Analysis of Interleukin-1 Signaling Alterations of Colon Adenocarcinoma Identified Implications for Immunotherapy

Abstract

Immune checkpoint inhibitors (ICIs) have made breakthrough progress in the treatment of various malignant tumors. However, only some patients receiving ICIs obtain long-lasting clinical effects, and some patients still do not achieve remission. Improving the treatment benefits of this part of the population has become a concern of clinicians. IL-1 signaling plays an important role in the tumor microenvironment (TME). However, the relationship between the IL-1 signaling mutation status and the prognosis of colon adenocarcinoma (COAD) patients receiving ICIs has not been reported. We downloaded the data of a COAD cohort receiving ICIs, including prognostic data and mutation data. Additionally, we downloaded the data of a COAD cohort from The Cancer Genome Atlas (TCGA) database, including clinical data, expression data and mutation data. Gene set enrichment analysis (GSEA) was used to assess differences in the activity of some key physiological pathways between the IL-1 signaling mutated-type (IL-1-MT) and IL-1 signaling wild-type (IL-1-WT) groups. The CIBERSORT algorithm was used to evaluate the contents of immune cells in the TME of COAD patients. The multivariate Cox regression model results suggested that IL-1-MT can be used as an independent predictor of a better prognosis in COAD patients receiving ICIs (P = 0.03, HR = 0.269, 95% CI: 0.082-0.883). Additionally, IL-1-MT COAD patients had significantly longer overall survival (OS) (log-rank P = 0.015). CIBERSORT analysis showed that the IL-1-MT group had high infiltration levels of activated dendritic cells (DCs), M1 macrophages, neutrophils, activated natural killer (NK) cells, activated CD4+ memory T cells and CD8+ T cells. Similarly, the IL-1-MT group had significantly upregulated immunogenicity, including in terms of the tumor mutation burden (TMB), neoantigen load (NAL) and number of mutations in DNA damage repair (DDR) signaling. GSEA showed that the IL-1-MT group was highly enriched in the immune response and proinflammatory mediators. Additionally, the expression levels of immune-related genes, immune checkpoint molecules and immune-related signatures were significantly higher in the IL-1-MT group than in the IL-1-WT group. IL-1-MT may be an independent predictor of a good prognosis in COAD patients receiving ICIs, with significantly longer OS in IL-1-MT COAD patients. Additionally, IL-1-MT was associated with significantly increased immunogenicity, activated immune cell and inflammatory mediator levels and immune response-related scores.

Keywords: IL-1; colon adenocarcinoma; immune checkpoint inhibitors; prognosis; tumor microenvironment.

Figure 1

Predictive values of clinical characteristics and the IL-1 signaling mutation status on ICI outcomes. (A) Forest plot of the results of the univariate Cox and multivariable Cox regression models. In the univariate Cox regression analysis, the factors with a p value below 0.05 were IL-1 signaling MT. Multivariate Cox regression analysis showed that the IL-1 signaling mutation status was an independent predictor of ICI therapy in COAD patients. The main part of the forest plot presents the risk ratio (HR) and 95% confidence interval (95% CI), where red dots indicate P < 0.05. The HR indicates predictors of favorable (HR < 1) or poor (HR > 1) OS. (B) Kaplan-Meier survival curves for the OS of 84 COAD patients in the ICI-treated cohort. We performed KM survival analysis on different subgroups of patients (IL-1 signaling mutation status). (C) Heatmap for the expression of markers related to IL1 signaling. (*P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; Mann-Whitney U test).

Figure 2

Genomic profiles of COAD patients in the ICI-treated (A) and TCGA-COAD (B) cohorts. The top 20 genes with the highest mutation frequencies and the corresponding clinical information are shown in the figure. The top five genes with the highest mutation frequencies in the ICI-treated cohort were APC (72%), KRAS (55%), TP53 (55%), PIK3CA (34%) and KMT2D (31%). The top five genes with the highest mutation frequencies in the TCGA-COAD cohort were APC (74%), TP53 (54%), TTN (53%), KRAS (41%) and MUC16 (30%). For the mutation type, yellow indicates splice site mutations, blue indicates missense mutations, orange indicates frameshift mutations, green indicates inframe indel mutations and brown indicates nonsense mutations. The IL-1 signaling mutation status, sex, age, sample type and MSI score are shown as patient annotations (the upper/lower bar plot). The left bar plot marks the mutation rate of each gene. (*P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001; Fisher’s exact test).

Figure 3

Comparison of clinical characteristics between IL-1-MT and IL-1-WT tumors. (A) Comparison of age between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD cohort. (B) Comparison of the MSI score between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD cohort. (C) Comparison of the sex proportions between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD cohort. (D) Comparison of the sample type proportions between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD cohort. (E) Comparison of age between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (F) Comparison of the sex proportions between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (G) Correlation analysis of the clinical stage proportions between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (*P < 0.05; **P < 0.01; ns, not significant).

Figure 4

IL-1-MT COAD is associated with an inflammatory TME. (A) Comparison of the fractions of 22 immune cells between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (B) Comparison of the immune-related signatures between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (C) Comparison of the expression levels of immune checkpoint molecules between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort. (D) Heatmap depicting the mean differences in immune-related gene mRNA expression between IL-1-MT and IL-1-WT tumors across different cancer types. The x-axis of the heatmap indicates different cancer types, and the y-axis indicates gene names. Each square represents the fold change or difference of each indicated immune-related gene between IL-1-MT and IL-1-WT tumors in each cancer type. Red indicates upregulation, while blue indicates downregulation (*P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; ns, not significant; Mann-Whitney U test).

Figure 5

The results of gene set enrichment analysis. The color of the curve corresponds to the font color of the pathway. GSEA of hallmark gene sets downloaded from the Molecular Signatures Database (MSigDB). Each run was performed with 1000 permutations. Enrichment results with significant differences (P < 0.05) between IL-1-MT and IL-1-WT tumors are shown.

Figure 6

IL-1-MT COAD is associated with enhanced tumor immunogenicity. Comparison of the mutation counts of nine DNA damage-related signaling pathways between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD (A) and TCGA-COAD (B) cohorts. Comparison of the TMB between IL-1-MT and IL-1-WT tumors in the ICI-treated COAD (C) and TCGA-COAD (D) cohorts. Comparison of the NAL between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort (E). Comparison of the MANTIS score between IL-1-MT and IL-1-WT tumors in the TCGA-COAD cohort (F). (*P < 0.05; **P < 0.01; ****P < 0.0001; ns, not significant; Mann-Whitney U test).