In this study, a large number of differential genes had been screened, and ceRNA network had been constructed to demonstrate some potential prognostic and therapeutic targets for CCA. The differentially expressed genes of CCA were enriched in the functions of many amino acid kinases, such as the Tyrosine Kinase Inhibitor (TKI), which have been widely used as target therapy for cancers. In addition, in three survival-predictable lncRNA related ceRNA networks, THAP7-AS1 and LINC00519 shared the same mRNAs targets, named ERBB3 and YWHAZ, members of the tyrosine kinase family. These indicated that amino acid inhibitors may become one of the strategies to enhance prognostic in CCA. It is undeniable that the limited sample size is a hindrance in the current ceRNA study.
Functional enrichment analysis of 106 mRNAs, downstream targets of LINC00519 THAP7-AS1 and AC090772.1, suggested they were enriched in the activation of the Toll like receptor signaling pathway that had been reported to modulate the immune microenvironment and promote the metastasis of CCA (23). Among them, RCN2, MECOM and MBNL3 had been reported to be involved in specific pathways as shown in the cancer pathway network map above (Fig. 7). What’s more, recently TWEAK/Fn14 signaling had also been reported to promotes CCA niche formation and progression (24). PI3K/AKT and EGF/EGFR axis had also been reported in many reports to promote the progression of CCA (25,26). Combined with current reports on the mechanism of these pathways in CCA and the mechanism of these three genes participating in these pathways in other specific cancers, we will verify the specific effects of these three genes in these pathways on CCA through cell experiments in the future.
It reported that LINC00519 could be a potential prognostic biomarker and promote the development of lung adenocarcinoma through positively regulated YAP1, which was a pivotal factor in Hippo pathway (27,28) In a recent published report, it pointed out that THAP7-AS1 can be activated by SP1 transcription, by improving the entry of CUL4B protein into the nucleus to inhibit the expression of miR-22-3p and miR-320a, and then activated the PI3K/AKT signaling pathway to promote GC progression (29). Therefore, the mechanism of these three lncRNAs in cholangiocarcinoma remained to be experimentally verified. However, based on published reports, LINC00519 and THAP7-AS1 may become potential drug targets and biomarkers for cholangiocarcinoma, and the four key networks in which they were involved may be able to explain the occurrence and progress of cholangiocarcinoma. As for AC090772.1, there was no relevant report so far, and it had great exploratory value.
AZ628, Crizotinib and paclitaxel were predicted effective drugs for MECOM, RCN2, MBNL3, respectively (Figure S3). Among them, AZ628 is a new type of pan-raf inhibitor, Crizotinib is an ALK tyrosine kinase inhibitor, and paclitaxel acts on the microtubule/tubulin system. Whether these treatments can improve CCA patient survive need further evidence.
Immune infiltration analysis (30) (Figure S4) was performed on six survival-related genes of MECOM, STMN1, RCN2, KIF23, LONRF3 and MBNL3, and in the result of immune infiltration, the expression level of these six genes in CCA may be related to the infiltration degree of a variety of immune cells including NK cells and macrophages. Studies had shown that high macrophages infiltration was associated with poor prognosis, and the activity of NK cells inhibited tumor growth. The increase of NK cells may be a promising method for the treatment of CCA. The high expression of RCN2 in CCA was positively correlated with the degree of infiltration of M0 macrophages and negatively correlated with the appearance of NK cells. Therefore, inhibiting the expression of RCN2 may inhibit the degree of infiltration of M0 macrophages and promote the degree of infiltration of NK cells to improve prognosis. Increased neutrophil infiltration may enhance tumor invasiveness (31). The expression of MBNL3 was positively correlated with the degree of neutrophil infiltration, suggesting that MBNL3 may affect tumor occurrence by reducing neutrophil infiltration. Changes in the expression of these genes may affect the infiltration of immune cells around tumor tissues, which in turn affects their invasion, differentiation and proliferation.
We also verified 6 survival-related mRNAs in the GEO database (Figure S5) and found that MBNL3 and LONRF3 down-regulated genes with p > 0.05. This may be due to the complexity of big data sources and the inconsistency of statistical methods.
We counted the recent research progress of these six prognostic genes in specific cancers. MECOM had been reported to be up-regulated and was related to the aggressive behavior of CCA. It may become a potential therapeutic target and classification basis for CCA (32). It had also been reported that the up-regulation of RCN2 can activate MYC signal and regulate the activation of EGFR-ERK pathway to promote the occurrence of Hepatocellular carcinoma (HCC) (33). What’s more, highly expressed STMN1 had been reported to promote the proliferation of CCA and lead to poor prognosis (34,35). In addition, knockdown of MBNL3 can almost eliminate the occurrence of HCC, and can up-regulate PXN and promote the occurrence of tumors [21]. And Many reports pointed out that KIF23 may become a prognostic predictor and biomarker of HCC (36,37). There was no report on the specific mechanism of MBNL3 and KIF23 in CCA and only one related research which indicated LONRF3 was a causative gene of pancreatic cancer (38). As HCC and CCA were close in anatomical position and similar in high risk factors for carcinogenesis, their pathogenesis may be related. The specific mechanisms of these survival-related genes in the occurrence and development of CCA were not yet clear, and had great exploratory value, and may become predictors and biomarkers for predicting the prognosis of patients with CCA.