Elevated glypican-1 expression is associated with an unfavorable prognosis in pancreatic ductal adenocarcinoma

Abstract

Pancreatic ductal adenocarcinoma (PDAC) is the most lethal cancer in humans, with a 5-year survival rate of <5%. Recently, glypican-1 (GPC1)-expressing circulating exosomes were found to be a promising diagnostic tool for PDAC. However, the aberrant expression of GPC1 has not been systematically evaluated in large-scale clinical samples of PDAC. Here, we performed a comprehensive analysis of GPC1 mRNA and protein expression features. Included in this study were 178 PDAC patients from the cancer genome atlas (TCGA) and 186 subjects whose tissues were used in immunohistochemical staining assays. We demonstrated that GPC1 mRNA was silenced in normal pancreata; however, it was re-expressed in PDAC tissues probably because of the promoter hypomethylation. The GPC1 protein was barely expressed in the normal and adjacent noncancerous pancreata. In tumor tissues, 59.7% (111/186) of the detected samples showed positive expression. Notably, GPC1 was elevated in 63.6% (34/55) of early stage cases. High levels of GPC1 were associated with poorer differentiation and larger tumor diameters. Kaplan-Meier analysis showed a significant difference in overall survival between the groups categorized by GPC1 expression (P = 0.0028). Multivariate analyses indicated that GPC1 was a significant risk factor for poor overall survival with a 1.82-fold increase in the hazard ratio (P = 0.0022). In conclusion, during pancreatic tumorigenesis, GPC1 was ectopically expressed and served as an independent poor prognostic factor. Our findings highlighted the alluring prospect of GPC1 as an early diagnostic and prognostic marker as well as a therapeutic target for PDAC.

Keywords: Biomarker; gene expression; glypican-1; pancreatic ductal adenocarcinoma; prognostic factor.

Figure 1

Gene expression of glypican family members in various tissues. (A) GPC1 is the most strongly expressed glypicans in human PDAC tissues (n = 178) according to the TCGA RNA sequencing data. (B) The correlation matrix plot of six glypicans in PDAC tissues. Spearman's rank correlation coefficients were calculated and are shown in each square. (C) The tissue‐specific expression of glypican family members according to the normalized RNA‐sequencing data downloaded from the Human Protein Atlas portal (www.proteinatlas.org). The normal pancreas tissues are highlighted by red arrows.

Figure 2

Clinicopathological characteristics of GPC1 mRNA presented in the TCGA RNA sequencing data from pancreatic adenocarcinoma. (A) The expression of GPC1 mRNA was inversely correlated with DNA methylation levels in PDAC. Based on the TCGA RNA‐sequencing and DNA methylation 27k bead array datasets, 178 pancreatic cancer cases had both sets of data available. Pearson correlation coefficients were calculated between the log2 transformed read per million values and methylation status of GPC1. (B–C) Kaplan–Meier curve of overall survival (B) and recurrence‐free survival (C) according to the GPC1 mRNA levels in PDAC tumor tissues. The cases were divided into two groups: a middle and high mRNA level group (upper 67th percentile) and a low mRNA level group (lower 33rd percentile). The log‐rank test was performed. (D) Clinicopathological characteristic analysis of GPC1 mRNA expression in patients with pancreatic adenocarcinoma. The short red line represents the median value in each group.

Figure 3

Representative immunohistochemistry images of GPC1 expression in normal pancreata (A), chronic pancreatitis (B), adjacent nontumor tissues (C), PDAC tumors (D), metastatic tumors in the liver (E), and metastatic lymph nodes (F). Original magnification: left panel, ×40; right panel, ×200.

Figure 4

GPC1 protein expression was associated with poor prognosis in PDAC. Immunohistochemical staining shows strong (A), weak (B), and negative immunoreactivity to GPC1 in PDAC tumor tissues. Original magnification: left panel, ×40; right panel, ×200. (C) Distribution of GPC1 protein levels determined by immunohistochemistry in the normal pancreata (n = 2), chronic pancreatitis (n = 4), nontumor pancreatic tissues (n = 169), PDAC (n = 186), metastases (n = 4), and metastatic lymph nodes (n = 7). The short red line represents the median value in each group. (D) Kaplan–Meier curve of PDAC patients with negative and positive GPC1 expression. The log‐rank test was performed.