| Epidemiological surveys indicate that liver cancer is the second leading cause of cancer-related death worldwide and is particularly prevalent in China.The annual death of HCC in China accounts for more than half of the world’s total number,it is a major public health problem in China and even in the world.At present,conventional treatments for HCC such as liver resection,liver transplantation and arterial embolization chemotherapy has been mature and routine,but the prognosis improvement also encounters a bottleneck period,and the 5-year overall survival rate remains unsatisfied.The main cause of this bottleneck is the low rate of early diagnosis,the high rate of early recurrence and metastasis after tumor resection,and the lack of other effective extraoperative adjuvant therapy for HCC.Therefore,it is of great clinical and social significance to further explore new biomarkers of the development and early recurrence of HCC,to study its specific mechanism and to explore its therapeutic value.Tumors have their specific genetic background,which is the intrinsic basis of their malignant phenotypes.At present,with the maturity and low price of gene detection and sequencing technology,a large number of cancer gene sequencing data are generated and shared.These datasets have been widely used in the detection of new markers,prediction of outcomes and molecular typing of tumors by means of rational statistical analysis,while similar applications in the study of HCC are still rare.Tumor cells have significant metabolic reprogramming compared with normal cells,and metabolic enzymes are the ultimate performers of this change.The treatment of metabolic enzymes has been applied in some tumors,but there are still few related studies in HCC.Therefore,based on the analysis of large public data of HCC,two main research parts were included in our study.One is to construct a multi-m RNA model that can effectively predict early recurrence of HCC after tumor resection;the other is to systematically screen key metabolic enzymes in HCC development,to further study their roles,mechanisms and therapeutic value in HCC.Both studies will effectively improve the prognosis of patients with HCC,and have certain clinical transformation value.Part I Construction of a multi-m RNA model for predicting early recurrence of hepatocellular carcinoma after tumor resection OBJECTIVE: Early recurrence after tumor resection is the main factor leading to the poor prognosis of HCC.Identifying patients with high risk of early recurrence and conducting more intensive follow-up and appropriate post-operative adjuvant therapy will effectively improve the prognosis of these patients.The purpose of this research part is to construct a multi-m RNA model for predicting early recurrence of HCC by analyzing the large public data of HCC.METHODS: Gene sets which were commonly differentially expressed in multiple HCC datasets were screened by GEO2 R and Venn diagram analyses;early recurrence-related genes were further screened,and LASSO analysis were performed to construct a multi-gene prediction model;the predictive value of the model was evaluated in GSE14520 and TCGA;an early recurrence prediction nomogram was constructed by combining the predictive model with clinical and pathological indicators,and the clinical benefit was evaluated.RESULTS: A total of 257 genes which were commonly differentially expressed in HCC datasets were identified;a prediction model of early recurrence risk was constructed,which consisted of 24 genes and related risk factors;the model can effectively predict early recurrence of HCC after tumor resection in GSE14520 and TCGA,and increase the prognostic predictive value of TNM and BCLC staging;the multi-m RNA signature integrated nomogram have more predictive value and higher clinical benefit.CONCLUSION: The multi-m RNA model can effectively predict the early recurrence of HCC after tumor resection.It can increase the predictive value of the commonly used staging system(TNM staging and BCLC staging)for early recurrence and benefit HCC patients.Part II The function,mechanism and therapeutic value of UCK2 in hepatocellular carcinomaOBJECTIVE: Metabolic enzymes are the ultimate executors of metabolic reprogramming in cancer cells.The therapeutic value of metabolic enzymes has been verified in some tumors,but there are few related studies in HCC.The purpose of this research part is to systematically screen out the key metabolic enzyme that may play an important role in the development of HCC,and to study its function,mechanism and therapeutic value in HCC,so as to provide a new potential target for the diagnosis and treatment of HCC.METHODS: All differentially expressed genes and metabolic genes in GSE14520 were collected,and the intersection of them was all differentially expressed metabolic genes;according to the prognostic data of GSE14520,UCK2 might be a key metabolic enzyme in the development of HCC;the expression of UCK2 in HCC and adjacent non-tumor tissues were detected by q RT-PCR,Western-blot and IHC tests;TCGA data analyses and clinical samples validation were performed to explore the specific mechanism of UCK2 overexpression;the prognostic value of UCK2 was studied in two independent HCC cohorts;UCK2 knockdown and overexpression cell lines were constructed for in vitro and in vivo functional studies about growth(CCK-8 test,Ed U test,clone formation test and subcutaneous xenograft model)and migration/invasion(transwell migration/invasion test,scratch wound healing test,intrahepatic metastasis model and lung metastasis model);through the targeted site-directed mutagenesis,the effect of loss of UCK2 metabolic activity on the growth and migration/invasion was studied in HCC cells;with phosphorylated antibody chip and Co IP tests,the specific molecular mechanism of UCK2 in HCC was studied;through drug sensitivity test and combined drug inhibition test in vitro and in vivo,the therapeutic value of UCK2 was explored in HCC.RESULTS: 442 differentially expressed metabolic genes were screened out,of which 155 were up-regulated and 287 were down-regulated;among the top 10 up-regulated metabolic genes,UCK2 could most effectively distinguish the prognosis of HCC in GSE14520,indicating that UCK2 might be a key metabolic enzyme in HCC;UCK2 was significantly up-regulated in HCC clinical samples,and this up-regulation might be due to the significant amplification of its genomic level;UCK2 could effectively predict the overall survival,recurrence-free survival and early recurrence-free survival of HCC;UCK2 significantly promoted the growth and migration/invasion of HCC cells in vitro and in vivo,and its promoting effect on growth,but not migration/invasion depended on its metabolic activity;UCK2 inhibited the ubiquitination of EGFR,slowed down the degradation of EGFR and further activated Akt phosphorylation to promote the migration/invasion of HCC cells;UCK2 maked HCC cells more sensitive to cytotoxic pyrimidine nucleotide analogues and EGFR inhibitors;UCK2 promoted the production of synergistic inhibitory effect of combined treatments with cytotoxic pyrimidine nucleotide analogues and EGFR inhibitors in HCC cells.CONCLUSION: UCK2,as a key metabolic enzyme in HCC,can promote the growth and migration/invasion properties of HCC cells through metabolic and non-metabolic mechanisms respectively.Treatments targeting its metabolic and non-metabolic activities or combined treatment of these two will effectively improve the prognosis of HCC patients,which provides a new target and new method for the diagnosis and treatment of HCC. |
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