrate that 4Ei-1 is likely to inhibit translation initiation by eIF4E cap binding by both antagonizing eIF4E cap binding and initiating eIF4E proteasomal degradation.",
keywords = "4Ei-1, 7-benzyl guanosine monophosphate, cancer, eIF4E, fibrotic diseases",
author = "Shui Li and Yan Jia and Jacobson, {Blake A} and Joel McCauley and Kratzke, {Robert A} and Bitterman, {Peter B} and Wagner, {Carston R}",
year = "2013",
month = "2",
day = "4",
doi = "10.1021/mp300699d",
language = "English (US)",
volume = "10",
pages = "523--531",
journal = "Molecular Pharmaceutics",
issn = "1543-8384",
publisher = "American Chemical Society",
number = "2",

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TY - JOUR

T1 - Treatment of breast and lung cancer cells with a N-7 benzyl guanosine monophosphate tryptamine phosphoramidate pronucleotide (4Ei-1) results in chemosensitization to gemcitabine and induced eIF4E proteasomal degradation

AU - Li, Shui

AU - Jia, Yan

AU - Jacobson, Blake A

AU - McCauley, Joel

AU - Kratzke, Robert A

AU - Bitterman, Peter B

AU - Wagner, Carston R

PY - 2013/2/4

Y1 - 2013/2/4

N2 - The development of cancer and fibrotic diseases has been shown to be highly dependent on disregulation of cap-dependent translation. Binding protein eIF4E to N7-methylated guanosine capped mRNA has been found to be the rate-limiting step governing translation initiation, and therefore represents an attractive target for drug discovery. Our group has found that 7-benzyl guanosine monophosphate (7Bn-GMP) is a potent antagonist of eIF4E cap binding (Kd = 0.8 μM). Recent X-ray crystallographic studies have revealed that the cap-dependent pocket undergoes a unique structural change in order to accommodate the benzyl group. Unfortunately, 7Bn-GMP is not cell permeable. Recently, we have prepared a tryptamine phosphoramidate prodrug of 7Bn-GMP, 4Ei-1, and shown that it is a substrate for human histidine triad nucleotide binding protein (hHINT1) and inhibits eIF4E initiated epithelial-mesenchymal transition (EMT) by Zebra fish embryo cells. To assess the intracellular uptake of 4Ei-1 and conversion to 7Bn-GMP by cancer cells, we developed a sensitive assay using LC-ESI-MS/MS for the intracellular quantitation of 4Ei-1 and 7Bn-GMP. When incubated with the breast cancer cell line MDA-231 or lung cancer cell lines H460, H383 and H2009, 4Ei-1 was found to be rapidly internalized and converted to 7Bn-GMP. Since oncogenic mRNAs are predicted to have the highest eIF4E requirement for translation, we carried out chemosensitization studies with 4Ei-1. The prodrug was found to chemosensitize both breast and lung cancer cells to nontoxic levels of gemcitabine. Further mechanistic studies revealed that the expressed levels of eIF4E were substantially reduced in cells treated with 4Ei-1 in a dose-dependent manner. The levels of eI4E could be restored by treatment with the proteasome inhibitor MG-132. Taken together, our results demonstrate that 4Ei-1 is likely to inhibit translation initiation by eIF4E cap binding by both antagonizing eIF4E cap binding and initiating eIF4E proteasomal degradation.

AB - The development of cancer and fibrotic diseases has been shown to be highly dependent on disregulation of cap-dependent translation. Binding protein eIF4E to N7-methylated guanosine capped mRNA has been found to be the rate-limiting step governing translation initiation, and therefore represents an attractive target for drug discovery. Our group has found that 7-benzyl guanosine monophosphate (7Bn-GMP) is a potent antagonist of eIF4E cap binding (Kd = 0.8 μM). Recent X-ray crystallographic studies have revealed that the cap-dependent pocket undergoes a unique structural change in order to accommodate the benzyl group. Unfortunately, 7Bn-GMP is not cell permeable. Recently, we have prepared a tryptamine phosphoramidate prodrug of 7Bn-GMP, 4Ei-1, and shown that it is a substrate for human histidine triad nucleotide binding protein (hHINT1) and inhibits eIF4E initiated epithelial-mesenchymal transition (EMT) by Zebra fish embryo cells. To assess the intracellular uptake of 4Ei-1 and conversion to 7Bn-GMP by cancer cells, we developed a sensitive assay using LC-ESI-MS/MS for the intracellular quantitation of 4Ei-1 and 7Bn-GMP. When incubated with the breast cancer cell line MDA-231 or lung cancer cell lines H460, H383 and H2009, 4Ei-1 was found to be rapidly internalized and converted to 7Bn-GMP. Since oncogenic mRNAs are predicted to have the highest eIF4E requirement for translation, we carried out chemosensitization studies with 4Ei-1. The prodrug was found to chemosensitize both breast and lung cancer cells to nontoxic levels of gemcitabine. Further mechanistic studies revealed that the expressed levels of eIF4E were substantially reduced in cells treated with 4Ei-1 in a dose-dependent manner. The levels of eI4E could be restored by treatment with the proteasome inhibitor MG-132. Taken together, our results demonstrate that 4Ei-1 is likely to inhibit translation initiation by eIF4E cap binding by both antagonizing eIF4E cap binding and initiating eIF4E proteasomal degradation.

KW - 4Ei-1

KW - 7-benzyl guanosine monophosphate

KW - cancer

KW - eIF4E

KW - fibrotic diseases

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U2 - 10.1021/mp300699d

DO - 10.1021/mp300699d

M3 - Article

VL - 10

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JF - Molecular Pharmaceutics

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