Dasatinib enhances cisplatin sensitivity in human esophageal squamous cell carcinoma (ESCC) cells via suppression of PI3K/AKT and Stat3 pathways

https://doi.org/10.1016/j.abb.2014.11.008Get rights and content

Highlights

  • Inhibition of Src by dasatinib sensitizes ESCC cells to cisplatin.

  • Dasatinib suppresses the PI3K/AKT and Stat3 pathways.

  • Dasatinib regulates the cisplatin-resistant molecules, such as ERCC1 and BRCA1.

Abstract

The clinical efficacy of cisplatin in esophageal squamous cell carcinoma (ESCC) treatment remains undesirable. Src, a non-receptor tyrosine kinase involved in multiple fields of tumorigenesis, recently has been indicated as a promising therapeutic target in the treatment of solid tumors including ESCC. However, whether inhibition of Src activity can increase cisplatin efficacy in ESCC cells remains unknown. The present study found that inhibition of Src by its inhibitor-dasatinib sensitized ESCC cells to cisplatin in vitro. Our data also suggest a likely mechanism for this synergy that dasatinib reduces expression of critical oncogenic members of the signaling pathways, such as AKT or Stat3, and cisplatin-resistant molecules, such as ERCC1 and BRCA1, under the control of Src. Furthermore, dasatinib could sensitize ESCC cells to another platin-based agent, carboplatin. Therefore, this study provides a potential target for improving cisplatin efficacy in ESCC therapy.

Introduction

Esophageal squamous cell carcinoma (ESCC)1 is one of the most common malignancies worldwide and occurs at a relatively high frequency in China [1]. Cisplatin is the most frequently used chemotherapeutic agent for ESCC treatment. Unfortunately, the clinical efficacy of cisplatin in ESCC treatment remains undesirable.

Src is a non-receptor tyrosine kinase involved in multiple fields of tumorigenesis including proliferation, migration, and angiogenesis, thus recently indicated as a promising therapeutic target in the treatment of solid tumors including ESCC [2], [3], [4], [5]. Molecular mechanistic studies reveal that hyperactivation of Src is central mediator in multiple signaling pathways of tumors, such as interacting with tyrosine kinase receptors and subsequently activating the Raf/MEK/MAPK pathway and PI3K/AKT pathway, or regulating pro-tumorigenic gene expression via transcription factors, such as Stat3 [6], [7], [8], [9]. Dasatinib, currently entering clinical trials for various malignancies to inhibit the Src activity, increases cisplatin cytotoxicity in non-small cell lung cancer (NSCLC) or breast cancer cells in vitro [10], [11]. However, there are still lacks of direct evidences that dasatinib enhances the cisplatin efficacy in ESCC treatment.

In this study, we sought to explore the potential correlation between inhibition of Src activation and cisplatin efficacy in ESCC cells. The results demonstrate a striking synergistic cytotoxic effect between Src inhibitor-dasatinib and cisplatin in ESCC cells. Our data also suggest a likely mechanism for this synergy that dasatinib reduces expression of critical members of the growth-promoting and apoptosis-resistant molecules under the control of Src. This study provides a potential target for improving cisplatin efficacy in ESCC therapy.

Section snippets

Antibodies and reagents

The sources of the antibodies were: anti-pAKT (Ser473), anti-pStat3 (Tyr705), anti-pERK1/2 (Thr202/Tyr204), anti-pSrc (Tyr416), anti-Src, anti-AKT, anti-Stat3, anti-MMP-9, and anti-GAPDH (Cell Signaling Technology). Antibodies against c-myc and VEGF were from Santa Cruz and Abcam, respectively. Dasatinib, LY294002 (PI3K/AKT inhibitor), U0126 (MEK inhibitor), cisplatin, carboplatin, etoposide, and SN38 were purchased from Selleck chemicals. Amrubicin was from Santa Cruz. S3I-201(Stat3 inhibitor)

Inhibition of Src activity sensitizes ESCC cells to cisplatin

We first evaluated the levels of phospho/total Src in ESCC cells and normal esophageal epithelial cells (NEEC). As shown in Fig. 1A, the levels of phospho/total Src was markedly higher in ESCC cell lines than that in NEEC. To explore the anti-proliferative effect of Src inhibitor-dasatinib in ESCC cells, we performed the MTS assay in human ESCC cell lines- KYSE410 and KYSE150. Dose–response curve in these cells showed that IC50 value of dasatinib in KYSE410 or KYSE150 cells was 91 or 115 nM,

Discussion

This study investigates the influence of inhibition of Src activity by dasatinib on cisplatin sensitivity in ESCC cells. The results indicate that dasatinib dramatically increases the growth-inhibitory effect of cisplatin in ESCC cell lines. Notably, we observe that the underlying mechanism of dasatinib-enhanced cisplatin cytotoxicity mainly via suppressing PI3K/AKT and Stat3 pathways, as well as inhibiting cisplatin-induced mRNA transcription of DNA repair and synthesis genes.

Our study shows

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Acknowledgments

This work is supported by the National Natural Fund of China (81021061, 81230047) and China Postdoctoral Science Foundation (2013M530555).

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