Long Non-Coding RNA Signatures Associated with Ferroptosis Predict Prognosis in Colorectal Cancer

Abstract

Background: Currently, colorectal cancer has become a common gastrointestinal malignancy that usually occurs in the colon and rectum, and ferroptosis plays a vital role in the pathology and progression of colorectal tumors.

Methods: A total of 627 patients (51 normal and 644 tumor samples) from The Cancer Genome Atlas (TCGA)-COAD and TCGA-READ were included in the study. Lasso and Cox's regression was employed to analyze the characteristic lncRNAs in colorectal cancer samples, and a distinctive prognostic model of ferroptosis-related lncRNAs was established. By analyzing the divergence between the high and low-risk groups of ferroptosis-related lncRNAs, 15 characteristic lncRNAs related to the prognosis of colorectal cancer were evaluated. Kaplan-Meier analysis, operation characteristic curve (ROC), nomogram, and gene set enrichment analyses (GSEA) further confirmed the validity of the characteristic prognostic model with ferroptosis-related lncRNAs.

Results: Kaplan-Meier analysis confirmed a high-risk group of ferroptosis-related lncRNA interrelated with a poor prognosis in colorectal cancer. AUC estimates of 1 -, 3 -, and 5-year survival rates for ferroptosis-related lncRNA characteristic models were 0.745, 0.767 and 0.789. GSEA analysis showed that immune and malignancy-related pathways were active in the high-risk score group. In addition, differential analyses of immune function, including Checkpoint, cytolytic, HLA, and T cell co-inhibition, differed significantly betwixt low - and high-risk groups.CD160, TNFRSF18, CD27, PDCD1, CD200R1, ADORA2A, TNFRSF14, LAIR1, CD244, CD40, TNFRSF4, CD70, TNFSF14, TNFRSF25, CD276, HHLA2, VTCN1, LAG3, TNFSF18, and other immune checkpoints had different expressions betwixt the high- and low-risk group.

Conclusion: Fifteen kinds of lncRNAs with different expressions (AP003555.1, AC099850.3, AL031985.3, LINC01857, STPG3-AS1, AL137782.1, AC124067.4, AC012313.5, AC083900.1, AC010973.2, ALMS1-IT1, AC013652.1, AC133540.1, AP006621.2, AC018653.3) were closely associated with poor prognosis of colorectal cancer. These indicators were significantly correlated with the overall survival (OS) rate and could be used as prognostic evaluation criteria.

Keywords: TCGA; colorectal cancer; ferroptosis; immune infiltration; lncRNAs.

Figure 1

GO and KEGG analysis of ferroptosis-related differentially expressed genes (DEGs) linked with TCGA-COAD and TCGA-READ data frames for colorectal cancer samples. (A) Perform a GO analysis of these DEGs. (B) KEGG analysis related to these DEGs.

Figure 2

A prognostic characteristic ferroptosis-related lncRNAs’ model assessment based on Colorectal cancer samples from the TCGA-COAD and READ data frame. (A) Survival curves for the two groups with high and low-risk scores. The year is represented by the abscissa, while the survival rate is represented by the ordinate. P<0.05, denotes statistical significance. (B) The AUC value has four different clinical parameters setting (a prognostic characteristic ferroptosis-related lncRNAs, age, gender, clinical stage). The abscissa denotes 1-specificity, while the ordinate represents sensitivity. AUC≈1.0 represents an ideal inspection index; AUC 0.7–0.9 indicates a high-test accuracy value; AUC=0.5 represents that the test has no diagnostic value. (C) The effect of high- and low-risk score groups in the prognostic ferroptosis-related lncRNAs model on survival time and survival status. The abscissa is for the patients to gradually improve their risk score; the high-risk score group is in red, and the low-risk score group is in green. The number of death samples increases the risk score. (D) The AUC values of a characteristic prognostic model of lncRNAs associated with ferroptosis at 1, 3, and 5-years. The ordinate represents sensitivity, and the abscissa represents 1-specificity. AUC≈1.0: An ideal inspection index; AUC 0.7–0.9: has a high-test accuracy value; AUC=0.5: the test has no diagnostic value. (E) DCA of 4 clinical parameter modes (a prognostic characteristic ferroptosis-related lncRNAs, age, gender, clinical stage). The risk threshold probability is represented by abscissa in the figure: in the risk assessment tool, the probability of a patient being diagnosed with colorectal cancer is recorded as Pi; when Pi reaches a certain threshold (denoted as Pt), it is defined as positive, and treatment measures are taken. The decision number model is based on the patient’s benefit after treatment and untreated loss (harm). The ordinate represents the net benefit, which is the patient’s benefit from treatment minus the loss caused by untreated. (F) The heat map shows the high and low-risk score groups related to the expression level of the prognostic ferroptosis-related lncRNAs among colorectal cancer samples. The abscissa represents the high- and low-risk score groups, pink represents the low-risk score group, blue represents the high-risk score group, and the ordinate represents the expression level of characteristic prognostic lncRNAs associated with ferroptosis in the samples; P<0.05, statistically significant.

Figure 3

Clinical correlation heatmap, with red denoting high-risk score group, and blue denoting low-risk score group. The main abscissa represents the high- and low-risk groups are represented by primary abscissa, with blue representing the low-risk score group, and red representing the high-risk score group. In addition, the abscissa stratification includes tumor TNM stage, age, gender, survival status, survival time, and pathological stage; the ordinate represents the expression level of characteristic prognostic lncRNAs associated with ferroptosis in the samples. T: the scope and size of the primary tumor; N: the dissemination of lymph nodes; M: Whether there is metastasis, M0 means that there is no metastasis, and M1 means that there is distant metastasis. Unknown: missing clinical data in the TCGA data frame; stge: stage; fustat: survival status; futime: survival time. *P<0.05, ***P<0.001.

Figure 4

Cox analysis of the ferroptosis-related lncRNA distinctive prognostic model. (A) Univariate Cox analysis, P<0.05, statistically significant. (B) Multivariate Cox analysis. P<0.05, statistically significant. (C) Relationship between characteristic prognostic ferroptosis-related lncRNAs and mRNAs expression.

Figure 5

Based on the total score the nomogram predicts the probability of survival (including the points of clinical factors and the risk score of characteristic ferroptosis-related lncRNAs). ***P<0.001.

Figure 6

GSEA was used to identify the ferroptosis-related lncRNAs. The high-risk group was considerably enriched in the intestinal immune network for IGA production, pathways in cancer, PPAR signaling, NOTCH signaling, STAT signaling, colon cancer signaling, MAPK signaling, and VEGF signaling.

Figure 7

Differences in immune function between high-risk and low-risk populations were analyzed using a heatmap (based on CIBERSORT, ESTIMATE, MCPcounter, ssGSEA, TIMER algorithm). The low-risk group is represented by blue on the horizontal axis; While the high-risk group is represented by red on the horizontal axis. The vertical axis represents the immune cells. Various hues represent various software forecasting approaches.

Figure 8

Immunological function and immune checkpoints in the high-risk and low-risk groups were compared. (A) Immune cell subsets and immune function difference between high-risk and low-risk groups. The immune-related activities in colorectal cancer are shown by the abscissa, and the scores of these functions are represented by the ordinate. *P<0.05, **P<0.01, ***P<0.001. (B) Perform immune checkpoint difference analysis on the high and low-risk scores group. The abscissa represents the immune checkpoint-related genes in colorectal cancer, and the ordinate represents its expression. *P<0.05, **P<0.01, ***P<0.001.

Figure 9

The expression of m6A-related genes in colorectal cancer samples from high-risk and low-risk score groups. The abscissa indicates m6A-related genes in colorectal cancer, and the ordinate reflects the amount of gene expression; *P<0.05, **P<0.01, ***P<0.001.