Blockade of endothelin receptor A enhances the therapeutic efficacy of gemcitabine in pancreatic cancer cells

https://doi.org/10.1016/j.bbrc.2020.04.118Get rights and content

Highlights

  • Gemcitabine promoted ET-1/ETAR axis signaling in pancreatic cancer cells.

  • ETAR overexpression is correlated with a poor prognosis in pancreatic cancer patients.

  • ETAR antagonist enhanced the growth-inhibiting effect of gemcitabine on pancreatic cancer cells.

Abstract

Pancreatic adenocarcinoma is currently one of the leading causes of cancer-related death worldwide. The high rate of mortality in pancreatic cancer patients is due to the inability to detect early-stage disease and the disease being highly refractory to therapy. Gemcitabine has been the standard chemotherapy for advanced pancreatic cancer patients for the last two decades. However, gemcitabine resistance develops within a few weeks of treatment, and the associated mechanism remains poorly understood. Therefore, a novel therapeutic strategy is needed to overcome the limited clinical efficacy of gemcitabine in pancreatic adenocarcinoma.

In this study, we demonstrated that ET-1/ETAR axis gene expression was upregulated in pancreatic cancer cells after treatment with gemcitabine. Additionally, ETAR expression was significantly higher in tumor tissues than in normal tissues, and patients with high ETAR expression had a notably worse overall survival rate than those with low ETAR expression. Furthermore, our results revealed that bosentan, an ETAR antagonist, enhanced the growth-inhibiting and proapoptotic effects of gemcitabine on pancreatic cancer cells.

Thus, our findings indicate that blockade of the ET-1/ETAR axis signaling pathway promotes the antiproliferative effect of gemcitabine on pancreatic cancer. Therefore, combination of ETAR blockade and gemcitabine serves as an effective therapeutic approach to achieve clinical benefits in pancreatic adenocarcinoma patients.

Introduction

Pancreatic adenocarcinoma is currently the third or fourth most common leading cause of cancer-related death worldwide [1,2]. The 5-year overall survival rate of pancreatic adenocarcinoma is the lowest among all cancer survival rates, and this disease has a very poor prognosis [3,4]. The high rate of mortality in pancreatic cancer patients is due to an inability to detect early-stage disease and the disease being highly refractory to therapy. In particular, the major reason for the poor survival outcome is resistance to chemotherapy [5,6].

Gemcitabine has been the standard chemotherapy for advanced pancreatic cancer patients for the last two decades [7,8]. However, gemcitabine resistance develops within a few weeks of treatment, and the underlying mechanism remains poorly understood [9]. To improve the limitation of gemcitabine chemotherapy, other drugs, such as paclitaxel, have been used in combination with gemcitabine, but side effects also increase with combination [10,11]. Therefore, appropriate elucidation of the molecular mechanisms underlying resistance to gemcitabine is crucial for the development of chemotherapy, and a novel therapeutic strategy is needed to overcome the limited clinical efficacy of gemcitabine in pancreatic adenocarcinoma.

The endothelin family consists of three isoforms, and endothelin-1 (ET-1) is the most abundant isoform in multiple tissues [12,13]. Endothelin ligands act through two subtypes of a G-protein-coupled receptor (GPCR), endothelin receptor A (ETAR) and endothelin receptor B (ETBR) [14]. ETAR is more highly specific for ET-1 than is ETBR [15]. ET-1 and ETAR are well known as key regulators of tumor progression in many human malignancies, including pancreatic adenocarcinoma, colorectal carcinoma, breast cancer and ovarian cancer [[16], [17], [18], [19]]. Recent studies have suggested that autocrine activation of the ET-1/ETAR axis is involved in regulating epithelial-to-mesenchymal transition (EMT), aggressive phenotypes, metastatic potential and drug resistance in human cancers [[20], [21], [22]].

Therefore, to elucidate the mechanism underlying chemoresistance to gemcitabine, we investigated ET-1/ETAR axis gene expression in human pancreatic cancer cells after treatment with gemcitabine and determined whether the ET-1/ETAR axis signaling pathway influences gemcitabine resistance.

Section snippets

Cell lines and culture

The established human pancreatic cancer cell lines AsPC-1 and Panc-1 were purchased from the Korean Cell Line Bank (KCLB; Seoul, Korea). AsPC-1 cells were maintained in RPMI-1640 medium supplemented with 1% penicillin-streptomycin, 1% sodium pyruvate and 10% fetal bovine serum (FBS), and Panc-1 cells were cultured in DMEM (high glucose) supplemented with 1% penicillin-streptomycin and 10% FBS at 37 °C in a humidified atmosphere of 5% CO2 following the manufacturer’s protocol.

RNA extraction and quantitative real-time PCR (qRT-PCR) analysis

Total RNA was

Gemcitabine promoted ET-1/ETAR axis signaling in pancreatic cancer cells

To evaluate the effect of ET-1/ETAR axis gene expression on pancreatic cancer cells after treatment with gemcitabine, we treated pancreatic cancer cells with gemcitabine (AsPC-1: 30 or 120 nM, Panc-1: 25 or 100 nM) for 24 h. ET-1 and ETAR mRNA expression levels were gradually upregulated in a dose-dependent manner by gemcitabine treatment in the pancreatic cancer cell lines (Fig. 1A). However, ETBR mRNA expression levels were not changed. ETAR protein expression levels were significantly

Discussion

Pancreatic adenocarcinoma remains a leading common cause of cancer-related death [1,2]. While gemcitabine is the standard chemotherapy for advanced and metastatic pancreatic cancers, drug resistance develops within a few weeks of treatment initiation [[7], [8], [9]]. Therefore, appropriate elucidation of the molecular mechanisms underlying resistance to gemcitabine is crucial for the development of chemotherapy for pancreatic adenocarcinoma. Recently, numerous studies have been performed to

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This research was supported by the National Research Foundation of Korea (NRF) grants funded by the Korean government (MSIT) (2017M3A9C8060558, 2018R1D1A1B07051154 and 2019R1A2C1086258) and the National Cancer Center grant (NCC-2010250).

References (46)

  • S. Bhargava et al.

    Selective inhibition of endothelin receptor A as an anti-angiogenic and anti-proliferative strategy for human pancreatic cancer

    J. Gastrointest. Surg.

    (2005)
  • S. Bhargava et al.

    Selective inhibition of endothelin receptor A as an anti-angiogenic and anti-proliferative strategy for human pancreatic cancer

    J. Gastrointest. Surg.

    (2005)
  • L. Rahib et al.

    Projecting cancer incidence and deaths to 2030: the unexpected burden of thyroid, liver, and pancreas cancers in the United States

    Cancer Res

    (2014)
  • R.L. Siegel et al.

    Cancer statistics, 2016

    Ca - Cancer J. Clin.

    (2016)
  • I. Garrido-Laguna et al.

    Pancreatic cancer: from state-of-the-art treatments to promising novel therapies

    Nat. Rev. Clin. Oncol.

    (2015)
  • G. Maity et al.

    CYR61/CCN1 regulates dCK and CTGF and causes gemcitabine-resistant phenotype in pancreatic ductal adenocarcinoma

    Mol. Canc. Therapeut.

    (2019)
  • J.E. Ying et al.

    Developments in metastatic pancreatic cancer: is gemcitabine still the standard?

    World J. Gastroenterol.

    (2012)
  • M. Gnanamony et al.

    Chemoresistance in pancreatic cancer: emerging concepts

    Oncol Lett

    (2017)
  • D.D. Von Hoff et al.

    Increased survival in pancreatic cancer with nab-paclitaxel plus gemcitabine

    N. Engl. J. Med.

    (2013)
  • M. Barton et al.

    Endothelin: 20 years from discovery to therapy

    Can. J. Physiol. Pharmacol.

    (2008)
  • Z. Wang et al.

    Endothelin promotes colorectal tumorigenesis by activating YAP/TAZ

    Cancer Res

    (2017)
  • T.F. Lüscher et al.

    Endothelins and endothelin receptor antagonists: therapeutic considerations for a novel class of cardiovascular drugs

    Circulation

    (2000)
  • L. Rosano et al.

    Combined targeting of endothelin A receptor and epidermal growth factor receptor in ovarian cancer shows enhanced antitumor activity

    Cancer Res

    (2007)
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