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EGCG inhibits growth of human pancreatic tumors orthotopically implanted in Balb C nude mice through modulation of FKHRL1/FOXO3a and neuropilin

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Abstract

Human pancreatic cancer is currently one of the fourth leading causes of cancer-related mortality with a 5-year survival rate of less than 5 %. Since pancreatic carcinoma is largely refractory to conventional therapies, there is a strong medical need for the development of novel and innovative cancer preventive strategies. The forkhead transcription factors of the O class (FOXO) play a major role in cell proliferation, angiogenesis, metastasis, and tumorigenesis. The objectives of this study were to examine whether FKHRL1/FOXO3a modulates antitumor activity of (−)-epigallocatechin-3-gallate (EGCG), an active ingredient in green tea, in pancreatic cancer model in vivo. PANC-1 cells were orthotopically implanted into Balb c nude mice and gavaged with EGCG after tumor formation. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of PI3K, AKT, ERK, and FOXO3a/FKHRL1 and its target genes were measured by the western blot analysis and/or q-RT-PCR. FOXO-DNA binding was measured by gel shift assay. EGCG-treated mice showed significant inhibition in tumor growth which was associated with reduced phosphorylation of ERK, PI3K, AKT, and FKHRL1/FOXO3a, and modulation of FOXO target genes. EGCG induced apoptosis by upregulating Bim and activating caspase-3. EGCG modulated markers of cell cycle (p27/KIP1), angiogenesis (CD31, VEGF, IL-6, IL-8, SEMA3F, and HIF1α), and metastasis (MMP2 and MMP7). The inhibition of VEGF by EGCG was associated with suppression of neuropilin. EGCG inhibited epithelial-mesenchymal transition by upregulating the expression of E-cadherin and inhibiting the expression of N-cadherin and Zeb1. These data suggest that EGCG inhibits pancreatic cancer orthotopic tumor growth, angiogenesis, and metastasis which are associated with inhibition of PI3K/AKT and ERK pathways and activation of FKHRL1/FOXO3a. As a conclusion, EGCG can be used for the prevention and/or treatment of pancreatic cancer.

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Abbreviations

bHLH:

Basic helix-loop-helix

Cbp:

CREB-binding protein

EC:

Endothelial cells

ECM:

Extracellular matrix

EGCG:

(−)-Epigallocatechin-3-gallate

ERK1/2:

Extracellular signal-regulated kinase

EMT:

Epithelial-mesenchymal transition

FOXO:

Forkhead transcription factors of the O class

HREs:

Hypoxia response elements

HIF:

Hypoxia-inducible factor

MMP:

Matrix metalloproteinases

NRP2:

Neuropilin-2

PI3K:

Phosphoinositide 3-kinase

PDA:

Pancreatic ductal adenocarcinoma

Pcaf:

p300/CBP-associated factors

SEMA 3F:

Semaphorin 3F

VEGF:

Vascular endothelial growth factor

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Acknowledgments

We thank our lab members for critical reading of the manuscript. This work was supported in part by the Grants from the National Institutes of Health (R01CA125262, RO1CA114469 and RO1CA125262-02S1) and Kansas Bioscience Authority.

Conflict of interest

The authors have declared no conflict of interest to disclose.

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Authors and Affiliations

  1. Department of Pathology and Laboratory Medicine, The University of Kansas Cancer Center, The University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA

    Sharmila Shankar

  2. Department of Biochemistry, University of Texas Health Science Center at Tyler, Tyler, TX, 75708, USA

    Luke Marsh

  3. Department of Pharmacology, Toxicology and Therapeutics, and Medicine, The University of Kansas Cancer Center, The University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA

    Rakesh K. Srivastava

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  1. Sharmila Shankar
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Correspondence to Rakesh K. Srivastava.

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Shankar, S., Marsh, L. & Srivastava, R.K. EGCG inhibits growth of human pancreatic tumors orthotopically implanted in Balb C nude mice through modulation of FKHRL1/FOXO3a and neuropilin. Mol Cell Biochem 372, 83–94 (2013). https://doi.org/10.1007/s11010-012-1448-y

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