ERAP2 is a novel target involved in autophagy and activation of pancreatic stellate cells via UPR signaling pathway

Abstract

Background/objectives

Pancreatic ductal adenocarcinoma (PDAC) is characterized by excessive desmoplasia and autophagy-dependent tumorigenic growth. Pancreatic stellate cells (PSCs) as a predominant stromal cell type play a critical role in PDAC biology. We have previously reported that autophagy facilitates PSC activation, however, the mechanism remains unknown. We investigated the mechanism of autophagy in PSC activation.

Methods

We compared gene expression profiles between patient-derived PSCs from pancreatic cancer and chronic pancreatitis using a microarray. The stromal expression of target gene in specimen of PDAC patients (n = 63) was analyzed. The effect of target gene on autophagy and activation of PSCs was investigated by small interfering RNAs transfection, and the relationship between autophagy and ER stress was investigated. We analyzed the growth and fibrosis of xenografted tumor by orthotopic models.

Results

In analysis of gene expression microarray, endoplasmic reticulum aminopeptidase 2 (ERAP2) upregulated in cancer-associated PSCs was identified as the target gene. High stromal ERAP2 expression is associated with a poor prognosis of PDAC patients. Knockdown of ERAP2 inhibited unfolded protein response mediated autophagy, and led to inactivation of PSCs, thereby attenuating tumor-stromal interactions by inhibiting production of IL-6 and fibronectin. In vivo, the promoting effect of PSCs on xenografted tumor growth and fibrosis was inhibited by ERAP2 knockdown.

Conclusions

Our findings demonstrate a novel mechanism of PSCs activation regulated by autophagy. ERAP2 as a promising therapeutic target may provide a novel strategy for the treatment of PDAC.

Introduction

Pancreatic cancer is an aggressive malignant tumor with a 5-year survival rate of 9% [1]. It is projected to become the second leading cause of cancer-related death by 2030 [2]. Surgery is the only potentially curative option and chemotherapy can extend overall survival [3,4]. Because the current treatments of pancreatic cancer are insufficient to improve the survival rate significantly, it is urgent to find a viable strategy to improve therapeutic effectiveness.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by excessive desmoplasia, in which pancreatic stellate cells (PSCs) as the predominant stromal cell type play a major role through tumor-stromal interactions [5]. When exposed to various stimuli in the tumor microenvironment, PSCs transdifferentiate from a quiescent to activated state represented by losing vitamin A-containing lipid droplets in the cytoplasm, undergoing morphological and functional changes, becoming “myofibroblast-like” cells, and producing large amounts of extracellular matrix (ECM) components, cytokines, and chemokines to facilitate pancreatic cancer cell (PCC) aggressiveness and therapeutic resistance [6].

We have previously shown that macroautophagy (hereafter referred to as autophagy) is associated with PSC activation [7]. Autophagy is a dynamic process involved in energy and nutrient homeostasis [8], and a broad spectrum of human diseases [9,10]. The roles of autophagy in cancer are contradictory in different cancer contexts [11,12]. In pancreatic cancer, production and secretion of activated PSCs is critical for cancer cell metabolism and aggressiveness, which is also regulated by autophagy [7,13]. Therefore, stroma-targeting therapy is a potentially promising strategy for pancreatic cancer [14,15]. However, the mechanism of PSC activation remains to be further elucidated [16]. Therefore, autophagy may fully or partially explain the mechanism of PSC activation.

Endoplasmic reticulum (ER) is an organelle where native folding and initial post-translational modifications occur [17]. Under conditions such as hypoxia, nutrient deprivation, and infection, the folding capacity of ER is exceeded, which is referred to as ER stress, resulting in accumulation of unfolded proteins in the ER. Correspondingly, the unfolded protein response (UPR) activates to restore ER protein-folding homeostasis [18]. Autophagy is initiated at the ER membrane and induced via the UPR pathway to relieve the burden of accumulating aberrant proteins during ER stress [[19], [20], [21]]. Furthermore, ER stress is implicated in activation of hepatic stellate cells (HSCs) in liver fibrosis [22]. Therefore, we hypothesized that ER activity is the crosstalk between autophagy and activation of PSCs.

Here, we applied a gene expression microarray and demonstrated that ER-resident target endoplasmic reticulum aminopeptidase 2 (ERAP2) is highly expressed in activated PSCs. We found that ERAP2 plays an important role in autophagy of PSCs, which regulates activation of PSCs through ER-derived autophagy. This process is mediated by ER stress and consequent UPR signaling pathways. Therefore, ERAP2 as a promising therapeutic target may provide a novel strategy for the treatment of PDAC.