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Ets-1 expression and gemcitabine chemoresistance in pancreatic cancer cells

Cellular & Molecular Biology Letters volume 16, pages 101–113 (2011)Cite this article

Abstract

Gemcitabine, a novel pyrimidine nucleoside analog, has become the standard chemotherapeutic agent for pancreatic cancer patients. The clinical impact of gemcitabine remains modest owing to the high degree of inherent and acquired resistance. There are various lines of evidence that confirm the role of Ets-1, a proto-oncoprotein, in tumor invasion, progression, and chemoresistance. This study examines a hypothesis that implicates Ets-1 in the development of gemcitabine-resistance in pancreatic cancer cells. Ets-1 protein expression was assessed in parental pancreatic cancer cells and their gemcitabine-resistant clones. Western blot analysis revealed elevated levels of Ets-1 protein expression in gemcitabine-resistant PANC1GemRes (4.8-fold increase; P < 0.05), MIA PaCa2GemRes (3.2-fold increase; P < 0.05), and Capan2GemRes (2.1-fold increase; P < 0.05) cells as compared to their parental counterparts. A time course analysis was conducted to determine the change in Ets-1 expression in the parental cells after incubation with gemcitabine. Reverse transcriptase quantitative real-time PCR (RT-qPCR) and Western blot analysis revealed a significant increase in Ets-1 expression. All the three parental cells incubated with gemcitabine showed elevated Ets-1 protein expression at 6 h. By 24 h, the expression level had decreased. Using small interfering RNA (siRNA) against Ets-1 in gemcitabine-resistant cells, we demonstrated a reversal in gemcitabine chemosensitivity and also detected a marked reduction in the expression of the Ets-1 target genes MMP1 and uPA. Our novel finding demonstrates the significance of Ets-1 in the development of gemcitabine chemoresistance in pancreatic cancer cells. Based on these results, a new siRNA-based therapeutic strategy targeting the Ets-1 genes can be designed to overcome chemoresistance.

Abbreviations

cDNA:

complementary DNA

Ct:

cycle threshold

dFdCTP:

2′,2′-difluorodeoxycytidine 5′-triphosphate (gemcitabine triphosphate)

DMEM:

Dulbecco’s modified Eagle’s medium

DMSO:

dimethyl sulfoxide

E26:

avian erythroblastosis virus

Ets-1:

E26 transformation specific sequence-1

FBS:

fetal bovine serum

GAPDH:

glyceraldehydes-3-phosphate dehydrogenase

MDR1:

multiple drug resistance-1

MMP1:

matrix metalloproteinase-1 (collagenase-1)

MMP3:

matrix metallopeptidase-1 (stromelysin-1)

MTT:

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

PDAC:

pancreatic ductal adenocarcinoma

RT-qPCR:

reverse transcriptase real-time quantitative PCR

siRNA:

small interfering RNA

uPA:

urokinase-type plasminogen activator

v-ets:

viral transforming gene of E26

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Authors and Affiliations

  1. Department of High Throughput Screening and Biotechnology, Piramal Life Sciences Ltd., 1A, Nirlon Complex, Goregaon-E, Mumbai, 400063, Maharashtra, India

    Amit Khanna & Giridharan Periyasamy

  2. Department of Biomarker Discovery, Piramal Life Sciences Ltd., 1A, Nirlon Complex, Goregaon-E, Mumbai, 400063, Maharashtra, India

    Debarshi Chakrabarti

  3. School of Biotechnology, Chemical and Bio-Medical Engineering, VIT University, Vellore, Tamil Nadu, India

    Amit Khanna & Kulandaivelu Mahalingam

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  1. Amit Khanna
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  2. Kulandaivelu Mahalingam
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  3. Debarshi Chakrabarti
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  4. Giridharan Periyasamy
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Correspondence to Giridharan Periyasamy.

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Khanna, A., Mahalingam, K., Chakrabarti, D. et al. Ets-1 expression and gemcitabine chemoresistance in pancreatic cancer cells. Cell Mol Biol Lett 16, 101–113 (2011). https://doi.org/10.2478/s11658-010-0043-z

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Key words

Cellular & Molecular Biology Letters

ISSN: 1689-1392

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