Exosomal transfer of p-STAT3 promotes acquired 5-FU resistance in colorectal cancer cells

Qian Zhang,Ka-Wo Chan,Huanliang Liu,Xiangling Yang,Huiliu Tan,Rui-Xian Liu,Lili Wei,Jingdan Zhang,Jiancong Hu

doi:10.1186/s13046-019-1314-9

Abstract

BackgroundAcquired resistance remains a limitation of the clinical use of 5-fluorouracil (5-FU). Because exosomes, are important vesicles participating in intercellular communication, their contribution to the development of acquired 5-FU resistance needs to be elucidated. In this study, we aimed to examine the underlying mechanisms of exosomes from 5-FU resistant cells (RKO/R) in sustaining acquired 5-FU resistance in sensitive cells (RKO/P).MethodsExosomes from a 5-FU-resistant cell line (RKO/R) and its parental cell line RKO/P were isolated and co-cultured with 5-FU-sensitive cells. Real-time cellular analysis (RTCA) and FACS analysis were used to examine cell viability and apoptosis. Exosomal protein profiling was performed using shotgun proteomics. Inhibitors and siRNAs were applied to study the involvement of selected proteins in 5-FU resistance. The effect of exosomal p-STAT3 (Tyr705) on the caspase cascade was examined by western blotting (WB) and high content analysis. Xenograft models were established to determine whether exosomal p-STAT3 can induce 5-FU resistance in vivo.ResultsOur results indicated that exosomes from RKO/R cells significantly promoted cell survival during 5-FU treatment. Proteomics and WB analysis results indicated that GSTP1 and p-STAT3 (Tyr705) were enriched in exosomes from RKO/R cells. Inhibition of p-STAT3 re-sensitized RKO/P cells to 5-FU via caspase cascade. Furthermore, p-STAT3 packaged by exosomes from RKO/R cells increased resistance of tumor cells to 5-FU in vivo.ConclusionsOur results reveal a novel mechanism by which p-STAT3-containing exosomes contribute to acquired 5-FU resistance in CRC. This study suggests a new option for potentiating the 5-FU response and finding biomarkers for chemotherapy resistance.

Highlights

Acquired resistance remains a limitation of the clinical use of 5-fluorouracil (5-FU)
To determine the capability of exosomes isolated from 5-FU resistant cell line (RKO/R) cells to transmit acquired 5-FU resistance to recipient cells, we first cultured recipient cells (RKO/P and HCT116) in medium containing 5-FU plus exosomes from RKO parental cell line (RKO/P) (Exo/P) or Exo/ R
The results showed that neither the morphology nor the survival rate of cells treated with exosomes from 5-FU resistant cells RKO/R (Exo/R) and a GSTP1 inhibitor changed significantly compared to those of cells treated with Exo/R and DMSO (Fig. 5a, b, c), excluding the possibility that 5-FU resistance is mediated by GSTP1 in exosomes

Summary

Introduction

Acquired resistance remains a limitation of the clinical use of 5-fluorouracil (5-FU). Most patients acquire resistance during treatment with a median survival of approximately 20 months [5, 6]. 5-FU-based chemotherapy has become the major hurdle to improve treatment efficacy. Some studies have demonstrated that cancer cells are resistant to chemotherapy intrinsically. Mutant p53 tumor suppressor gene is found in many human tumors and its activity is associated with 5-FU resistance [7]. Some members of Bcl-2 family, such as Bcl-xL and Bax, are related to 5-FU resistance in colorectal cancer [8]. Overexpression of Astrocyte elevated gene-1 (AEG-1) increases 5-FU resistance in human hepatocellular carcinoma (HCC) [9]. While other studies have indicated that drug resistance could be acquired. Tumor microenvironment includes many stromal cells and plays a critical role in

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