IGF-1R is frequently overexpressed in primary CRC tumors
The analysis of the IGF-1R gene expression between diverse cancer types and adjacent normal tissues across The Cancer Genome Atlas (TCGA) cohort revealed high gene expression in COAD (n = 458) compared to the adjacent normal tissues (n = 41) (P < 0.05, Fig. 1A) from Sangerbox dataset. Similarly, upregulated IGF-1R gene expression was also observed in primary COAD in GEPIA2 dataset (n = 275, Fig. 1B) and UALCAN portal dataset (n = 286, P < 0.05, Fig. 1C). Furthermore, the IHC results showed significantly higher levels of IGF-1R in tubular adenoma, villous adenoma, and CRC tissues compared to the adjacent normal tissues, as assessed by the Mann-Whitney U test, and the difference between adenoma and CRCs was statistically significant (Fig. 1D, Table 1). These results demonstrated that IGF-1R is commonly overexpressed in CRC and may play a critical role in the growth and malignant transformation of precancerous polyps.
Livin expression is upregulated in CRC
Next, we analyzed the Livin (also known as BIRC7) expression in primary CRC tissues from TCGA cohort. The gene expression was significantly upregulated in primary COAD compared to the adjacent normal tissues in the Sangerbox dataset (P < 0.001, Fig. 2A). Specifically, the upregulation of Livin gene expression in COAD was respectively validated in two datasets (GEPIA2 and UALCAN) of paired tumor and adjacent normal samples from TCGA study (Fig. 2B and 2C). To further analyze Livin expression and validate these findings, we conducted Livin IHC in a cohort of 100 patients with localized CRCs, adenoma, and adjacent normal tissues. The staining showed that the level of Livin protein in the colorectal adenoma and cancer groups was significantly higher than that of the adjacent normal tissues; however, no significant difference was detected between the colorectal adenoma and cancer groups (Fig. 2D, Table 1). These results demonstrated that the high level of Livin protein is related to the occurrence of CRC, and it may not be a crucial factor in the progression of adenoma to CRC.
Overexpression of IGF-1R is an independent predictor of oncogenic function in CRC patients
We further evaluated the clinicopathological significance of the IGF-1R gene and protein expression in CRC patients. A significant correlation was established between IGF-1R gene expression and clinicopathologic features, including age (61-80 years), gender (male), histological subtype (adenocarcinoma), individual cancer stages (stage 3), nodal metastasis status (N1 and N2), and TP53-mutant (Fig. 3A-E and 3G). In addition, the promoter methylation level of IGF-1R was closely related to COAD. We also evaluated the correlation between IGF-1R protein expression and clinicopathological features in colorectal cancer. Consistent with the TCGA study, IGF-1R protein overexpression was associated with the depth of invasion, Dukes’ stage, lymph node metastasis, and distant metastasis in patients with CRC (all P < 0.05, Table 2), and with the increase in Dukes’ stage, the expression level of IGF-1R protein increased significantly (Fig. 3H, Table 2). However, no correlation was established between IGF-1R protein expression and age, gender, diameter, location, histological subtype, and differentiation (Table 2). These results demonstrated that IGF-1R might be used as a biomarker to evaluate the condition of CRC patients.
Clinical application value of Livin expression in CRC
We also analyzed the association between Livin gene and protein expression and the clinicopathological features of CRC. As shown in Fig. 4A-G, a significant correlation was established between Livin gene expression and clinicopathological features, including age (41-60, 61-80, and 81-100 years), gender (male and female), histological subtype (adenocarcinoma), individual cancer stages (1, 2, and 3), nodal metastasis status (N0, N1, and N2), and TP53 mutation status (TP53-mutant and TP53-non-mutant), while no correlation was established in the promoter methylation level of Livin. Moreover, Spearson’s chi-square test analysis showed that Livin protein overexpression was an independent risk factor for stage and metastasis, but was not associated with age, gender, tumor size, primary tumor location, pathological type, histological type, or depth of invasion (Fig. 4H, Table 2).
IGF-1R and Livin are not prognostic markers of CRC
The correlation between IGF-1R and Livin expression and patients’ overall survival (OS) and disease-free survival (DFS) was estimated by Kaplan-Meier analysis. The survival map showed that both IGF-1R and Livin are not significantly correlated with COAD from TCGA dataset (Fig. 5A and 5B). We sorted the patient cases from TCGA-COAD with survival data into the high- and low-expression of IGF-1R and Livin groups. As shown in the Kaplan-Meier survival curves, CRC patients with high IGF-1R or Livin expression showed a longer OS and DFS than those with low expression based on the log-rank test (P > 0.05 for both OS and DFS, Fig. 5C-F). Collectively, these findings indicated that neither IGF-1R nor Livin serve as an independent new biomarker for the prognosis of CRC patients. Moreover, a marked correlation was established between IGF-1R and Livin expression in COAD by Spearman’s chi-squared test analysis (R = 0.2, P = 0.00048, Fig. 6).