## 4 Discussion
In ESCC, SPP1 was associated with poor prognosis in patients with locally advanced ESCC receiving preoperative chemoradiotherapy [30]. Matrix metalloproteinases (MMPs) are widely regarded as important regulators of the tumor microenvironment [31]. According to the gene structure and substrate specificity of the enzyme, MMPs can be divided into six different subgroups. Collagenases (MMP1, 8, 13, 18) are able to degrade type I, II, and III collagen. Gelatinases (MMP2 and MMP9) cleave denatured type IV collagen. Matrix lysoszymes (MMP3, 7, 10, 11, 26, 27) can effectively degrade type III, IV, V collagen, proteoglycan, glycoprotein, and gelatin, etc. MMP12 is also known as macrophage metalloelastase. MMP14, 15, 16, 17, 24, 25 are closely related to the activation of MMP2. Additionally, other unclassified MMPs (MMP19, 20, 21, 22, 23, 28). In the early stage ESCC tissues, MMP10 is highly expressed and is associated with poor prognosis in patients (Figure S3). MMP10 expression is also closely related to lymph node metastasis, TNM stage, and tumor invasion [32]. MMP13 is a proteolytic enzyme belonging to the endopeptidase family of extracellular matrix degradation and it is characterized by zinc binding motif at its catalytic site [33]. MMP13 is produced by many types of cancer and is a key regulator in the process of human malignant tumor metastasis [34; 35; 36]. It is overexpressed in a variety of malignancies [37]. MMP13 was originally identified from overexpressed breast cancer, and its role in breast tumorigenesis has been reported [38; 39; 40]. TP53 has a high mutation rate in esophageal cancer. Studies have shown that 59-93% of patients with esophageal cancer have a TP53 mutation [41; 42], suggesting that TP53 may be involved in the regulation of MMP expression level in esophageal cancer. Previous studies have shown that Ajuba promotes the migration and invasion of esophageal squamous cell lines KYSE450 and KYSE510 by up-regulating the expression of MMP10 and MMP13 [43]. COL1A1, as a type of group I collagen, is abnormally highly expressed in breast cancer [44], gastric cancer [45], non-small cell lung cancer [46], and is associated with the progression and prognosis of cancer. It is an important protein component in the adhesion of the tumor cell and extracellular matrix [47]. In ESCC, [PERSON] et al. found that COL1A1 enhanced cell proliferation, migration, and invasion [48]. Combined with PPI network analysis, survival analysis, and the ROC curve, the results showed that SPP1, MMP13, MMP10, and COL1A1 were four reliable predictors of diagnosis and prognosis of esophageal cancer. Meanwhile, we analyzed the MMP13, MMP10, SPP1, and COL1A1 expression profile in numerous human solid tumors via Oncomine and TIMER2 databases. The results demonstrated that MMP13, MMP10, SPP1, and COL1A1 gene expression were higher in breast cancer, colorectal cancer, head and neck cancer, liver cancer, stomach cancer, and others, than in their matched adjacent normal tissues. This opens up possibilities for follow-up pan-cancer analysis. However, there are some limitations in our study. In public databases, datasets on ESCC are relatively scarce. More relevant clinical samples should be collected for ESCC research. The reliability of the analyses of molecular mechanism and immune infiltration levels, especially the correlation between immune signatures and these key genes, is not supported by experiments in vivo and in vitro. These key genes need to be verified in a larger cohort of ESCC. Some studies have pointed out that there is a correlation between ESCC and Human papillomavirus (HPV) [49; 50]. However, others hold the opposite view [51]. A study suggested that the overexpression of p16 could be used to predict HPV infection [52]. We analyzed the correlation between p16 expression and the expression of MMP10, MMP13, SPP1, and COL1A1 in ESCC samples through the GEPIA2 database and showed that these genes are not correlated (Table S6). This result may be related to the study sample size.