Clinical Value of COL11A1 Expression in Colorectal Cancer
To examine the potential relationship between COL11A1 expression and CRC prognosis, we compared the expression of COL11A1 by analyzed two databases from Oncomine and TCGA, and found that COL11A1 mRNA was highly expressed in colorectal cancer tissues as compared to the normal tissues (Figure 1A-B). We further quantified the expression level of COL11A1 in 118 CRC tissues from patients with matched normal tissues. In agreement with our data in Figure 1A-B, the expression of COL11A1 mRNA was higher in cancer tissues than that in normal tissues (Figure 1C). This suggests COL11A1 correlates with CRC occurrence.
We next evaluated the relationship between COL11A1 expression and different clinicopathological features by using UNCLA database, expression of COL11A1 was positively associated with tumor stage (Figure 1D), node metastasis status (Figure 1E) and histological subtypes (Figure 1F). A high expression of COL11A1 was strongly related with overall survival (OS) (Figure 1G) and disease free survival (DFS) (Figure 1H). To further validate if COL11A1 could affect CRC patients’ survival, we re-analyzed the clinical data from 118 CRC patients. The five-year OS was significantly shorter in the COL11A1-high group than the COL11A1-low group (Figure 1I). In addition, univariate and multiple Cox regression analyses in Figure 1J-K revealed that COL11A1 expression was an independent risk predictor for CRC patients’ survival. Hence, our data indicate that COL11A1 is clinically associated with CRC patient outcome.
Genomic Alteration of COL11A1 in CRC
Genomic alteration is one of important factor for the occurrence of CRC. To determine the frequency and type of COL11A1 alteration in CRC, we used TCGA data with 562 CRC cases from cBioPortal database to examine the genetic alteration. COL11A1 was altered in 54 (10%) of 526 patients with CRC (Figure 2A) and genetic mutation was the most common form (Figure 2B). In addition, we next wanted to study the biological interaction network of COL11A1 in CRC. Though cBioPortal analyses, we showed COL11A1-neighboring genes that were altered at frequencies >5% (Figure 2C). The 20 most frequently altered neighbor genes were further used to explore the biological function network. Analysis of significantly enriched gene ontology (GO) terms indicated that these genes encoded proteins mainly localized to extracellular region, plasma membrane and extracellular exosome. These proteins primarily involved in extracellular matrix organization and cell adhesion, while they also served as metal ion binding, structural molecule activity and identical protein binding (Figure 3A-C). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed enrichment in focal adhesion, PI3K-AKT signaling pathways and ECM-receptor interaction (Figure 3D).
COL11A1 Expression Is Correlated With Immune Infiltration Level in Colon Cancer
Tumor-infiltrating lymphocytes are an independent predictor of sentinel lymph node status and survival in cancers. Tumor purity is an important factor that influences the analysis of immune infiltration in clinical tumor samples by genomic approaches, and TIMER and GEPIA have most of the homologous data from TCGA[26; 23]. By analyzing data from TIMER, we found that COL11A1 had a negative correlation with tumor purity in CRC. And COL11A1 expression level also correlated with poorer prognosis and high infiltration in COAD, as the results showed that COL11A1 had positive related with B cell (r=0.025, p=2.07e-10), CD8+ T cell (r=0.213, p=1.54e-5), CD4+ T cell (r=0.345, p=1.13e-12), macrophage (r=0.54, p=5.98e-32), neutrophil (r=0.422, p=9.93e-19) and dendritic cell (r=0.44, p=1.9e-20) (Figure 4). This finding strongly indicates COL11A1 plays a crucial role in immune infiltration in COAD, especially in those of macrophages, neutrophil and dendritic cell.
Correlation Analysis between COL11A1 Expression and Immune Marker Sets
In order to analyze the correlation between COL11A1 and the diverse immune infiltrating cells, we further investigate the relationships between COL11A1 and immune marker sets of various immune cells of COAD in the GEPIA and TIMER databases. We studied the associations between COL11A1 expression and immune marker genes of different immune cells, such as CD8+ T cells, T cells (general), B cells, monocytes, TAMs, M1 and M2 macrophages, neutrophils, NK cells and DCs in COAD, and the different functional T cells included Th1 cells, Th2 cells, Tfh cells, Th17 cells, Tregs and exhausted T cells. The results showed that expression of COL11A1 was significantly correlated with most immune marker sets of various immune cells and different T cells in COAD (Table 1，uploaded in the supplementary material.).
Interestingly, we also found that COL11A1 expression had strong positive correlations with the expression levels of most marker sets of monocytes (CD86, CSF1R), TAMs (CCL2, CD68 and IL10), M2 macrophages (CD163, VSIG4 and MS4A4A) in COAD (p<0.0001) (Figure 5 and Table 2). We also analyzed the correlations though GEPIA analyses, and the results between COL11A1 and markers of monocytes and TAMs are similar to those in TIMER (Table 2, uploaded in the supplementary material). In addition to bioinformatics analysis, we further used clinical data to examine the relationships between COL11A1 expression and the expression levels of marker sets of monocytes (CD86, CSF1R), TAMs (CCL2, CD68 and IL10), M1 (COX2 and IRF5) and M2 macrophages (CD163, VSIG4 and MS4A4A) in COAD, and the results were in agreement with that in TIMER and GEPIA (Figure 6A-D). These findings suggest that COL11A1may regulate macrophage polarization in COAD.
In addition, we also found significant positive correlations between COL11A1 and the marker genes of Treg (FOXP3, CCR8, STAT5B, and TGF-) and T cell exhaustion (PD-1, CTLA4, LAG3, and TIM-3) (Table 1). FOXP3 plays an important role in Treg cells which leads to the suppression of cytotoxic T cells attacking tumor cells. For PD-1, as a crucial immunosuppressive molecule that regulates T cell exhaustion, has a positive association with COL11A1 expression, suggesting that high COL11A1 expression plays an important role in PD-1 mediating T cell exhaustion. Therefore, these results further confirm the findings that COL11A1 is specifically correlated with immune infiltrating cells in COAD which suggests that COL11A1 plays a vital role in immune escape in the colon cancer microenvironment.