Identification Of Super-enhancer-related Genes In Hepatobiliary Malignant Tumors And Related Targeted Intervention Research

Gallbladder cancer(GBC)is a primary malignant tumor of the epithelium of the gallbladder.Most patients are already in the advanced stage of the disease when they are diagnosed,losing the chance of radical surgery.GBC has a very high degree of malignancy and is not sensitive to conventional chemotherapy.Currently,as to GBC,the available treatment options are limited,making the prognosis remaining rather poor.Therefore,in order to find effective therapeutic targets,it is of great significance to study the mechanism of gallbladder carcinogenesis.Super-enhancers are enhancers with strong transcriptional activity across the genome.By enriching a large number of transcription factors and cofactors,they are mainly responsible for regulating the expression of genes that determine cell identity,including key genes related to cell differentiation and critical oncogenes of tumor cells.It was found that tumor cells were extremely sensitive to the targeted inhibition of the super-enhancer machinery,which could be attributed to the dependence of tumor cells on the key oncogenes regulated by the super-enhancer.The study of super-enhancers in GBC is still blank.Depicting super-enhancer-associated molecular mechanism in the onset and progression of gallbladder cancer may contribute to the discovery of potential therapeutic targets and brings hope for the improvement of prognosis of patients with GBC.In this study,the super-enhancer landscapes of two gallbladder cancer cell lines and two normal gallbladder tissues were first described by CHIP-seq assay against H3K27ac antibody.By comparing the super-enhancer landscapes of gallbladder carcinoma cells and normal gallbladder tissues,we identified 1521 GBC-SE genes.Functional enrichment analyses showed that GBC-SE genes were significantly enriched in tumor-related biological processes such as cell adhesion and tumor-related signaling pathways such as AMPK signaling pathway,suggesting that they may play an important role in gallbladder carcinogenesis.Super-enhancer machinery plays an important role in building and maintaining the function of super-enhancers,and our study further found that gallbladder cancer cells were rather sensitive to the CDK7 inhibitor THZ1.The IC50 of THZ1 in NOZ and GBC-SD cell was 26.8nM and 277.9nM,respectively.Both in vitro and in vivo experiments confirmed that THZ1 could significantly inhibit the progression of gallbladder cancer.Further experiments confirmed that THZ1 treatment effectively suppressed the phosphorylation of the Ser-2,Ser-5 and Ser-7 residues of the RNA Pol Ⅱ C-terminal domain(CTD)in GBC cells,and CDK7 knockdown could also inhibit GBC cell proliferation.Finally,we integrated CHIP-seq and RNA-seq data to analyze the key oncogenes associated with super-enhancers in gallbladder cancer.The results showed that 220 and 26 candidate super-enhancer associated oncogenes were identified in NOZ and GBC-SD cells,respectively.MTFP1 was then verified to be inhibited by THZ1 in GBC cancer cells,and knockdown of MTFP1 indeed inhibited GBC cell proliferation.Hepatocellular carcinoma(HCC)is one of the most common and lethal cancers worldwide.Most patients are diagnosed at an advanced stage,at which point the radical surgical treatments are not available.For advanced HCC,patients have limited options for treatment.Therefore,a better understanding of the detailed molecular mechanism underlying HCC tumorigenesis is urgently needed.In this study,by analyzing CHIP-seq data of HCC cells and combining with the transcriptome data of HCC from TCGA database,a number of candidate oncogenes related to super-enhancers in HCC were identified.GO analyses indicated that the tumor-related biological processes such as cell migration,epithelial-mesenchymal transformation(EMT)and cell differentiation were significantly enriched in the candidate oncogenes related to super-enhancers,indicating that they played an significant role in liver carcinogenesis.Then,we performed prognostic analyses using the TCGA dataset,and found that 42 candidate oncogenes were associated with poor prognosis in patients with HCC,consistent with their role as oncogenes.By hierarchical clustering,we found that the above 42 prognostic super-enhancer related candidate oncogenes could divide HCC patients into two subgroups(cluster 1 and cluster 2),among which the prognosis of patients in the cluster 2 subgroup was worse than those in the cluster 1 subgroup.Finally,we focused on the SLC family gene SLC7A6 to further investigate its expression in HCC and its clinical significance.The results showed that the expression of SLC7A6 in HCC was significantly increased,and its high expression was positively correlated with later clinical TNM stage,worse clinical grade and higher AFP level.In terms of prognosis,we found that high expression of SLC7A6 was associated with poor prognosis in patients with HCC.Furthermore,multivariate cox regression analysis suggested that high expression of SLC7A6 served as an independent predictor of poor prognosis in HCC patients.