EC is one of the most lethal malignancies of the digestive tract worldwide and has high morbidity and mortality rates. Traditional methods are unsatisfactory. Hence, clinicians and physicians are paying more attention to molecular biomarkers for EC diagnosis and prognosis. There are two main histological subtypes of EC: EAC and ESCC. Studies at the molecular level are usually based on one of these two subtypes of EC. In addition, diagnostic and prognostic studies are usually separated. In brief, the identified biomarkers are not broadly applicable. However, no evidence has shown that different molecular therapy methods should be developed that apply to different histological types of EC.
We aimed to identify the risk genes among PCGs that can be applied in both diagnosis and OS prediction. The feasibility of our aim was explored in advance via network analysis. After confirmation of the applicability of the design, a comprehensive study was generated to identify biomarkers via weighted coexpression network analysis, GO enrichment analysis, and LASSO Cox regression analysis. The reproducibility of the network and preservation of the modules were also validated in the analysis. These procedures are crucial, as they are prerequisites to ensure that the results are repeatable. Finally, CMA1 was identified as a biomarker, and it was widely explored and comprehensively compared with other biomarkers identified in other studies of EC. The results show that CMA1 is a promising candidate among these biomarkers due to its superior performance in both diagnostic and prognostic roles.
Downregulation of CMA1 expression was observed in various cancers, and the difference was relatively high between tumor and nontumor tissues in the EC dataset (Fig. 8A). Tumor infiltrating lymphocytes are prognostic biomarkers in various cancers.[27, 28] TIMER analysis revealed a correlation between the infiltration of many types of immune cells, especially dendritic cells, and the expression of CMA1, which indicated the possibility that CMA1 influences tumor progression through immune infiltration.
Mast cells are recognized as a crucial part of the immune microenvironment in tumor tissues and modulate tumor progression by releasing protumorigenic and antitumorigenic molecules,[29] including transforming growth factor-beta, tumor necrosis factor-alpha, interleukin-8, FGF-2, and VEGF.[30] Increased activity of chymase and tryptase secreted by mast cells was observed at all stages of tumor progression and was accompanied by an increase in the number and size of blood vessels.[31]
In a breast cancer immunotyping model based on tumor-infiltrating immune cell subsets published in 2020,[32] patients with high B cell, NK cell, CD8 + cell, activated CD4 + memory T cell levels and low γδT cell and activated mast cell levels had a better prognosis than those with other infiltration patterns, so activated mast cells are unfavorable factors in terms of breast cancer patient survival. Moreover, an in vitro animal experiment showed that coinjection of mast cells and HER2-positive breast cancer cells increased tumor engraftment and outgrowth in mice.[33] Additionally, a high level of mast cells in extratumoral benign prostate tissues was considered a risk factor for a poor prognosis in prostate cancer patients.[34] Mechanistically, it was observed that tumor-derived microvesicles from non-small-cell lung cancer (NSCLC) cells are internalized by mast cells, which enhances the migratory ability of mast cells and promotes the release of TNF-alpha and MCP-1.[35] In EC, however, there are some conflicting reports about the effect of mast cell infiltration on prognosis. Tinge et al found that the number of mast cells was not related to prognosis in EC patients,[36] but Fakhrjou et al pointed out that high mast cell density in the invasive edge of the tumor correlated with tumor progression and poorer survival.[37] Further studies are required to confirm the prognostic significance of mast cell infiltration in malignancies due to the complicated functions of mast cells.
Located at 14q11.2 in a cluster of genes encoding other proteases, CMA1 encodes a chymotryptic serine proteinase belonging to the peptidase family S1.[38] It is secreted by mast cells and has been reported to play an essential role in the degradation of the extracellular matrix, the regulation of submucosal gland secretion, and the generation of vasoactive peptides. In the heart and blood vessels, the major convertor from angiotensin I to the vasoactive peptide angiotensin II is not angiotensin converting enzyme but CMA1, so CMA1 aroused the interest of experts in heart[39] and hypertension research.[40]
In the field of oncology, however, CMA1 is not well studied. There are only a few studies concerning CMA1 in oncology. It was reported in 2015 that chymase has angiogenic effects and can induce tumor growth.[41] Interestingly, miR-9 induced an elevation of CMA1 expression in both P815 (a mouse malignant mast cell line expressing activating KIT mutations) and mouse bone marrow-derived mast cells, supporting that miR-9 promotes metastasis by increasing the expression of proteases crucial for physical remodeling of the extracellular matrix.[42] Chymase-positive mast cells seem to be a biomarker of poor prognosis in breast cancer,[43] gastric cancer,[30] and lung cancer.[44]
In different malignancies, the mRNA levels of CMA1 have various prognostic implications. Elevated CMA1 mRNA expression levels were found in benign prostate hyperplasia tissue compared to prostate cancer tissue or normal prostate tissue.[45] Of note, the CMA1 mRNA levels were lower in oral squamous cell carcinoma tissue than in normal tissue, and patients with high CMA1 expression had apparently better prognoses (p = 7*10− 6).[46] The CMA1 mRNA levels were elevated in the normal mucosa compared to the normal tissues and tumor tissues of colorectal carcinoma patients.[47] The upregulation of THBS2 and SPARC at the same time also implied that inflammatory activation of stromal fibroblasts is not only an integral part of colorectal carcinoma but can facilitate its invasion of the surroundings.[47] Increased CMA1 mRNA was associated with an unfavorable prognosis in gastric cancer but a favorable prognosis in ovarian cancer.[48] Mechanistically, CMA1 may influence the prognosis of cancer patients by altering the immune microenvironment. In our EC research, the CMA1 expression level was positively correlated with the infiltration of dendritic cells, regulatory T cells, and exhausted T cells (Fig. 8B-C). The metastasis promotion potential of dendritic cells is related to inducing Treg cell cytotoxicity and reducing the cytotoxicity of CD8 + T cells. In addition, overexpression of CMA1 was correlated with poorer OS in gastric cancer patients at lymph node stages 1–3 but not in patients without lymph node metastasis. This discovery strongly suggests that the CMA1 expression level is a potential predictive marker of metastasis.[48] However, if CMA1 plays only a driving role in tumor development, it is difficult to explain its controversial prognostic implications across different tumors. Further studies are required to uncover the intricate regulatory network between CMA1 and the immune microenvironment and develop it as a potential immune therapy target.