Exosomal S100A4 derived from highly metastatic hepatocellular carcinoma cells promotes metastasis by activating STAT3

Haoting Sun,Yan Fu,Kaili Zhang,Mo Chen,Beiyuan Hu,Tiantian Zou,Chaoqun Wang,Qiongzhu Dong,Luyu Yang,Yan Zheng,Lunxiu Qin,Yuanyuan Sheng,Xiaomei Gao,Xudong Ren,Yan Geng,Qin Luo,Shican Yan

doi:10.1038/s41392-021-00579-3

Abstract

Intercellular cross-talk plays important roles in cancer progression and metastasis. Yet how these cancer cells interact with each other is still largely unknown. Exosomes released by tumor cells have been proved to be effective cell-to-cell signal mediators. We explored the functional roles of exosomes in metastasis and the potential prognostic values for hepatocellular carcinoma (HCC). Exosomes were extracted from HCC cells of different metastatic potentials. The metastatic effects of exosomes derived from highly metastatic HCC cells (HMH) were evaluated both in vitro and in vivo. Exosomal proteins were identified with iTRAQ mass spectrum and verified in cell lines, xenograft tumor samples, and functional analyses. Exosomes released by HMH significantly enhanced the in vitro invasion and in vivo metastasis of low metastatic HCC cells (LMH). S100 calcium-binding protein A4 (S100A4) was identified as a functional factor in exosomes derived from HMH. S100A4rich exosomes significantly promoted tumor metastasis both in vitro and in vivo compared with S100A4low exosomes or controls. Moreover, exosomal S100A4 could induce expression of osteopontin (OPN), along with other tumor metastasis/stemness-related genes. Exosomal S100A4 activated OPN transcription via STAT3 phosphorylation. HCC patients with high exosomal S100A4 in plasma also had a poorer prognosis. In conclusion, exosomes from HMH could promote the metastatic potential of LMH, and exosomal S100A4 is a key enhancer for HCC metastasis, activating STAT3 phosphorylation and up-regulating OPN expression. This suggested exosomal S100A4 to be a novel prognostic marker and therapeutic target for HCC metastasis.

Highlights

Metastasis and recurrence, causing about 90% of deaths of cancer patients, are the most significant characteristics of malignant cancers.[1,2] Tumor recurrence occurs in over half of hepatocellular carcinoma (HCC) patients at 5 years after resection and is a common cause of poor prognosis.[3]
We investigated the protein profile of exosomes derived from HCC cells with diverse metastatic potential
In addition to metastatic promotion, in vitro sphere formation rates of MHCC97-L and HepG2 cells were elevated by S100A4rich exosomes treatment (Supplementary Fig. S5a), so was the in vivo tumor initiation ability of MHCC97-L (Supplementary Fig. S5b). These findings further demonstrate the functional roles of exosomal S100 calcium-binding protein A4 (S100A4) in enhancement of the metastatic potential and stemness of HCC cells

Summary

1234567890();,: INTRODUCTION

Metastasis and recurrence, causing about 90% of deaths of cancer patients, are the most significant characteristics of malignant cancers.[1,2] Tumor recurrence occurs in over half of hepatocellular carcinoma (HCC) patients at 5 years after resection and is a common cause of poor prognosis.[3]. In addition to metastatic promotion, in vitro sphere formation rates of MHCC97-L and HepG2 cells were elevated by S100A4rich exosomes treatment (Supplementary Fig. S5a), so was the in vivo tumor initiation ability of MHCC97-L (Supplementary Fig. S5b) Taken together, these findings further demonstrate the functional roles of exosomal S100A4 in enhancement of the metastatic potential and stemness of HCC cells. When OPN was knocked down or the cells were treated by the STAT3 inhibitor S3I-201, the abilities of invasion and migration of HepG2 and PLC cells were significantly inhibited, even after S100A4rich exosome treatment (Supplementary Fig. S6a–d). Similar results were observed with Western blot (Fig. 5f) These results support that S100A4rich exosomes from HMH promote the metastatic potential of LMH, and exosomal S100A4 is a key enhancer for HCC metastasis, by activating STAT3 phosphorylation and upregulating OPN expression (Fig. 6a). The combination of exosomal S100A4 and OPN levels had a better prognostic performance than exosomal S100A4 or OPN alone (Table 2)

DISCUSSION

Findings

MATERIALS AND METHODS