The tumorigenesis of colon adenocarcinoma is commonly known as a multi-stage process, which involves a series of genetic changes interacting with the tumor microenvironment (TME)[14, 15]. This study was conducted to analyze mutations in COAD samples from the TCGA dataset and ICGC dataset. We found a high mutation frequency of MUC4 in both cohorts. We were able to show that mutation in MUC4 was significantly correlated with increased TMB and indicated a worse prognosis. Moreover, samples with MUC4 mutations were enriched in the NK cell mediated cytotoxicity signaling pathway. The result of immune cells infiltration demonstrated that MUC4 mutated samples were more infiltrated in memory CD8 T cells, activated NK cells and M1 macrophages, but less infiltrated in resting memory CD4 T cells.
Some researches have revealed the relationship between tumor mutation burden (TMB) and immunotherapy response.MUC4 exhibited a relatively high mutation rate in COAD patients. Mucins are important glycoproteins, falling into two categories: the membrane-bound mucins and the secreted mucins[16, 17]. MUC4 is a useful biomarker of tumor progression[18, 19]. Its biological effects are mainly used in lung cancer, pancreatic cancer, ovarian and oesophageal malignancy [20–24]. MUC4 is expressed in epithelial cells of the digestive tract, as well as some goblet cells[25]. Abnormal MUC4 expression contributes to the progression of esophageal cancer[21] and colon cancer[26, 27]. Besides, an increased expression in MUC4 demonstrates a poor survival outcome, specifically in the early stages of colon cancer [28]. Due to the size of the MUC4 gene is relatively large, this may increase its mutation probability. Thus, MUC4 is the most common mutated gene in the state of stress exposure, for example, aging or nicotine treatment[29, 30]. MUC4 mutation was significant in Renal Carcinoma and was related to survival outcomes. The mutation of MUC4 could be tightly linked with smoking in kidney cancer [13] [31]. In present study, we revealed that COAD samples with a high rate of MUC4 mutation had a significantly worse survival outcome and were highly correlated with increased TMB. TMB has attracted attention as a biomarker for predicting effectiveness of immune checkpoint inhibitors, higher TMB may respond favorably to anti-PD-1/PD-L1 immunotherapy[32, 33]. However, recent studies [33, 34] confirmed that high TMB is a poor prognostic factor. The possible reasons are that high TMB tends to be associated with chromosomal instability that could depressed immune activities[35].
We used GSEA analysis to explore the potential biological function of MUC4 mutation in COAD. The result showed that the NK cell-mediated cytotoxicity pathway was the main driving mechanism for MUC4 mutation influencing the outcome for COAD patients. Together these functional analyses suggested that the MUC4 mutation had an important function in regulating the immune response in COAD.
Previous studies have indicated that the tumor immune microenvironment(TIM)has a significant meaning in predicting prognosis and evaluating therapeutic efficacy[8]. In our study, we found that that MUC4 mutation was significantly related to increased CD8+ T cells, NK cells and M1 macrophages. We hypothesized that a relatively higher proportion of CD8+ T cells in samples with MUC4 mutated was due to an increased number of tumor neoantigens, which strongly activated immune response. Interestingly, the correlation analysis result revealed that CD8+T cells had a significant negative relation with resting memory CD4+ T cells. We also discovered a higher level of activated NK cells infiltrated in samples with MUC4 mutated, indicating that MUC4 mutation could stimulate activation of NK cells,however, more data are needed for various kinds of tumors. Studies show that NK cells were major players in the early detection, control of tumor metastasis in colon cancer[36–38]. Previous studies reported that M2-tumor-associated macrophages (M2-TAMs) were common in malignant tumors, which creates a favorable microenvironment for tumor progression[39, 40]. Meanwhile, M2-TAMs were found to be linked with poor survival outcome, while M1-TAMs point to a contrary conclusion [41–43]. M1 macrophages could be transformed into M2 with tumor progression. Interestingly, our study found that MUC4 mutated samples were more infiltrated in M1 macrophages. This is probably caused by an early staging of the majority cases in the TCGA datasets. Therefore, further studies are required to verify this hypothesis.