Identification of Cysteine Protease Inhibitor CST2 as a Potential Biomarker for Colorectal Cancer

Abstract

Additional biomarkers for the development and progression of colorectal cancer (CRC) remain to be identified. Hence, the current study aimed to identify potential diagnostic markers for CRC. Analyses of cysteine protease inhibitor [cystatins (CSTs)] expression in CRC samples and its correlation with cancer stage or survival in patients with CRC demonstrated that CRC tissues had greater CST1 and CST2 mRNA expression compared to noncancerous adjacent tissues, while higher CST2 mRNA expression in CRC tissues was correlated with advanced stages and disease-free survival in patients with CRC, encouraging further exploration on the role of CST2 in CRC. Through an online database search and tissue microarray (TMA), we confirmed that CRC samples had higher CST2 expression compared to noncancerous adjacent tissue or normal colorectal tissues at both the mRNA and protein levels. TMA also revealed that colorectal adenoma, CRC, and metastatic CRC tissues exhibited a significantly increased CST2 protein expression. Accordingly, survival analysis demonstrated that the increase in CST2 protein expression was correlated with shorter overall survival of patients with CRC. Moreover, our results found a significant upregulation of CST2 in multiple cancer tissues. Taken together, these findings suggest the potential role of CST2 as a diagnostic and prognostic biomarker for CRC.

Keywords: Biomarker; CST2; Colorectal cancer; Oncogene; Tissue microarray.

Figure 1

Upregulation of CST1 and CST2 at the mRNA level in colorectal cancer tissues. CST mRNA expression was analyzed using the (A) Colon adenocarcinoma (COAD) and (B) Rectum adenocarcinoma (READ) datasets from the GEPIA database. *P < 0.05, tumor vs. normal tissues. T, tumor tissues; N, normal tissues.

Figure 2

Higher expression of CST2 in advanced stage colorectal cancer. CST mRNA expression at different colorectal cancer stages was compared based on the GEPIA database.

Figure 3

High CST2 mRNA expression was correlated with shorter disease-free survival in patients with colorectal cancer. The correlation between overall survival (A) and disease-free survival (B) in patients with CRC and CST mRNA expression in CRC tissues was analyzed through the GEPIA database. Survival was analyzed using the log-rank test.

Figure 4

CST2 upregulation at the protein level in colorectal cancer tissues. Microscopic observation (A) and scatter chart (B) of CST2 protein in 71 pairs of colorectal cancer tissues and noncancerous adjacent tissues, which was determined by immunohistochemistry-based tissue microarray. Image magnification at ×40 or ×200. *P < 0.05, tumor vs. normal tissues. Microscopic observation (C) and scatter chart (D) of CST2 protein in 7 pairs of CRC tissues, normal, and adjacent tissues determined using immunohistochemistry-based tissue microarray. Image magnification at ×40 or ×200. *P < 0.05, tumor vs. normal tissues, tumor vs. adjacent.

Figure 5

CST2 upregulation at the protein level in both colorectal adenoma and carcinoma tissues. Microscopic observation (A) and scatter chart (B) of CST2 protein in normal, adenoma, primary and metastatic colorectal tissues determined using immunohistochemistry-based tissue microarray. Image magnification at ×40 or ×200. *P < 0.05, colorectal cancer vs. normal tissues.

Figure 6

High CST2 protein expression was correlated with shorter overall survival in patients with colorectal cancer. (A) Representative image of high- and low-CST2 expression. Magnification at ×40 or ×200. (B) Correlation between CST2 protein (immunohistochemistry-based tissue microarray) expression and overall survival was analyzed using Kaplan-Meier plots in 94 patients with colorectal cancer (P < 0.05).

Figure 7

CST2 was highly expressed in multiple cancer tissues. CST2 expression in multiple cancer tissues and normal tissues was compared based on the STAD database. *P < 0.05, **P < 0.005, ***P < 0.001, tumor vs. normal tissues.